1. health and fitness
2. disease

Products and Services Catalog
IDEXX Technology
IDEXX People
Introduction
Introduction
*FlockChek, HerdChek, CHEKIT and xChek are trademarks
or registered trademarks of IDEXX Laboratories, Inc. in the
United States and/or other countries.
Microsoft and Windows are registered trademarks of
Microsoft Corporation.
All other products, company names and logos are trademarks
or registered trademarks of their respective holders.
Catalog
Products and Services Catalog
Every day around the world, thousands of laboratories and clinics rely on
IDEXX instrumentation, microtiter well assays and software for accurate
assessment of infectious diseases in poultry, swine, horses and ruminants—
including cattle, sheep, goats and deer. Since its beginning in 1984,
IDEXX has been dedicated to working with veterinarians and researchers
worldwide to bring the most advanced veterinary diagnostic products into
their laboratories and practices.
This strong commitment to veterinary medicine has led to the development
of a team of dedicated IDEXX customer and technical service staff who help
veterinarians, producers, researchers and technicians around the world.
IDEXX products have been developed by on-site research-and-development
scientists, whose experiences span such diverse technologies as DNA
probe, immunoassays, clinical chemistries, microbiology, hematology,
molecular biology and cellular biology.
IDEXX has established a strong global presence in veterinary medicine,
maintaining offices throughout the world and offering products to customers
in more than 50 countries. In 2004, IDEXX welcomed Bommeli Diagnostics
into its organization. Located in Bern, Switzerland, Bommeli produces and
distributes state-of-the-art ELISAs worldwide—their CHEKIT* ELISA product
line stems from the Bommeli IBR total antibody ELISA, the first commercially
available test. The combined resources of IDEXX and Bommeli allow us
to offer improved customer and technical service and a wider range of
diagnostic products.
Introduction
1-1
Technology
IDEXX Technology
AGID
The agar gel immunodiffusion (AGID) test involves the diffusion of antigen
and antibody through an agar gel matrix. At areas of concentration
equilibrium, precipitin lines of identity can be visualized in positive sera,
whereas no precipitin lines of identity form with negative sera.
FlockChek*, HerdChek* and CHEKIT* ELISA Kits
IDEXX offers cost-effective microwell tests to aid in the detection of livestock and poultry diseases. Samples are introduced into wells coated with
antibody or antigen, and allowed to incubate. After washing and the
subsequent addition of detector reagents, results are then read using an
ELISA plate reader. The reader can be connected to a computer with IDEXX
xChek* software installed. xChek calculates test results and provides
various data summaries and graphical reports that are helpful in herd and
flock disease management.
DNA Probe Kits
The polymerase chain reaction (PCR) amplification procedure utilizes
synthetic DNA molecules that are specific for infectious agents to amplify
small numbers of target DNA molecules. This highly sensitive and specific
technique can be used for the direct detection of infectious agents in
various clinical specimens.
HerdChek* PCFIA Systems
Introduction
Particle concentration fluorescence immunoassay (PCFIA) is based on the
use of submicron polystyrene particles as the solid phase for performing
rapid and sensitive fluorescent immunoassays. Particles coated with
antigens react with sample and fluorescent conjugate in the wells of a
patented 96-well vacuum filtration plate. Separation of bound versus free
fluorescence is achieved by vacuum filtration. The solid-phase particles are
then washed to remove unbound antibodies and the total particle-bound
fluorescence is quantitated by front-surface fluorimetry.
1-2
People
IDEXX People
Research and Development/Technical Support
The IDEXX research and development organization continues to provide an
important competitive advantage. This team of more than 150 professionals
continuously develops new products and supports more than 350 existing
products. The IDEXX Production Animal Services division has aligned key
individuals within the R&D team to assist with our customers’ evolving needs.
Production/Distribution
IDEXX Laboratories operates two locations that manufacture products to high standards: Westbrook, Maine, USA, a USDA-licensed facility that is anticipated to be ISO 9001-2000 certified in 2006, and the ISO-certified
Bern, Switzerland, plant. Products are qualified worldwide by appropriate national reference
laboratories prior to sale and distribution. IDEXX has state-of-the-art
distribution facilities in the United States and Europe, and maintains a
worldwide network of distributors to facilitate product delivery to customers.
Sales and Product Support
IDEXX has established a superb sales force throughout the world. We have
representatives in North America, Europe, South America and Asia ready to
listen to customer needs and provide innovative solutions. Customer and Technical Services
Our customer and technical service representatives are committed to
understanding and responding to the needs of customers worldwide. They
are available for your ordering needs, technical questions, and equipment
and software installations.
Introduction
1-3
Introduction
Avian Encephalomyelitis (AE)
Avian Influenza (AI)
Avian Leukosis Virus (LLAg, LLAb, ALV-J)
Avian Pneumovirus (APV)
Chicken Anemia Virus (CAV)
Infectious Bronchitis Virus (IBV)
Infectious Bursal Disease—Gumboro Disease (IBD, IBD-XR)
Mycoplasma (MG, MS, MG/MS, MM)
Newcastle Disease Virus (NDV, NDV-T)
Ornithobacterium rhinotracheale (ORT)
Pasteurella multocida (PM, PM-T)
Reovirus (REO)
Reticuloendotheliosis Virus (REV)
Salmonella enteritidis (SE)
Poultry Test Kits
Poultry Test Kits
AE
Avian Encephalomyelitis Antibody
Test Kit
FlockChek* AE Antibody ELISA: 5 plates, serum samples, indirect format
Avian encephalomyelitis (AE) is a viral infection primarily
affecting young birds. The disease is characterized by
a variety of neurological signs, including incoordination,
ataxia and tremors of the head and neck. An assessment
of immune status, as well as serologic identification of
AE, requires a measurement of antibody to AE in serum.
Enzyme-linked immunosorbent assays (ELISAs) have
proven efficacious in the quantification of antibody levels to AE, and facilitate the monitoring of immune status in large flocks.
Poultry Test Kits
2-1
AI
Avian Influenza Antibody Test Kit
FlockChek* AI Antibody ELISA: 5 plates, serum samples, indirect format
Poultry Test Kits
Strains detected include:
H7N2
H1N7
H7N3
H5N9
H11N6 H3N8
H5N2
H4N8
H10N7
H8N4
H14N5 H6N5
H5N1
2-2
H13N6
H9N2
H2N9
H12N5
Domestic and wild avian species are affected by avian
influenza viruses. The disease is characterized by a wide
range of responses, from virtually no clinical signs to high
mortality. Respiratory signs are common, along with a drop
in egg production, greenish diarrhea, bloodstained nasal
and oral discharges, and cyanosis and edema of the head,
comb and wattle.
Due to the variation and severity of clinical symptoms,
serological testing produces significant advantages to
detection of infected flocks. Monitoring the exposure of a
flock to influenza viruses is facilitated by the measurement
of antibody to avian influenza virus in serum.
ALV
Lymphoid leukosis, the most common manifestation of the avian leukosis/
sarcoma group of viruses, produces a variety of neoplastic diseases, including
erythroblastosis, myelocytomatosis, myeloblastosis and others. Not all infected
birds will develop tumors. Infection can occur horizontally from bird to bird by
direct or indirect contact, or vertically from an infected hen to her eggs as virus
is shed into the albumin of the egg. In addition, vertical transmission may occur
from virus incorporated in the DNA of a germ cell. Viremia in the hen is strongly
associated with the transmission of virus congenitally. Enzyme immunoassays
have proven efficacious in the detection of both leukosis antibody and antigen.
Avian Leukosis Virus
Antibody Test Kit
FlockChek* LL Antibody ELISA: 5 plates, serum samples, indirect format
This enzyme-linked immunosorbent assay (ELISA) is
designed to detect antibody to avian leukosis virus
(ALV) subgroups A and B in chicken serum. Antibody to
subgroups C–E and J are not detected.
Avian Leukosis-J Virus
Antibody Test Kit
FlockChek* ALV-J Antibody ELISA: 5 plates, serum samples, indirect format
ALV-J is an avian retrovirus first isolated in meat-type
chickens in the late 1980s, and is designated as a unique
subgroup partly based on the envelope glycoprotein (gp85).1
Clinically, ALV-J causes predominantly myeloid leukosis,
with variable tumor frequency across chicken lines.1,2 As
with other avian leukosis viruses, ALV-J is transmitted both
vertically (congenital infection of the egg albumin and the chick embryo) and horizontally (through close contact
with infected chicks).2,3 This test was designed as a
screening tool using serum samples from flocks 10 weeks of age or older.
Avian Leukosis Virus
Antigen Test Kit
FlockChek* LL Antigen (p27) ELISA: 5 plates; albumin, cloacal swab and
serum samples; antigen-capture format
The ALV antigen test detects p27, an antigen common
to all subgroups of ALV, including endogenous viruses.
The recommended sample types are light albumin and
cloacal swabs. While serum has been validated for use
on the ALV antigen test, it is not a recommended sample
for the detection of exogenous virus because of potential
interference from endogenous sequences.4
2 Payne LN, Fadly AM. Neoplastic diseases/Leukosis/Sarcoma group. In: Calnek BW, et al, eds. Diseases of Poultry. 10th ed. Ames, Ia: Iowa State
University Press; 1997:414–466.
3 Payne LN. HPRS-103: A retrovirus strikes back. The emergence of subgroup J avian leukosis virus. Avian Pathology. 1998;27:36–45.
4 Payne LN, Gillespie AM, Howes K. Unsuitability of chicken sera for detection of exogenous ALV by the group-specific antigen ELISA. Veterinary
Record. May 1993:555–557.
2-3
Poultry Test Kits
1 Payne LN, et al. A novel subgroup of exogenous avian leukosis virus in chickens. Journal of General Virolog. 1991;72:801–807.
APV
Avian Pneumovirus
Antibody Test Kit
Poultry Test Kits
FlockChek* APV Antibody ELISA‡:
Detects A, B and/or C avian pneumovirus
serotypes, 5 plates (solid), chicken and
turkey serum, indirect format
Avian pneumovirus (APV), previously known as turkey
rhinotracheitis virus, causes acute, highly contagious
upper respiratory tract diseases in turkeys and chickens.
Typical signs of the disease include sneezing, nasal
discharge, conjunctivitis, swollen sinuses and a marked
drop in egg production. The virus can also infect chickens
without exhibiting any clinical signs. The FlockChek* Avian
Pneumovirus Antibody Test Kit is designed to detect APVspecific antibodies in serum samples.
‡
Available for sale outside the U.S.
2-4
CAV
Chicken Anemia Virus
Antibody Test Kit
FlockChek* CAV Antibody ELISA: 5 plates, serum samples, blocking format,
1:10 dilution for detection, titer calculation
available for 1:100 dilution for monitoring
vaccine response
Chicken anemia virus (CAV) is an important pathogen
of poultry and has been found in broilers, breeders and
specific-pathogen-free flocks. Virus isolation is difficult and
time-consuming. Screening for the presence of antibody
will indicate exposure to the virus. The enzyme-linked
immunosorbent assay (ELISA) has been utilized to detect
antibody against CAV. This method is quite useful in largescale testing of flocks for exposure to CAV.
Poultry Test Kits
2-5
IBV
Infectious Bronchitis Virus
Antibody Test Kit
Poultry Test Kits
FlockChek* IBV Antibody ELISA: 5 plates, serum samples, indirect format
2-6
Infectious bronchitis virus (IBV) is a highly contagious
viral disease of chickens that is usually manifested as a
respiratory condition. An assessment of immune status,
as well as serological identification of IBV, requires a
measurement of antibody to IBV in serum. Enzyme-linked
immunosorbent assay (ELISA) systems have proven
efficacious in the quantification of antibody levels to IBV, and
facilitate the monitoring of immune status in large flocks.
IBD
Infectious bursal disease (IBD), or Gumboro disease, is a viral disease
affecting chickens in a subclinical form (early bursa atrophy) that may lead to
a temporary or permanent immunosuppression. The clinical form in chickens
may appear at 3–6 weeks of age. The bursa becomes swollen and then
quickly regresses to a small size, leading to suppression of the immune
system. Symptoms include anorexia, incoordination and depression. Affected
birds are more susceptible to a variety of infectious agents, including
Staphylococcus, Clostridium, E. coli and the respiratory viruses. Coinfection
with CAV can enhance the immunosuppression condition. Losses may
approach 20% in an infected flock, and subsequent flocks may become
infected from a contaminated living environment. An assessment of immune
status, as well as serologic identification of IBD, requires a measurement of
antibody to IBD in serum.
Infectious Bursal Disease Antibody
Test Kits
FlockChek* IBD Antibody ELISA: 5 plates, serum samples, indirect format
Enzyme-linked immunosorbent assay (ELISA) systems are
efficacious in the quantification of antibody levels to IBD,
facilitate the monitoring of immune status in flocks, and aid
in determining the appropriate time for vaccination.
FlockChek* IBD-XR Antibody ELISA
(Extended Range): 5 plates, serum
samples, indirect format, extended titer range, enhanced detection of variant strains
This ELISA’s extended titer range allows for the differentiation of IBD viruses and enhanced detection of IBD variant strains.
Poultry Test Kits
2-7
Mycoplasma
Poultry flocks are susceptible to respiratory infections from a variety of agents,
including Mycoplasma spp. The usual types of infection from Mycoplasma spp
are chronic respiratory disease, airsacculitis, sinusitis and synovitis. In many
cases, however, the infection may be identified only through serological and culture methods. Monitoring a flock for exposure to Mycoplasma is facilitated
by the measurement of antibody to Mycoplasma in serum.
Mycoplasma gallisepticum Test Kits
FlockChek* MG Antibody ELISA:
5 plates, serum samples, indirect format,
chicken and turkey samples
FlockChek* MG/MS Antibody ELISA:
5 plates, serum samples, indirect format,
plates coated with both MG and MS
antigens, chicken and turkey samples
FlockChek* MG DNA Probe: 30 tests
per kit, tracheal swab, PCR** DNAamplification procedure, chicken and
turkey samples
Mycoplasma gallisepticum (MG) is associated with chronic
respiratory disease in chickens and infectious sinusitis
in turkeys. The symptoms generally seen are coryza,
coughing, nasal exudate and respiratory rales. Economic
losses due to M. gallisepticum infection include reduced
egg production, poor eggshell quality, lowered hatchability
of chicks and downgraded meat quality.
The FlockChek* Mycoplasma gallisepticum DNA Probe
Test Kit is a nonradioactive probe-based test utilizing PCR
amplification for the specific detection of Mycoplasma
gallisepticum genomic DNA from chicken and turkey
tracheal swab samples.
Mycoplasma meleagridis Antibody
Test Kit
FlockChek* MM Antibody ELISA: 5 plates, serum samples, indirect format,
turkey samples
Mycoplasma meleagridis (MM) is the cause of egg-transmitted disease of turkeys in which the primary lesion is an
airsacculitis in the progeny. Other manifestations include
decreased hatchability, skeletal abnormalities and poor
growth performance. The organism is a specific pathogen of
turkeys. MM is distributed worldwide, and the incidence of
MM-associated airsacculitis is very high (20–65%) in naturally
infected cull poults.
Mycoplasma synoviae Test Kits
FlockChek* MS Antibody ELISA:
5 plates, serum samples, indirect format,
chicken and turkey samples
Poultry Test Kits
FlockChek* MG/MS Antibody ELISA:
5 plates, serum samples, indirect format,
plates coated with both MG and MS
antigens, chicken and turkey samples
FlockChek* MS DNA Probe: 30 tests
per kit, tracheal swab, PCR** DNAamplification procedure, chicken and
turkey samples
2-8
Mycoplasma synoviae (MS) is a known pathogen associated
with the development of synovitis and chronic respiratory
disease in chickens and turkeys. Clinical symptoms include
joint swelling, coryza and respiratory rales. Economic
losses due to M. synoviae infection include reduced egg
production, lowered hatchability of chicks and downgraded
meat quality. The FlockChek* Mycoplasma synoviae
DNA Probe Test Kit is a nonradioactive probe-based test
utilizing PCR** amplification for the specific detection of
Mycoplasma synoviae genomic DNA from chicken and
turkey tracheal swab samples.
NDV
Newcastle Disease Virus
Antibody Test Kits
FlockChek* NDV Antibody ELISA
for chickens: 5 plates, serum samples,
indirect format
FlockChek* NDV Antibody ELISA
for turkeys: 5 plates, serum samples,
indirect format
Newcastle disease is a highly contagious and sometimes
fatal illness affecting poultry. The severity of the disease
is a function of the virulence of the infecting viral strain.
Lentogenic strains infect the trachea, lungs and air
sacs, and interfere with egg production. Mesogenic and
velogenic strains are manifested through incoordination,
paralysis, swelling of tissue around the eyes, diarrhea
and eventual death. An assessment of immune status,
as well as serological identification of Newcastle disease
virus (NDV), requires a measurement of antibody to NDV
in serum. Enzyme-linked immunosorbent assay (ELISA)
systems have proven efficacious in the quantification of
antibody levels to NDV, and facilitate the monitoring of
immune status in large flocks.
Poultry Test Kits
2-9
ORT
Ornithobacterium rhinotracheale
Antibody Test Kit
Poultry Test Kits
FlockChek* ORT Antibody ELISA‡: 5 plates, serum samples, indirect format,
chicken and turkey samples
Ornithobacterium rhinotracheale has been associated with
respiratory disease, increased mortality, retarded growth
and decreased egg production in avian species worldwide.
Viral or bacterial infections by IBV, APV and NDV are able
to trigger an ORT infection. An assessment of immune
status, as well as serologic identification of ORT, requires
a measurement of antibody to ORT in serum. Enzymelinked immunosorbent assay (ELISA) systems have proven
efficacious in the quantification of antibody levels to ORT,
and facilitate the monitoring of immune status in large
flocks. The FlockChek* ORT Test Kit detects serological
response to ORT serotypes A–M.
‡
Available for sale outside the U.S..
2-10
PM
Pasteurella multocida
Antibody Test Kits
FlockChek* PM Antibody ELISA
for chickens: 5 plates, serum samples,
indirect format
FlockChek* PM Antibody ELISA
for turkeys: 5 plates, serum samples,
indirect format
Fowl cholera, caused by Pasteurella multocida (PM)
infection, is a commonly occurring disease of birds. In
the acute form, its usual symptom is a septicemia with
associated high morbidity and mortality. Chronic localized
infections can also occur, either following an acute exposure
or resulting in infection with an organism of low virulence.
Clinical signs of acute infections are typical of bacterial
septicemia, whereas the signs of chronic disease are
typically related to the anatomic location of the infection.
An assessment of immune status, as well as serologic
identification of PM, requires a measurement of antibody in
serum. Enzyme-linked immunosorbent assay (ELISA) have
proven efficacious in the quantification of antibody levels
to other diseases, and facilitate the monitoring of immune
status in large flocks.
Poultry Test Kits
2-11
REO
Avian Reovirus
Antibody Test Kit
Poultry Test Kits
FlockChek* REO Antibody ELISA: 5 plates, serum samples, indirect format
2-12
Avian reoviruses are ubiquitous among poultry populations
and have been reported to be responsible for viral
arthritis (tenosynovitis), runting/stunting, malabsorbtion
syndrome and feed passage in birds 4–16 weeks of
age. The incidence of reovirus infection in older birds is
high, but clinical symptoms are not seen in most birds.
An assessment of immune status, as well as serologic
identification of avian reovirus, requires a measurement
of antibody to reovirus in serum. Enzyme-linked
immunosorbent assays (ELISAs) have proven efficacious in
the quantification of antibody levels to other diseases, and
facilitate the monitoring of immune status in large flocks.
REV
Reticuloendotheliosis Virus
Antibody Test Kit
FlockChek* REV Antibody ELISA: 5 plates, serum samples, indirect format
Reticuloendotheliosis virus (REV) is a retrovirus that is
morphologically similar to, but genetically distinct from, the
avian leukosis/sarcoma viruses. Runting disease has been
reported in chickens through the application of vaccines
contaminated with REV. In addition, REV can produce disease
in experimentally infected chickens that is pathologically
indistinguishable from lymphoid leukosis. An assessment of
infection with REV requires a measurement of antibody to REV
in serum. Enzyme-linked immunosorbent assays (ELISAs) are
especially suitable for screening chicken flocks.
Poultry Test Kits
2-13
SE
Salmonella enteritidis
Antibody Test Kit
FlockChek* SE Antibody ELISA: 5 plates, serum and egg yolk samples,
blocking format
Salmonellosis is one of the most important zoonotic
diseases. The major factor in the success of Salmonella
spp as virtually universal pathogens is their ability to adapt
to almost any kind of host. Salmonella-contaminated food
products are a major source of human infection. Control
programs based on serological monitoring and isolation of
Salmonella aim not only toward the reduction of infection
prevalence, but also could serve as valuable tools to
change routines to decrease food contamination.
Salmonella enteritidis (SE) is an important pathogen of
poultry and has been isolated from broilers, breeders and
commercial egg-laying flocks. Bacteriological identification
of positive birds is difficult due to the intermittent excretion
of the organism. The presence of antibody does not always
signify infection, but is indicative of previous exposure.
Poultry Test Kits
The enzyme-linked immunosorbent assay (ELISA) has
shown utility in the detection of antibody to Salmonella in
poultry, and is particularly useful in large-scale monitoring
of flocks for Salmonella enteritidis infection. The FlockChek*
SE Test Kit can be used as an initial screening method
for the presence of Salmonella enteritidis antibodies.
Because the SE ELISA is a g,m flagellin-based assay,
other Salmonella serotypes that share the epitopes of g,m
flagellin can potentially yield positive results. Therefore,
positive ELISA screening results must always be confirmed
by standard bacteriological methods.
2-14
Actinobacillus pleuropneumoniae (App)
Classical Swine Fever Virus (CSFV Ab, CSFV Ag)
Foot-and-Mouth Disease (FMD)
Mycoplasma hyopneumoniae (M. hyo.)
Porcine Reproductive and Respiratory Syndrome (PRRS 2XR)
Pseudorabies Virus—Aujeszky’s disease (PRV gB, PRV-V, PRV-S, PRV g1 (gE))
Swine Influenza (SIV H1N1, SIV H3N2)
Swine Salmonella
Swine Test Kits
Swine Test Kits
App
APP-ApxIV Antibody Test Kit
CHEKIT* APP Antibody ELISA‡:
2 plates or 10 plates (strips), unique
serological screening method covering
all serotypes, no cross-reaction with
other bacteria, differentiates infected from
vaccinated animals (DIVA), results in less
than three hours
Swine pleuropneumonia caused by Actinobacillus
pleuropneumoniae (App) is a highly contagious pulmonary
disease that may cause important economic losses. Clinical
signs of the acute disease are dyspnea, coughing, anorexia,
depression, fever and sometimes vomiting. In the absence
of treatment, the disease can progress very rapidly and
death can occur within a few hours. Chronic infections are
characterized by cough and pleuritis in the lungs.
It is important to differentiate infection from disease. Indeed,
many herds are infected with App without presenting any
clinical evidence of the disease. Carrier pigs harbor App in
their nasal cavities and/or tonsils. These animals represent
the major source of dissemination of the infection between
herds. Mixing animals infected with virulent App strains
together with immunologically naive animals, inappropriate
management, concurrent infections (e.g., PRRS,
pseudorabies) and/or stress conditions may be responsible
for the sudden appearance of severe clinical disease.
Fifteen serotypes have been described, which variously
express four different proteinaceous cytotoxins (ApxI,
ApxII, ApxIII, ApxIV) belonging to the RTX toxin family.
There are many lines of evidence that suggest these toxins
play a predominant role in the pathogenicity of App with
different activities (hemolytic, cytotoxic, etc.). Due to their
strong antigenic properties, they have been used in the
serodiagnosis of App infections. However, other bacteria
express toxins that serologically cross-react with ApxI, ApxII
or ApxIII.
In contrast, ApxIV, a fourth RTX determinant, has been found
to be common and specific to all serotypes.
Swine Test Kits
‡
Available for sale outside the U.S.
3-1
CSFV
Classical Swine Fever Virus
Test Kits
HerdChek* CSFV Antibody ELISA‡: 5 plates (solid or strips), serum or plasma
samples, overnight protocol with optional
two-hour result
CHEKIT* CSF SERO Antibody ELISA:
10 plates (strips), serum or plasma
samples, overnight protocol, blocking
(competitive) format
CHEKIT* HCV Antigen ELISA: 2 and 10 plates (strips); serum, plasma or
leukocyte samples; short and overnight
protocols
CHEKIT* CSF Marker ELISA: 10 plates (strips), serum or plasma
samples, differentiates infected from
vaccinated animals (DIVA)
Classical swine fever virus (CSFV), or hog cholera, causes
serious losses in the pig industry because it is highly
pathogenic and may cause widespread deaths. Pigs
infected with highly virulent CSFV strains may shed high
amounts of virus before showing clinical signs of the
disease. Animals that survive an acute or subacute infection
develop antibodies and will no longer spread the virus.
Moderately virulent, less pathogenic strains may lead to
chronic infection when pigs excrete the virus continuously or
intermittently until death. Congenital infection may result in
abortion, mummified fetuses, stillborn and/or weak piglets,
or embryonic malformations.
The most frequent outcome of congenital infection with low
virulent strains is the birth of persistently infected piglets
spreading the virus without the signs of the disease in the
absence of immune response. All animals shedding the
virus need to be identified and removed from the herd to
prevent the spread of infection. Pigs may also be infected
with bovine viral diarrhea virus (BVDV) or border disease
virus (BDV). Although these infections in pigs are usually
mild and self-limiting, it is important to reliably distinguish
them from the CSFV infection.
Swine Test Kits
The field experience and EU regulation actually in place are
also asking for a DIVA system that permits the differentiation
between SCF-negative animals and animals vaccinated with
an E2 subunit vaccine (Erns-negative) from CSF-infected
animals (Erns-positive).
‡
Available for sale outside the U.S.
3-2
FMD
Foot-and-Mouth Disease
3ABC Antibody Test Kit
CHEKIT* FMD Antibody ELISA‡: 2 and
10 plates (strips), specific to screening
serum or plasma samples from swine,
differentiates infected from vaccinated
animals (DIVA)
Foot-and-mouth disease (FMD) is a highly contagious
viral disease. It can spread rapidly and affects both
domesticated and wild ruminants, as well as pigs. FMD has an economically devastating impact on affected
countries because trade barriers are imposed in countries
where the disease occurs.
The FMD genome encodes a single polyprotein that is
cleaved during the translation. Mature structural and
nonstructural viral protein derive after a cascade of further
cleavages by viral proteases.
Vaccine produced according to the guidelines of the OIE
are depleted of nonstructural proteins like 3ABC during
purification steps. Antibody to 3ABC is generally accepted
as the most reliable single indicator for viral replication.
Detection of anti-3ABC antibodies has been described upon
challenge in animals with and without previous vaccination,
confirming the potential of 3ABC as a marker for the
surveillance of vaccinated populations.
Swine Test Kits
‡
Available for sale outside the U.S.
3-3
M. hyo.
Mycoplasma hyopneumoniae
Antibody Test Kit
Swine Test Kits
HerdChek* M. hyo. Antibody ELISA: 5 plates, serum and plasma samples,
indirect format, results in less than two hours
3-4
Enzootic pneumonia, or mycoplasma pneumonia of swine,
a chronic disease with a high morbidity and a low mortality,
is caused by Mycoplasma hyopneumoniae. The clinical
signs include a chronic nonproductive cough, retarded
growth, slow onset, and spread and repeated occurrence of
the disease. The IDEXX M. hyo. antibody ELISA allows rapid
screening for the presence of antibodies to Mycoplasma
hyopneumoniae, which can be an indicator of exposure
to the agent. Monitoring the immune status of a herd with
regard to M. hyo. can play an important role in the control of
this disease.
PRRS
Porcine Reproductive and Respiratory
Syndrome Antibody Test Kit
HerdChek* PRRS 2XR Antibody
ELISA: 5 plates (strips), serum samples,
utilizes recombinant PRRS and normal
host cell (NHC) antigens, results in less
than two hours
Porcine reproductive and respiratory syndrome (PRRS)
is a swine disease that causes reproductive problems,
respiratory disease and mild neurological signs. Due to the
general clinical symptoms presented in most cases, diagnosis
is often confused with swine influenza, pseudorabies
(Aujeszky’s disease), classical swine fever virus, parvovirus,
encephalomyocarditis, Chlamydia and Mycoplasma. A
major component of the syndrome is reproductive failure,
resulting in premature births, late-term abortions, piglets born
weak, increased stillbirths, mummified fetuses, decreased
farrowing rates and delayed return to estrus. These aspects
of the syndrome have been observed to last 1–3 months.
Respiratory disease is another significant feature of the
syndrome that will most likely affect pigs less than 3–4 weeks
of age. Respiratory signs can occur in most stages of the
production cycle.
The PRRS 2XR ELISA can be used to develop herd profiles
that provide important decision-making information to better
manage PRRS-exposed herds. The IDEXX xChek* software
provides the ability to assess the significance of changes in
herd profiles over time. You can perform additional statistical
and/or graphical analyses and export the data to Microsoft*
Excel files.
Swine Test Kits
3-5
PRV
Pseudorabies virus (PRV/Aujeszky’s disease) is caused by a type 1 porcine
herpesvirus. Infections with the highest mortality rate are those affecting suckling
pigs born to a susceptible sow. Baby pigs in the fatal course of the disease exhibit
difficulty breathing, fever, hypersalivation, anorexia, vomiting, diarrhea, trembling and depression.
Within this age group, the final stages of infection are commonly characterized by
ataxia, nystagmus, running fits, intermittent convulsions, coma and death. Death
usually occurs 24–48 hours following apparent clinical symptoms. The clinical
events of the disease in weaning and fattening pigs are essentially the same
except the course of the disease is usually protracted 4–8 days.
The mortality rate in mature pigs may reach 2%, however, losses do not usually
occur. While the clinical course in pregnant swine is virtually the same as in mature
pigs, there is variation due to transplacental infection of fetuses. Transplacental
infection with PRV can occur, and depending upon the stage of gestation, one
of the following sequelae may occur: resorption, premature expulsion, birth of
macerated fetuses, stillbirth, or birth of weak, infected pigs.
An assessment of the exposure to pseudorabies virus via natural infection or
vaccine is facilitated by a measurement of antibody in the serum.
Pseudorabies Virus
Antibody Test Kits
The HerdChek* PRV gB Test Kit is a blocking immunoassay
for the detection of antibodies in swine serum or plasma to
the gB antigen of the pseudorabies virus (PRV/Aujeszky’s
disease). The presence of antibodies indicates exposure to
the pseudorabies virus.
HerdChek* PRV g1 (gE) Antibody
ELISA: 6 (strips) and 30 plate (solid),
serum samples, differentiates infected
from vaccinated animals (DIVA), results in less than two hours
The HerdChek* Anti-PRV g1 (gE) Test Kit is an immunoassay
for the detection of antibodies in swine serum to the g1 (gE)
antigen of pseudorabies virus (PRV/Aujeszky’s disease).
The presence of antibodies indicates exposure to field
strains of PRV and/or vaccines containing g1 antigen. This
differential test is intended for use in herd management and
pseudorabies eradication applications.
Swine Test Kits
HerdChek* PRV gB Antibody ELISA:
6 plates (solid), serum or plasma
samples, blocking format, results in less
than two hours
3-6
SIV
Swine influenza is an acute infectious respiratory disease of swine caused
by type A influenza viruses. The disease is characterized by a sudden onset,
coughing, dyspnea, fever and prostration, followed by rapid recovery. Swine Influenza Virus (H1N1)
Antibody Test Kit
HerdChek* SIV (H1N1) Antibody
ELISA: 5 plates, serum samples, indirect
format, results in less than two hours
The IDEXX SIV (H1N1) antibody ELISA allows rapid
screening for the presence of antibodies to subtype H1N1
swine influenza, which can be an indicator of exposure to
the virus. Monitoring the immune status of a herd, maternal
antibody decay, vaccine immune response, and field virus
interaction with vaccination program with regard to SIV
(H1N1) can play an important role in the control of swine
influenza virus.
Swine Influenza Virus (H3N2)
Antibody Test Kit
HerdChek* SIV (H3N2) Antibody
ELISA: 5 plates, serum samples, indirect
format, results in less than two hours
The IDEXX SIV (H3N2) antibody ELISA allows rapid
screening for the presence of antibodies to subtype H3N2
swine influenza, which can be an indicator of exposure to
the virus. Monitoring the immune status of a herd, maternal
antibody decay, vaccine immune response, and field virus
interaction with vaccination program with regard to SIV
(H3N2) can play an important role in the control of swine
influenza virus.
Swine Test Kits
3-7
Salmonella
Swine Salmonella
Antibody Test Kit
HerdChek* Swine Salmonella
Antibody ELISA‡: Detects antibodies
against the most common serotypes (B, C1, D) isolated in Europe, Asia and
America; 5 plates (strips), 30 plates
(solid); serum, plasma or meat juice
samples; rapid testing method—short
(<two hours) and overnight protocols;
results available as S/P and OD% values
using IDEXX xChek* software
Salmonellosis is one of the most important zoonotic
diseases. The major factor in the success of Salmonella
spp as virtually universal pathogens is their ability to adapt
to almost any kind of host. Salmonella-contaminated food
products are a major source of human infection. Control
programs based on serological monitoring and isolation of
Salmonella aim not only toward the reduction of infection
prevalence, but also could serve as valuable tools to
change routines to decrease food contamination.
The great majority of clinical disease in swine is caused by
S. cholerasuis in the U.S. and by S. typhimurium in Europe.
S. typhimurium is mostly a latent disease in pigs, and
causes diseases only in conjunction with other cofactors.
The primary host and main reservoir of S. cholerasuis is
swine. While infections with S. cholerasuis can cause severe
clinical disease in swine, it is of minor importance in food
safety. Human infections with S. cholerasuis are rare due to
a limited number of infected swine at the abattoir.
Swine Test Kits
The HerdChek* Swine Salmonella Antibody Test Kit allows
rapid screening for the presence of antibodies to a broad
range of Salmonella serogroups, indicating exposure of the
swine herd to the bacteria. Since serological screening is easy to run in large scale, it is useful for estimating the herd prevalence.
‡
Available for sale outside the U.S.
3-8
Bovine Leukemia Virus/Enzootic Bovine Leukosis (BLV/EBL)
Bovine Viral Diarrhea Virus (BVDV)
Brucella abortus (B. abortus)
(Bovine, Ovine)
Brucella ovis (B. ovis)
Caprine Arthritis Encephalitis Virus/Maedi-Visna Virus (CAEV/MVV)
(Caprine, Ovine)
Chlamydia
(Bovine, Caprine, Ovine)
Foot-and-Mouth Disease (FMD)
(Bovine, Ovine)
Infectious Bovine Rhinotracheitis (IBR)/ Bovine Herpesvirus-1 (BHV-1)
Mycobacterium paratuberculosis (M. pt.)—Johne’s disease (Bovine, Ovine)
Neospora caninum
(Bovine, Caprine, Ovine)
Q Fever (Bovine, Caprine, Ovine)
Toxotest (Toxoplasma gondii)
(Caprine, Ovine)
Transmissible Spongiform Encephalopathies (TSEs)
Bovine Spongiform Encephalopathy (BSE)
Bovine Spongiform Encephalopathy-Scrapie (BSE-Scrapie)
(Bovine, Caprine, Ovine)
Chronic Wasting Disease (CWD)
(Cervid)
Ruminants
RuminantsTest
Test
Kits
Kits
Ruminants Test Kits
BLV/EBL
Bovine Leukemia Virus/Enzootic
Bovine Leukosis Antibody Test Kits
HerdChek* BLV Screen/Verification
Antibody elisa: 5 screen/1 verification
plate (strips), serum or plasma samples,
results in less than three hours with
optional overnight protocol, indirect format
HerdChek* BLV Verification elisa:
6 verification plates (strips), serum
samples
CHEKIT* Leukotest Screening Milk
Antibody ELISA‡: 10 plates (solid), milk
samples (individual or pool up to 250),
short or overnight protocol, monophasic
kit, results in less than three hours with
optional overnight protocol, indirect format
Bovine leukemia virus (BLV) is a retrovirus that may cause
lymphosarcoma in cattle. The virus resides in blood lymphocytes
where circulating antibodies are unable to neutralize it. Therefore,
once an animal is infected with BLV, it is infected for life.
BLV is economically significant to the producer because of
premature culling or death as a result of lymphosarcoma.
Another concern is the condemnation of carcasses at slaughter,
which has a significant economic impact on the dairy and cattle
industries. Losses from export restrictions are another economic
concern of BLV infection. Countries that have bovine leukosis
control programs require BLV-free certification prior to shipping
cattle to their regions. Moreover, exporters of semen are under
increasing pressure to ensure that their product is from a BLVfree animal in a BLV-free herd.
CHEKIT* Leukotest Verification Milk
Antibody ELISA‡: 10 plates (solid), milk
samples (individual or pool up to 250),
biphasic kit
CHEKIT* Leucose Serum Antibody
ELISA‡: 10 plates (strips), serum or
plasma samples, monophasic kit
Ruminants Test Kits
‡
Available for sale outside the U.S.
4-1
BVDV
Bovine Viral Diarrhea Virus
Test Kits
HerdChek* BVDV Antibody ELISA‡:
5 plates (strips); serum, plasma or milk
samples; overnight protocol with optional
two-hour assay; indirect format
HerdChek* BVDV Antigen ELISA—
Leukocyte‡: 5 plates (strips); leukocytes,
tissues, nasal swabs; antigen-capture
format; overnight protocol with optional
three-hour assay
HerdChek* BVDV Antigen ELISA—
Serum Plus‡: 2 plates (strips) or 5 plates
(strips and solid); serum, plasma, whole
blood and ear-notch tissue samples;
antigen-capture format; overnight protocol
with optional two-hour assay
Ruminants Test Kits
HerdChek* BVDV Antigen ELISA—
Ear-Notch and Serum†: 5 plates (strips),
serum and ear-notch tissue samples,
antigen-capture format, results in less
than four hours
Bovine viral diarrhea virus (BVDV), border disease virus
(BDV) and hog cholera virus (classical swine fever virus,
CSFV) are the three member viruses of the genus Pestivirus
within the family Flaviviridae. BVDV is one of the most
important pathogenic viruses in cattle, causing considerable
losses in the dairy and beef industries. The virus crosses the placenta in infected pregnant cows,
causing reproductive losses due to abortions, stillborn
calves or calves that die early in life. Some calves that
survive are immunotolerant to the virus and may excrete
large amounts of virus, infecting other animals in the herd.
The carrier animals often die of mucosal disease in the first
two years of life.
Persistently infected virus shedders can be detected using
the BVDV antigen ELISA. Testing for antibodies to BVDV is a
useful tool for herd-screening for BVDV prevalence and for
monitoring BVDV-free herd status.
‡
Available for sale outside the U.S.
†
Available for sale in U.S. and Canada only
4-2
B. abortus
Brucella abortus
Antibody Test Kits
HerdChek* B. abortus Antibody PCFIA:
10,560 tests per kit, serum samples,
recognized as an official test in the U.S.
(bovine)
Brucellosis in cattle is a disease caused by Brucella abortus (B. abortus), a facultative, intracellular bacterium. The major
mode of disease transmission is ingestion of B. abortus
organisms that may be present in tissues of aborted fetuses,
fetal membranes and uterine fluids. In addition, infection
may occur as the result of cattle ingesting B. abortuscontaminated feed or water. Infection in cows has also
occurred through venereal transmission of the organism by
infected bulls.
CHEKIT* Brucellose Serum Antibody
ELISA‡: 10 plates (strips), serum and
plasma samples (individual or pool up to
10), short and overnight protocols (bovine, ovine)
Abortion is the outstanding clinical feature of the disease.
If the carrier state develops in a majority of infected cows
in a herd, the clinical manifestations may be reduced milk
production, dead calves at term and a higher frequency of
retained placentas. Disease in the bull may produce infections
of the seminal vesicles and testicles, resulting in shedding of
the organisms in semen.
▲
CHEKIT* Brucellose Milk Antibody
ELISA‡: 10 plates (solid), milk samples
(individual or pool up to 250), short and
overnight protocols (bovine)
Diagnosis is based on serological (serum/milk) and
bacteriological procedures. While a positive bacteriological
finding is the most definitive diagnosis, several weeks may
be required to obtain final culture results. The success of
disease eradication is dependent upon rapid and accurate
identification.
The CHEKIT B. abortus Test Kit includes chemical reagents
(wash, substrate, stop solutions) that can also be used with
CHEKIT kits that test for other abortive diseases (Q fever,
neosporosis, toxoplasmosis, Chlamydia).
Ruminants
RuminantsTest
Test
Kits
Kits
Available for sale outside the U.S.
‡
4-3
B. ovis
Brucella ovis Antibody Test Kit
▲
CHEKIT* Brucella ovis Antibody
ELISA: 2 plates (strips), serum and
plasma samples of sheep, short protocol
(60 minutes at 37°C), ready-to-use
reagents
Ovine brucellosis is caused by a bacterium called Brucella
ovis (B. ovis). All breeds of sheep are susceptible to
brucellosis, which may cause considerable economic loss
due to increased culling of rams, reduced lamb marking
percentages and extended lambing seasons. The disease
produces inflammation of the epididymis in rams and of
the placenta in pregnant ewes. Ovine brucellosis has been
recorded in sheep from Australia, New Zealand, the United
States, South Africa and Europe.
The infection of pregnant ewes leads to abortion and
neonatal lamb deaths due to microscopic and macroscopic
changes occurring in the placenta and fetus. Infection does
not persist in nonpregnant ewes, but may persist for several
months in pregnant ewes. This characteristic has an impact
in the control of the disease.
In many countries, the routine use of vaccines against
brucellosis is no longer allowed. Brucellosis eradication
programs are now based exclusively on the serological
screening of sheep and cattle herds to detect and remove
infected animals.
The CHEKIT* Brucella ovis ELISA provides a rapid, simple,
sensitive and specific method for detecting antibodies
against Brucella ovis in serum and plasma of sheep.
Ruminants Test Kits
The CHEKIT Brucella ovis Test Kit includes chemical
reagents (wash, substrate, stop solutions) that can also be
used with CHEKIT kits that test for other abortive diseases
(B. abortus, neosporosis, toxoplasmosis, Chlamydia).
4-4
CAEV/MVV
Caprine Arthritis Encephalitis Virus/
Maedi-Visna Virus Antibody ELISA Test Kits
CHEKIT* CAEV/MVV Antibody
ELISA‡: 2 plates (strips), serum and
milk confirmation, ready-to-use reagents
(caprine, ovine)
CHEKIT* CAEV/MVV Antibody ELISA‡:
2- and 10-plate (strips) screening of
serum, plasma and milk samples of goats
and sheep; monophasic kit; short protocol
(caprine, ovine)
Maedi-visna virus (MVV) infection of sheep is characterized
by slowly progressive arthritis, pneumonia, mastitis and
encephalomyelitis. The causative agent of the disease is a
virus classified as lentivirus, a subgroup of retroviruses. MVV
infection is spread all over the world in many countries.
This classification also includes caprine arthritis encephalitis
virus (CAEV), which manifests itself in adult goats mainly in
the form of severe arthritis of the carpal joint (“big knee”).
Animals with CAEV become emaciated despite an intact
appetite, and show poor milk yield. Serologic surveys show
that CAEV is widely disseminated in goat herds on different
continents.
By far, the most important source of infection, however, is
the milk of positive animals. In fact, if the kids are fed with
communal milk from the herd, all the kids can be infected
by just a single positive goat. An effective vaccination
does not exist. For this reason, eradication programs are
based on colostrum deprivation, separation and serological
detection of infected animals.
Specific antibodies in serum for both diseases can be
measured by the same ELISA.
Ruminants Test Kits
‡
Available for sale outside the U.S.
4-5
Chlamydia
Chlamydiophila abortus Antibody ELISA
Test Kit
CHEKIT* Chlamydia Antibody ELISA‡:
2 plates (strips), serum and plasma
samples of ruminants, results within 2.5
hours, adapted to automation (bovine,
caprine, ovine)
Chlamydiophila abortus is known as the responsible agent for
abortion, fertility problems, broncopneumonia and clinically
inapparent infections in cattle and sheep populations.
Chlamydiophila-infected animals show no clinical illness
prior to abortion. Pathogenesis commences when
chlamydial invasion of placentomes produces a
progressively diffuse inflammatory response and tissue
necrosis. Some animals may shed Chlamydiophila spp in
vaginal fluids for more than two weeks before abortion and
up to more than two weeks after abortion. This may explain
the higher incidence of abortion in newly infected herds,
since the susceptibility to infection varies in relation to the
physiological status of the animal.
The success of eradication and surveillance programs
depends on the quality and the effectiveness of available
diagnostic tools. The isolation of the microorganism is
difficult and the diagnosis is mainly based on serological
methods as agent isolation remains a difficult and timeconsuming task.
For this reason, the ELISA is easy to implement, and the
analysis of small numbers of samples, as well as largescale screening, is facilitated with this method.
Ruminants Test Kits
The CHEKIT Chlamydia Test Kit includes chemical reagents
(wash, substrate, stop solutions) that can also be used with
CHEKIT kits that test for other abortive diseases (B. abortus,
neosporosis, toxoplasmosis, Q fever).
‡
Available for sale outside the U.S.
4-6
FMD
Foot-and-Mouth Disease
3ABC Antibody ELISA Test Kit
CHEKIT* FMD Antibody ELISA‡: 5 plates (strips), specific to screening
serum or plasma samples, differentiates
infected from vaccinated animals (DIVA)
(bovine, ovine)
Foot-and-mouth disease (FMD) is a highly contagious
viral disease. It can spread rapidly and affects both
domesticated and wild ruminants, as well as pigs. FMD has an economically devastating impact on affected
countries because trade barriers are imposed in countries
where the disease occurs.
The FMDV genome encodes a single polyprotein that
is cleaved during the translation. Mature structural and
nonstructural viral protein derive after a cascade of further
cleavages by viral proteases.
Vaccine produced according to the guidelines of the OIE
are depleted of nonstructural proteins like 3ABC during
purification steps. Antibody to 3ABC is generally accepted
as the most reliable single indicator for viral replication.
Detection of anti-3ABC antibodies has been described upon
challenge in animals with and without previous vaccination,
confirming the potential of 3ABC as a marker for the
surveillance of vaccinated populations.
Ruminants Test Kits
‡
Available for sale outside the U.S.
4-7
IBR/BHV-1
Infectious bovine rhinotracheitis (IBR) is a highly contagious, infectious disease
that is caused by bovine herpesvirus-1 (BHV-1). In addition to causing respiratory
disease, this virus can cause conjunctivitis, abortions, encephalitis and generalized
systemic infections. Although clinical findings may be highly suggestive of IBR, no
real pathognomonic signs provide a clinical diagnosis for IBR. Therefore, laboratory
confirmation is necessary to identify BHV-1 infection. Confirmation of exposure to
BHV-1 is facilitated by measurement of antibody in serum, plasma or milk. Countries
that have IBR control programs require IBR-free certification prior to shipping cattle to
their regions. ELISA is the test procedure of choice in many European IBR programs.
Infectious Bovine Rhinotracheitis (IBR)/
Bovine Herpesvirus-1 (BHV-1) gE Antibody
ELISA Test Kit
HerdChek* IBR gE Antibody ELISA‡:
6 plates (strips) or 30 plates (solid);
serum, plasma or milk samples;
blocking format; overnight sample
incubation; differentiates infected from
vaccinated animals (DIVA) with BHV-1
gE-deleted vaccines
The HerdChek* IBR gE Test Kit is an enzyme-linked
immunosorbent assay (ELISA) that can be an effective
differential test distinguishing naturally infected from
vaccinated cattle when used in IBR control programs
together with gE-deleted IBR vaccines.
Infectious Bovine Rhinotracheitis
(IBR)/Bovine Herpesvirus-1 (BHV-1) gB
Antibody ELISA Test Kit
HerdChek* IBR gB Antibody ELISA‡: 5 plates (strips) or 30 plates (solid);
serum, plasma or milk samples; blocking
format; results in less than four hours;
optional overnight protocol
The HerdChek* IBR gB Test Kit is an enzyme-linked
immunosorbent assay (ELISA) for the detection of
glycoprotein-B-specific antibodies to bovine herpesvirus-1
(BHV-1) in bovine samples using IBR gB-specific monoclonal
antibodies.
Infectious Bovine Rhinotracheitis (IBR)/
Bovine Herpesvirus-1 (BHV-1) Antibody
ELISA Test Kits
Ruminants Test Kits
CHEKIT* Trachitest Serum Screening
Antibody ELISA‡: 10 plates (strips
and solid), serum or plasma samples
(individual or pool of up to 10), short and
overnight protocols, monophasic format
The CHEKIT*-Trachitest Serum and CHEKIT* BHV1 Bulk
Milk enzyme-linked immunosorbent assay (ELISA) test kits
provide a rapid, simple, sensitive and specific method for
detecting antibodies against bovine herpesvirus-1 (BHV-1).
CHEKIT* Trachitest Serum Verification
Antibody ELISA‡: 10 plates (strips
and solid), serum or plasma samples
(individual or pool of up to 10), short and
overnight protocols, biphasic format
CHEKIT* BHV1 Bulk Milk Antibody
ELISA‡: 10 plates (strips and solid), milk
(individual or pool of up to 50), overnight
protocol
4-8
‡
Available for sale outside the U.S.
M. pt.
Mycobacterium paratuberculosis
Antibody ELISA Test Kits
HerdChek* M. pt. Antibody ELISA‡:
2 plates (strips), 5 plates (solid), serum or plasma samples, indirect format,
results in less than three hours (bovine,
caprine, ovine)
HerdChek* M. pt. Antibody ELISA:
5 plates (strips), serum or plasma
samples, indirect format, results in less
than three hours (bovine)
Johne’s disease is a chronic, debilitating enteritis of
cattle and other ruminants caused by the organism
Mycobacterium paratuberculosis (M. pt.). Every year, Johne’s disease costs the dairy and livestock industries
millions of dollars. Animals may carry the disease for
months or years without showing clinical signs. This slow
progression of the disease may reduce milk production
and compromise the health of other animals in the herd,
which may lead to further financial loss. Implementing a
testing and management program may have significant cost
savings when compared with the potential revenue lost if an
infected herd is not effectively managed.
Ruminants Test Kits
‡
Available for sale outside the U.S.
4-9
Neospora
Neospora caninum Antibody
ELISA Test Kits
HerdChek* Neospora caninum Antibody
ELISA: 2 plates (strips), serum samples,
indirect format, results in less than two
hours (bovine)
CHEKIT* Neospora caninum Antibody
ELISA‡: 2 plates (strips), serum or plasma
samples, short protocol, ready-to-use
reagents (bovine, caprine, ovine)
Neospora caninum is a protozoal (Apicomplexan) parasite that
has been described to cause abortion and neonatal morbidity
in cattle, sheep, goats and horses. It has been reported as
one of the most important causes of abortion in dairy cattle.
The dog has been identified as the definitive host. Primary
transmission is through congenital infections and the spread of Neospora oocysts.
Use of ELISA testing allows herd veterinarians to assess
infection or exposure status and monitor immune status in
Neospora caninum-vaccinated herds.
Ruminants Test Kits
The CHEKIT Neospora Test Kit includes chemical reagents
(wash, substrate, stop solutions) that can also be used with
CHEKIT kits that test for other abortive diseases (B. abortus,
Q fever, toxoplasmosis, Chlamydia).
‡
Available for sale outside the U.S.
4-10
Q Fever
Q Fever Antibody ELISA
Test Kit
CHEKIT* Q Fever Antibody ELISA‡: 2 plates (strips), serum and plasma
samples of ruminants, results within 2.5
hours, adapted to automation (bovine,
caprine, ovine)
C. burnetii has a worldwide occurrence with an increased
prevalence in countries with dense cattle, sheep and goat
populations. There are two independent infection cycles:
wildlife and domestic, via ticks.
The animal infection, also called coxiellosis, is characterized
by a subclinical phase with a relatively rare and sudden
epidemic appearance of abortion.
Use of ELISA testing allows for the serological diagnosis of Q fever by detecting C. burnetii-specific antibodies. In fact,
due to ease and rapidity, ELISA tests have become the major
techniques applied for herd monitoring on a routine basis.
The CHEKIT Q Fever Test Kit includes chemical reagents
(wash, substrate, stop solutions) that can also be used with
CHEKIT kits that test for other abortive diseases (B. abortus,
neosporosis, toxoplasmosis, Chlamydia).
Ruminants Test Kits
‡
Available for sale outside the U.S.
4-11
Toxotest
Toxoplasma gondii Antibody ELISA
Test Kit
CHEKIT* Toxotest Antibody ELISA‡: 2 plates (strips), serum and plasma
samples of ruminants, results within 2.5
hours, adapted to automation, common
incubation times, ready-to-use conjugate
(caprine, ovine)
Toxoplasma gondii is an intestinal coccidium that parasitizes
members of the cat family as definitive hosts and has a
wide range of intermediate hosts. In most cases, infection is asymptomatic, but devastating disease can occur. Cats—definitive hosts—are infected by eating rodents—
intermediate hosts.
Oocysts are spread in the feces. If the oocysts are
ingested by intermediate host mammals, such as rodents,
they develop in the digestive tract. It is here that they
are engulfed by macrophages. Within the macrophage,
tachyzoites develop and travel to various parts of the body
(liver, brain, heart, spleen) via the bloodstream.
The intermediate host may be eaten by a predator, such as
a cat. In addition to public health problems, this zoonosis is
a frequent cause of abortions in sheep and goat herds.
Ruminants Test Kits
The CHEKIT Toxotest Kit includes chemical reagents
(wash, substrate, stop solutions) that can also be used with
CHEKIT kits that test for other abortive diseases (B. abortus,
neosporosis, Q fever, Chlamydia).
‡
Available for sale outside the U.S.
4-12
TSEs
Bovine spongiform encephalopathy (BSE), chronic wasting disease (CWD)
and scrapie belong to the family of transmissible spongiform encephalopathies
(TSEs). Clinically affected animals exhibit ataxia and excessive salivation, along
with other behavioral disturbances. These animals show a progressive loss of
physical condition that precedes death. The modes of transmission and the
causative agents of TSEs are poorly understood. The disease is characterized
by accumulation of a proteinase K-resistant form of the prion protein (PrPSc) in
the medulla oblongata (obex) region of the brain, as well as other tissues (lymph
nodes, spleen, tonsils, etc.).
Bovine Spongiform Encephalopathy
Antigen Test Kit
HerdChek* BSE Antigen EIA: 5 plates
(strips), obex samples, very simple
sample preparation and EIA protocol—no
proteinase K digestion, less equipment
required, all room-temperature, less
hands-on time, results in 2.5 hours
The IDEXX BSE test kit is an antigen-capture enzyme
immunoassay (EIA) for the detection of the abnormal
conformer of the prion protein (PrPSc) in bovine postmortem tissues.
Bovine Spongiform EncephalopathyScrapie Antigen Test Kit
HerdChek* BSE-Scrapie Antigen
EIA‡: 5 plates (strips); obex, spleen
and lymph node samples; very simple
sample preparation and EIA protocol—no
proteinase K digestion, less equipment
required, all room-temperature, less
hands-on time, results in 2.5 hours
HerdChek* CWD Antigen EIA: 5 plates
(strips), retropharyngeal lymph node tissue
samples, very simple sample preparation
and EIA protocol—no proteinase K
digestion, less equipment required, all
room-temperature, less hands-on time,
results in less than 3.5 hours
The IDEXX BSE-Scrapie Test Kit is an antigen-capture
enzyme immunoassay (EIA) for the detection of the
abnormal conformer of the prion protein (PrPSc) in bovine,
caprine and ovine postmortem tissues.
Chronic Wasting Disease
Antigen Test Kit
The IDEXX CWD Test Kit is an antigen-capture enzyme
immunoassay (EIA) for the detection of the abnormal
conformer of the prion protein (PrPSc) in postmortem white-tailed and mule deer retropharyngeal lymph node
tissue. Additional species elk claim is in the late stages of development.
Ruminants Test Kits
‡
Available for sale outside the U.S.
4-13
Ruminants Test Kits
Equine Infectious Anemia (EIA AGID, EIA cELISA)
Ruminants
EquineTest
Test
Kits
Kits
Equine Test Kits
EIA
Equine Infectious Anemia Antibody
Test Kits
EIA Antibody Agar Gel
Immunodiffusion (AGID): 180 tests;
serum samples; 24-hour assay read time; bright, clean lines for easy reading;
virtually no nonspecific lines, leading to high specificity
EIA Antibody cELISA: 92 tests, serum samples, competitive ELISA format, 45-minute assay time, visual test interpretation with optional
spectrophotometric results
Equine infectious anemia (EIA) virus causes a persistent
infection in horses, resulting in periodic episodes of fever,
anemia, thrombocytopenia, leukopenia and weight loss. The virus may be transferred in utero or horizontally by
biting flies, contaminated needles or mother’s milk. Once a
horse is infected with EIA, it will test positive for antibody to
the virus in serological tests and remain infected for life.
Equine Test Kits
5-1
Equine Test Kits
xChek* Software
Ruminants
xChek*Test
Software
Kits
xChek* Software
xChek
*
An Invaluable Tool for
Managing Data
Requirements: Microsoft* Windows*
95 (or later), 32 MB of RAM (64 MB
recommended), 30 MB of available hard-disk space (100 MB recommended),
CD-ROM drive, VGA or compatible display
monitor, Microsoft* Windows*-compatible
printer
Software Features: User-friendly
interface, extensive database merging
options, quick and easy report e-mailing
capabilities, numerous reports for flock
and herd profiling, reliable baselines for
preventive medicine
Developed in 1996, IDEXX xChek* software was the first
Microsoft* Windows*-based software for ELISA result
interpretation in the poultry and livestock industries.
Built on the strong foundation of its predecessors, the
FlockChek* and HerdChek* software programs, the xChek*
program allows rapid input and reporting of individual and
population-based data. It is a comprehensive, user-friendly
diagnostic tool for the interpretation and management of
data pertaining to the health of flock and herd animals.
xChek* software is widely used in private, public and
government sectors. On a daily basis, veterinarians,
laboratory personnel, production managers, farmers and
field service employees around the world use xChek* to
monitor the health of production animals. xChek* data
can be shared between users by hard-copy printouts and
through database export.
The xChek* software collects, organizes and reports pointin-time ELISA information that can be stored, analyzed and
evaluated to determine immediate implications on the flock
or herd health status. Supporting information on animals,
owners and other testing methods performed can also be
stored to assist with data interpretation. xChek* reports can
be used for historical information look-up and trend analysis,
resulting in reliable baselines for monitoring and preventive
medicine programs.
xChek* can search quickly through large amounts of
information and organize it into easy-to-use formats. xChek*
offers several reports that can be customized to meet the
individual user’s needs. If ELISA information needs to be
combined with other diagnostic information, xChek* can
export Microsoft* Excel and text files in predefined or userdefined formats.
xChek* Software
6-1
xChek* Software
Introduction
7-1
ELISA Technology
7-2
ELISA Kits
7-3
ELISA Equipment
7-5
Equipment Maintenance and Calibration
7-6
Reagent Handling and Preparation
7-7
Handling and Preparing Kit Components
7-8
Quality Control
7-9
Sample Handling
7-10
Pipetting Methods
7-12
ELISA Plate Timing
7-15
ELISA Plate Washing
7-16
Reading Plates and Data Management
7-18
ELISA Troubleshooting
7-19
Appendices
A: Gravimetric Pipette Calibration Procedure
B: FlockChek* and HerdChek* Inventory
Control Tracking Chart
7-23
7-24
C: Laboratory Tracking Chart
7-25
D: Maintenance and Calibration Schedule
7-26
E: Quality Control Quick Check
7-27
ELISA Technical Guide
ELISA Technical Guide
ELISA
Introduction
IDEXX Laboratories manufactures diagnostic test kits for the detection of
diseases in ruminants, horses, swine and poultry.
The enzyme-linked immunosorbent assay (ELISA) is one of the most
sensitive and reproducible technologies available. These assays are
rapid, simple to perform and easily automated. IDEXX introduced the first
commercial poultry ELISA for infectious bursal disease (IBD) in 1985 and
the first commercial livestock ELISA for Aujeszky’s disease/pseudorabies in
1986, enhancing the way laboratories test production animals today.
As with any assay, the reproducibility and reliability of ELISAs are dependent
upon proper technique and attention to detail. This ELISA technical guide
will increase your awareness of ELISA techniques and help you maintain
proficiency with this methodology.
Check your package insert for specific instructions for each assay you
perform. Periodically, improvements and revisions are made to kit and
package inserts. Therefore, it is important to review the protocol on a regular
basis. If you have questions concerning any of the following information,
call IDEXX Technical Services for assistance. Within the United States,
call 1-800-548-9997 or 1-800-943-3999. Outside of the United States,
call 1-207-856-0895 and select the Production Animal Services Technical
Support option. Or visit our Web site at www.idexx.com/production/contact/
contactpas.jsp.
ELISA Technical Guide
7-1
ELISA
ELISA Technology
ELISA formats provide the
ability to:
• Test a large number of
samples at the same time
• Automate the procedure using
robotics or other types of
automated equipment
• Computerize the calculation
and reporting of results
The ELISA is a rapid test used for detecting and quantifying antibodies or
antigens against viruses, bacteria and other materials. This method can be
used to detect many infectious agents affecting poultry and livestock.
In ELISA technology, the solid phase consists of a 96-well polystyrene
plate, although other materials can be used. The function of the solid
phase is to immobilize either antigens or antibodies in the sample, as they
bind to the solid phase. After incubation, the plates are washed to remove
any unbound material. Conjugate is then added to the plate and allowed to incubate.
The conjugate consists of either an antigen or antibody that has been labeled
with an enzyme. Depending upon the assay format, the immunologically
reactive portion of the conjugate binds with either the solid phase or the
sample. The enzyme portion of the conjugate enables detection.
The plates are washed again and an enzyme substrate (hydrogen peroxide
and a chromogen) is added and allowed to incubate. Color develops in the
presence of bound enzyme and the optical density is read with an ELISA
plate reader.
ELISA Formats
ELISAs are divided into three main formats—indirect, blocking (competitive)
and antigen-capture (direct).
ELISA Technical Guide
Indirect Format
In the indirect format, the sample antibody is sandwiched between the
antigen coated on the plate and an enzyme-labeled, anti-species globulin
conjugate. The addition of an enzyme substrate-chromogen reagent causes
color to develop. This color is directly proportional to the amount of bound
sample antibody. The more antibody present in the sample, the stronger the
color development in the test wells. This format is suitable for determining
total antibody level in samples (Newcastle disease virus, B. abortus, etc.).
Indirect ELISA
7-2
ELISA
Blocking (Competitive) Format
In this format, the specific sample antibodies compete with, or block, the
enzyme-labeled, specific antibody in the conjugate. The addition of an
enzyme substrate-chromogen reagent causes color to develop. This color
is inversely proportional to the amount of bound sample antibody. The more
antibodies present in the sample, the less color development in the test
wells (CAV, CSFV Ab, etc.).
Blocking ELISA
Antigen-Capture (Direct) Format
In the antigen-capture format, the antigen in the sample is sandwiched
between antibodies coated on the plate and an enzyme-labeled conjugate.
The antibody conjugate can be either monoclonal or polyclonal. The
addition of an enzyme substrate-chromogen reagent causes color to
develop. This color is directly proportional to the amount of the target
antigen present in the sample (CSFV Ag, LLAg, etc.).
ELISA Kits
NOTE: Do not mix or use components from different kit lot numbers.
7-3
ELISA Technical Guide
An ELISA kit is a set of standardized reagents and microwell plates manufactured for a specific test. IDEXX ELISA kits may contain some or all of the following components: coated plates (solid and/or strip plates),
sample diluent, controls, wash concentrate, conjugate, substrate and stop
solution. The kits are manufactured in batches or lots. Each component
of each kit lot is optimized and manufactured to work as a unit. The kits
pass many quality-control tests conducted by IDEXX, numerous worldwide
reference laboratories, and/or the USDA before they are approved and
released for sale.
ELISA
ELISA Kits continued
IDEXX kits are manufactured
in batches or lots according to
strict quality standards. Each
component or reagent in a kit
lot is optimized to work with the
other reagents contained in the
kit. This includes measurements
of sensitivity, specificity and
repeatability. Therefore, it is very
important not to mix reagents
from different kit lots.
Coated Plates
The 96-well plates are made of polystyrene and coated with either
inactivated antigen or antibody. This coating is the binding site for the
antibodies or antigens in the sample. Unbound antibodies or antigens in the sample are washed away after incubation.
For some assays, normal host cell (NHC) antigens and pathogen-specific
antigens are coated in alternating rows or columns. The NHC antigens
are used to assess whether antibodies against tissue culture components
present in vaccines are contributing to sample reactions. When using these
plates, each sample is added to both a sample well and an NHC well.
Sample Diluent
Most assays require a specific dilution of the sample. Samples are added to
the sample diluent and mixed prior to putting them onto the coated plates.
Controls
The positive control is a solution that contains antibody or antigen. The
negative control is a solution without antibody or antigen. The controls
help to normalize or standardize each plate. Controls are also used to
validate the assay and to calculate sample results. In most kits, the controls
are prediluted and ready to use. Be sure to follow the instructions in the
package insert.
Conjugate
All kit components have an
expiration date.
ELISA conjugates are enzyme-labeled antibodies or antigens that react
specifically to plate-bound sample analytes. Unbound conjugate is washed
away after incubation and before the addition of substrate. The optical
density of the colorimetric substrate is directly proportional to the quantity of
bound enzyme present.
Substrate
For peroxidase conjugates, the substrate is a mixture of hydrogen peroxide
and a chromogen that reacts with the enzyme portion of the conjugate to
produce color.
Wash Concentrate
The wash concentrate is a buffered solution containing detergent used to
wash away unbound materials from the plates.
ELISA Technical Guide
Stop Solution
The stop solution stops the enzyme-substrate reaction and, thereby, the
color development.
7-4
ELISA
ELISA Equipment
There is a large selection of
equipment available. When
purchasing a plate reader, call
IDEXX Technical Services to
make sure the xChek* software
has the proper interface.
Equipment for ELISA testing is widely available. Readers, washers and
pipettes are available as manual or automated systems. Some of the
factors affecting equipment selection are the number and types of tests and
samples, technical training of staff, and financial considerations. Below is a
brief outline of some equipment available for performing ELISA testing.
Pipettes
• Single-channel, fixed-volume and adjustable-volume (1–20 µL, 10–100 µL, 20–200 µL, etc.)
• Multichannel, 8- and 12-channels
• Semi-automated dispensing units
• Fully automated systems that can process multiple plates
Multichannel pipette and
single-channel pipette
Dilutors
• Single-channel
• Multichannel
• Automated dispensing units
Washer Systems
• Manual systems that wash one row or column at a time
• Semi-automated systems that handle one strip or plate at a time
• Fully automated systems that can process multiple plates
Semi-automated wash system
ELISA Plate Readers
• Manual readers that read one row or well at a time
• Semi-automated readers that read one plate at a time
• Fully automated systems that can process multiple plates simultaneously
Other
• Humidity chamber (not required for all ELISA tests)
• Plate sealers for assays that have long incubation times (to avoid
evaporation)
Manual wash system
ELISA Technical Guide
Plate reader
7-5
ELISA
Equipment Maintenance and Calibration
Be sure to label your pipettes
with the calibration date and
keep a log for the calibration and
maintenance of all your equipment.
The maintenance and calibration of your laboratory equipment is extremely
important in obtaining accurate and reproducible results.
The Maintenance and Calibration Schedule (Appendix D) should be used
as a guideline. Adjust maintenance routines according to the amount of
daily testing performed in your laboratory. Always refer to your equipment
manufacturer’s guide for their recommendations.
Calibration Protocols
Equipment always needs to be in proper calibration. Equipment that is out of
calibration can produce false or inaccurate results. Refer to the Maintenance
and Calibration Schedule (Appendix D) and your manufacturer’s instructions
for the proper calibration protocol and required frequency.
Options for Calibrating Pipettes
• Perform the gravimetric method outlined in Appendix A.
Pipette with calibration label
• Use a commercial automated calibration system like the PCS®‚ produced
by Artel. See Appendix A, Gravimetric Pipette Calibration Procedure, for
more information.
• Send the pipette to the manufacturer; see your owner’s manual for
instructions.
• Send the pipette to a pipette calibration service.
Sending pipettes out for service is beneficial when repair or maintenance
is necessary. However, this practice provides only a limited level of quality
control, which can be increased with in-house calibration.
ELISA Technical Guide
Operator technique and laboratory environment are two critical variables
that determine how a pipette will perform when used on your bench top.
A thorough quality-control program must include a quantitative account of
these effects. It is beneficial to have a method in place that allows you to
perform regular, routine performance verifications on your own pipettes. By
doing so, you will be able to track pipettes that are drifting out of tolerance.
When this happens, the failing pipette should be sent out for corrective
maintenance or repair by a qualified service before it compromises your
laboratory data and productivity.
7-6
ELISA
Reagent Handling and Preparation
Receiving Kits
When you receive your ELISA kits, record the date on your Inventory Control
Tracking Chart (Appendix B) and on the kit boxes. Inspect them for damage
and store them at 2°–7°C. When using kits from your inventory, use the firstin-first-out (FIFO) method. In other words, use the kits that are the oldest (or
will expire) first. Individual kit components may have longer expiration dates
than the actual date on the outer kit box. However, you should go by the
expiration date on the outer label of the kit. If you do not use an entire kit,
mark the date it is opened and each time it is used thereafter. This way, you
can keep track of how many times it moves or cycles from the refrigerator to
room temperature. Keep the number of cycles to a minimum by batching (or
accumulating) samples into larger groups whenever possible.
Label your kit with the date it
was received.
The contamination of reagents
may compromise your test
results. Labeling your reagent
reservoirs and using a separate
one for each reagent will help
minimize the risk.
Labeled reservoirs
General Reagent Handling
Be sure to check your package insert for guidelines on handling and
preparing reagents. Some test kits recommend that all reagents and plates
be brought to room temperature (18°–25°C) prior to use; others indicate that
only specific reagents be brought to room temperature. When you need
to bring a kit to room temperature (18°–25°C), take it out of the refrigerator
and take the kit components out of the kit box at least 2–3 hours before
beginning the assay.
Measure all reagents using sterile or clean vessels. Be careful to measure
only what is needed for the number of plates being run. This will help to
maintain the integrity of the reagents. Do not return reagents to the original
stock bottles. We strongly recommend using disposable pipettes and
reservoirs when handling reagents to minimize the risk of contamination.
However, if you choose to reuse any disposable device, use a separate
reservoir for each reagent and be sure to label them. Also, wash and
thoroughly rinse the wells with deionized or distilled water after each use.
Change and discard the disposable reservoirs as frequently as possible.
Never use the same reservoir for conjugate and substrate, even if it has
been washed.
ELISA Technical Guide
7-7
ELISA
Handling and Preparing Kit Components
Refer to the package insert for
specific details on the kit you
are using.
Do not exchange components
between kit lot numbers, even
if kits are of similar type. Test
results may be severely and
adversely affected.
Plates
Most assay plates are provided in resealable bags that include a desiccant.
In some kits, the plates are packaged individually. When opening a bag
that contains several plates, remove only the number of plates you need,
reseal the bag and return it to the refrigerator for storage. If possible, the
plates that were taken from the original bag should be stored in another bag
with desiccants in it. If a partial plate is used, aspirate all the liquid from the
used wells and cover them with sealing tape. Place several desiccants in
the bag. Be sure to mark the plate accordingly and store it separately from
unused assay plates. Some kits have the plates formatted into strips. We
recommend labeling each strip before use to prevent confusion.
Sample Diluent and Wash Concentrate
Make sure the sample diluent and wash concentrate have come to room
temperature (18°–25°C) before use. These are usually the largest bottles in
a kit and require the most time to equilibrate. If the wash concentrate still
shows crystal formation after reaching room temperature, mix it by inverting
it several times.
Controls
Seal, label and store partially used
plates in a bag with desiccant.
Most kits are formulated with prediluted controls. However, some require
that you dilute them in the same manner as your sample. Controls should be
added to the plate in the same method and at the same time as the samples.
Conjugate
If the kit requires you to prepare a “working” conjugate solution, be sure to
follow the instructions closely. Prepare only what you immediately need, and
do not save leftover solution for future use. If conjugates are contaminated or
improperly stored, they may lose enzymatic activity or may have an apparent
increase in background. Most kits supply a ready-to-use conjugate.
Substrate
Our ELISA test kits include a ready-to-use substrate. The chemical activity
of the substrate will be compromised if it is exposed to light or comes into
contact with metal. Protect this solution by storing it in a dark container until
ready for use.
Stop Solution
ELISA Technical Guide
Be sure to use the stop solution included in the kit. Follow any safety
precautions in the package insert. The stop solution should be at room
temperature before use. If the stop solution shows crystal formation after
reaching room temperature, mix it by inverting several times. The stop
solution may crystalize at lower temperatures. Before use, make sure that it
is completely dissolved and appears clear.
7-8
ELISA
Quality Control
In-House Controls
We recommend using in-house assay controls to monitor your ELISA
techniques and kit performance over time.
Tracking chart for controls
Because sera are generally received in small quantities, controls will need
to be made by pooling samples. Collect negative and positive samples
separately. When sufficient quantities of each have been collected, pool
similar samples together. Mix the pooled samples thoroughly. In small
quantities, perform the serial dilution of positive sera in negative serum.
Assay each dilution according to standard kit protocol (same sample
dilution as described in the kit package insert). Select the dilution that is
most comparable to the sample-to-positive (S/P) or sample-to-negative
(S/N) values that you want to monitor. Make large quantities of that dilution.
Prefilter the prepared controls using a 0.45-micron filter membrane; you
may then choose to filter with a 0.20-micron filter membrane (optional). Put
a small amount of the freshly made in-house control (volume enough for
1–2 tests) into airtight vials, label, date and store frozen at -70°C if possible.
Keep a record of this preparation in a notebook for reference.
To use this control, thaw, mix and dilute it in the same manner as a routine
sample. Run it on every plate next to the kit controls. Do not refreeze your
in-house control. You can store it for 3–5 days at 4°C.
To help troubleshoot questionable
results, record and graph the
laboratory temperature.
Record your results on the Laboratory Tracking Chart (Appendix C) and
graph them. Any variations or trends should alert you to review your
technique and quality-control measures.
Monitoring Temperature
ELISA tests are sensitive to temperature extremes. Try to maintain a
laboratory temperature of 18°–25°C. Avoid running assays under or near
air vents as this may cause excessive cooling, heating and/or evaporation.
Also, do not run assays in direct sunlight as this may cause excessive heat
and evaporation. Cold bench tops may affect your assay and should be
avoided by placing several layers of paper towels or some other insulating
material under the assay plates during incubation.
Laboratory temperature
tracking chart
Record and track the temperature during each assay. If your laboratory’s
temperature fluctuates from morning to afternoon, record this on your
tracking chart. If you have conditions that are difficult to control, it is a good
idea to use a temperature control chamber to incubate your plates. Using
ELISA plate covers will help control evaporation and accidental spills.
Quality Control Check
ELISA Technical Guide
Use the Quality Control Quick Check (Appendix E) to troubleshoot any problems.
7-9
ELISA
Sample Handling
Incoming Sample Quality
Sample quality can have a significant impact on final assay results. Most labs
have no choice regarding the quality of incoming samples. In many cases,
the sample diluent formulation compensates for variations in sample quality.
Gross fungal or bacterial contamination can have adverse effects on
the antibody or other protein components of a sample and may have an
undesirable effect on test results. If sample quality is highly questionable,
obtaining a fresh sample is strongly advised, when possible.
Meat juice sample
Take sample from the area indicated.
Serum/Plasma Samples
Serum samples with trace hemolysis (light-red color) and moderate lipemia
(milky appearance) may have little or no effect on ELISA results. Avoid using
samples that are heavily hemolyzed (dark-red color) or grossly lipemic.
Check your package insert for information. When serum is on the clot, be
careful not to aspirate any of the clotted material or blood cells.
Meat Juice Samples
Meat juice samples should be as clean as possible. Remove debris and
lipids from the sample when pipetting.
Milk Samples
Whole milk samples can be used after centrifugation for 15 minutes at 2000 x g, or left overnight if refrigerated (2°–8°C). The sample intended for
the assay should be drawn from below the cream layer.
Egg Yolk Samples
Collect samples with a clean tuberculin syringe and mix the diluted samples
thoroughly by vortexing.
ELISA Technical Guide
Blood sample
Take sample from the area indicated.
Milk sample
Take sample from the area indicated.
7-10
Other Sample Types
Refer to your package insert for sample handling, preparation and storage
of other sample types (e.g., albumin, cloacal swabs).
ELISA
Sample Handling continued
Avoid numerous freeze-thaw
cycles, as this may damage
the antibodies or antigens in
the sample. We recommend no
more than 3–5 cycles.
Storing Samples
Be sure samples are properly stored. In general, serum samples should
be refrigerated at 2°–7°C for no more than 3–5 days. If samples need to be
stored for a longer period, they should be removed from the clot and frozen
to a minimum of -20°C. Make sure all stored samples are properly labeled
and sealed to prevent evaporation. Evidence of lyophilization (concentration
of the sample) can be seen as crystalization and is common in self-defrosting
freezers. This should be avoided because the integrity of the sample will most
likely be compromised.
Using Frozen Samples
Frozen samples can be thawed at room temperature or in a refrigerator. All
thawed samples need to be thoroughly mixed prior to dilution to ensure that
the proteins are dispersed throughout the sample. Mix by gentle vortexing
or inverting at least five times. Frothing or over-mixing of samples will cause
denaturation of serum proteins.
Light hemolysis
Dark hemolysis
ELISA Technical Guide
Unmixed thawed sample
Proteins settled on bottom of tube;
mix prior to taking sample
7-11
ELISA
Pipetting Methods
Use standard (forward) pipetting
for the preparation of sample
dilutions, and reverse pipetting
for the addition of diluted
samples, controls and reagents.
Two pipetting methods used for ELISA are standard (forward) and reverse.
Not all pipettes are capable of reverse pipetting. Refer to the instructions
included with your pipette for details.
Use standard (forward) pipetting for the preparation of sample dilutions, and
reverse pipetting for the addition of diluted samples, controls and reagents.
Careful pipetting is crucial for accurate test results. Become familiar with the
pipette and both methods before running actual tests. Be sure to use the correct pipette and tip (volume capacity) for the volume being transferred.
Pipetting Technique
1. Draw the calibrated volume of sample into the tip.
2. Touch the side of the tube with the tip to remove the excess liquid.
3. Ensure that you have the proper volume of sample in the tip.
When using a multichannel pipette, if the wells on your plate are empty,
position the tips into the lower corner of each well, making contact with the
plastic. If the wells on your plate contain liquid, position the tips above the
liquid, making contact with the plastic.
Pipetting a Sample
Draw up the calibrated
volume of sample into the tip.
The drop on the tip needs
to be removed.
Touch the side of the tube
with the tip to remove the
excess liquid.
Proper Pipetting
ELISA Technical Guide
Proper position to dispense reagents into
empty wells using a multichannel pipette;
in the lower corner of each well
Proper position to dispense reagents into
wells containing liquid using a
multichannel pipette; above the liquid
7-12
Ensure that you have the
proper volume of sample
in the tip.
ELISA
Pipetting Methods continued
Standard (Forward) Pipetting and Sample Preparation
1. Put a new tip on a single-channel pipette and make sure that it is on tight.
2. Press the plunger to the first stop.
3. Some manufacturers recommend that you prewet the tip by aspirating
and expelling an amount of the sample. Check the instructions that came
with your pipette.
4. Draw the calibrated volume of sample into the tip and pause for one
second with the tip still in the sample. Be careful not to place the tip too
deeply into the sample.
5. Touch the tip to the side of the sample container to remove any excess
liquid on the outside of the tip.
6. Dispense the sample into the measured diluent by depressing the
plunger past the first stop to the second stop. Be careful not to place the
tip too deeply into the sample diluent.
For samples less than or equal to 10 µL: After dispensing the sample
into the diluent, rinse the pipette tip in the diluent by pushing the plunger
down 2–3 times before ejecting the tip.
7. Mix samples with a multichannel pipette prior to dispensing samples onto
the plate. You can do this by pushing the plunger down 3–6 times.
8. Eject the tip into a waste container.
Reverse Pipetting Using a Multichannel Pipette
1. Put new tips on the pipette. Make sure they are on tight and straight.
2. Press the plunger past the first stop and halfway to the second stop.
3. Draw the liquid in a slow motion, being careful not to draw any air
bubbles into the tips. Check for consistency of volume in the tips.
4. Touch the tips to the edge of the reagent reservoir to remove excess
liquid on the outside of the tips.
a. If the wells on your plate are empty, position the tips into the lower
corner of each well, making contact with the plastic.
Bent tips; needs to be replaced
b.If the wells on your plate contain liquid, position the tips above the
liquid, making contact with the plastic.
5. Slowly dispense the liquid into the wells by depressing the plunger to the
first stop. Be careful not to splash liquid out of the wells, and make sure
there are no drops left on the tips.
7. Eject the tips into a waste container.
NOTE: Reverse pipetting uses more reagent/volume (=“dead volume”).
7-13
ELISA Technical Guide
6. To repeat, hold the plunger at the first stop and continue with step 3.
ELISA
Pipetting Methods continued
Automated equipment
uses more reagent/volume
than semi-automated.
Check your manufacturer’s
recommendations for purging
and priming your system.
Automated Dilution Systems and Competitive Assays
For those systems and assays using neat samples or lower dilution factors, the
sample can be put directly into the wells of the coated plates.
Follow the sequence below:
1. Add the diluent to the plate.
2. Add the sample into the diluent.
ELISA Technical Guide
3. Mix by tapping the plate or repeating pipetting.
7-14
ELISA
ELISA Plate Timing
To minimize incubation time
between controls and samples,
rack the controls with the samples
and add them to your plate using
a multichannel pipette.
Adding Samples and Controls
Incubations for assay plates should be timed as precisely as possible.
Usually the process of adding samples to the plate requires the most
time. When you dispense samples onto the plate, it is critical to keep the
time difference between the first and the last sample to a minimum.
Use a multichannel pipette whenever possible to minimize the time
interval from the start of the plate to the end. If the time interval becomes
too long or you are interrupted while adding samples to the plate, put a
positive control and/or your in-house control at the end of the plate and
compare these results with the controls at the beginning of the plate.
For tighter control over the time differentiation from when the controls
and samples are added, you can put your controls in a tube that is
racked in position with your samples. Then use a multichannel pipette
and put the controls onto the plate at the same time you are adding the
samples.
Multiple Plate Runs
When timing multiple plates, it is important to keep track of the time
interval from the first plate to the last plate in the run. Keep your batch
sizes small enough so your processes do not overlap. You do not want
to be washing a plate (unless it is done by an automated washer) while
another needs to have conjugate added. If possible, use a timer for
every plate.
ELISA Technical Guide
Use several timers when incubating multiple plates.
7-15
ELISA
ELISA Plate Washing
After tapping out plates, check
paper towels for any evidence of
color. This may indicate that the
plates were not washed properly
and there are reagents remaining
in or around the wells.
Automated or Semi-Automated Systems
In general, an automated or semi-automated wash system in proper working
order will provide more consistent washing than manual methods. Check
that all the dispensing needles are dispensing with a smooth, steady stream
and that all aspiration ports aspirate uniformly.
Make sure your wash system is properly cleaned and maintained. Refer to
the Equipment Maintenance and Calibration section in this guide (page 7-6)
and your owner’s manual for proper maintenance. The plate-washing
technique should be consistent from plate to plate and from row to row
within a plate. Avoid prolonged soak times unless specifically recommended
in the package insert.
Prepare the wash solution according to the package insert. Use only the
wash solution included with, or recommended for, your kit.
Aspirate reagents from the plate before dispensing the wash solution.
Follow the specific recommendations in your package insert for the
number of washes to use at each step of an assay. Most assays require
approximately 300–350 µL per well per wash. Be careful to fill the wells
above the level of the reagents. Do not allow wells to overflow. If this occurs,
the test results may be invalid.
Microbial growth in wash system
tube; needs to be replaced
Do not allow the plate to dry between plate washings and prior to the
addition of reagents.
After the final aspiration, tap out any remaining liquid onto several layers of
paper towels.
ELISA Technical Guide
When testing milk, albumin, whole blood or yolk samples, take extra care to
inspect the wells. Because of their protein or fat composition, these sample
types are sometimes more difficult to wash from the wells and may require
the maximum recommended number of wash cycles.
7-16
ELISA
ELISA Plate Washing continued
Manual or Semi-Manual Systems
Work quickly so the time from washing the first well/row to the last is
minimal. If the time is too long, the empty wells may dry out and the last
wells will have a longer incubation than the first wells.
Washing by multichannel pipette or wash bottles should be avoided if
at all possible as this gives insufficient washing that results in high and
inconsistent background.
Overflowing plate; this can
contaminate other wells
Make sure to aspirate all the liquid from the wells by placing the aspiration
needles at the bottom and in the corners of the wells. Do not scrape the
surface of the plate as this will remove the antigen/antibody bound to the
surface and cause inconsistent or inaccurate results. After aspiration, wells
should not dry before the addition of the next reagent.
After tapping out the plates, check the paper towels for any evidence of
color. This may indicate that the plates were not washed properly and there are reagents remaining in or around the wells.
Proper position of manual washer needles for dispensing
wash solution
Proper position of manual washer needles for aspirating liquid
ELISA Technical Guide
7-17
ELISA
Reading Plates and Data Management
Reading Plates
The last step in an ELISA is to read and interpret the results. For most
assays, the optical density (amount of color) of the solution on the plate
is read with a spectrophotometer, commonly known as a plate reader.
There are many models and manufacturers of plate readers; refer to the
manufacturer’s instructions for details of operation.
The package insert specifies which wavelength is required for the assay.
Most assays specify the absorbance reading at 450 nm or 650 nm. Most
assays are optimized using a plate reader equipped with a 650-nm filter.
Other filters can be used, but will result in lower optical density (OD) values.
The use of 630-nm or 620-nm filters will lower the OD values of both the
controls and samples, but will do so equivalently across the entire plate. The use of these alternative filters will not affect the test results.
Plates should be read as soon as possible following the addition of stop
solution. Absorbance readings may drift if excessive time elapses between
stopping the reaction and reading the plates.
Data Management
ELISA Technical Guide
IDEXX provides the xChek* software to assist you in the collection and
management of the data from your ELISA assays. The xChek software
interfaces with most common plate readers to read the plate, send the
optical densities to the computer and calculate the results. An IDEXX
Technical Services representative can assist you in learning more about this software.
The xChek* software interfacing with the ELISA plate reader
7-18
ELISA
ELISA Troubleshooting
This information is intended to help you troubleshoot your ELISA procedure. If you need assistance, contact IDEXX Production Animal Services Technical
Services at 1-207-856-0895, option 2; or 1-800-548-9997, option 2.
NOTE: The conditions described here may not pertain to every ELISA kit
because performance requirements vary for individual assays. Be sure to
check your package insert for specifications.
Issue:
High background or excessive color development
(high optical density (OD) readings)
Possible Causes
Poor-quality water was used
to wash plates or to prepare
wash solution.
Substrate solution
has deteriorated.
There was insufficient
washing or poor
washer performance.
Washer system had
microbial contamination.
Wash system contained an
alternate wash formulation.
Reader was malfunctioning or
not blanked properly;
this is a possible cause if the
OD readings were high and
the color was not dark.
Laboratory temperature was
too high or too low.
Check the water quality. If it is questionable, try substituting an alternate water
source, such as bottled distilled water, to wash plates or prepare the wash solution.
Make sure the substrate is colorless prior to addition to the plate.
Try using the highest number of washes recommended for the assay. Make sure
that at least 350 µL of wash solution is dispensed per well per wash. Verify the
performance of the washer system. Have the system repaired if any ports drip,
dispense or aspirate poorly.
Clean out microbial contamination by flushing the system with a dilute solution of
bleach (10% by volume) followed by a large amount of distilled or deionized water.
Prime the system with the appropriate wash solution before use. The tubing may
need to be changed if the contamination is heavy.
Be sure each unique wash solution is properly labeled. Prime the system
thoroughly when switching between solutions.
Verify the reader’s performance using a calibration plate and check the lamp
alignment. Verify the blanking procedure, if applicable, and reblank.
Maintain the room temperature within 18°–25°C. Avoid running assays near heat
sources, in direct sunlight or under air vents.
Ensure that the correct reagents were used, that working solutions were prepared
correctly and that contamination has not occurred.
7-19
ELISA Technical Guide
Reagents were intermixed,
contaminated or
prepared incorrectly.
Recommended Actions
ELISA
ELISA Troubleshooting continued
Issue:
Insufficient color development (low optical density (OD) readings)
Possible Causes
Laboratory temperature
was too low.
Wash solution was prepared
incorrectly or the wrong wash
solution was used.
Recommended Actions
Maintain the room temperature within 18°–25°C. Avoid running assays under air
conditioning vents or near cold windows.
Be sure to use the wash solution recommended for the kit and that it is prepared
correctly. Label each unique wash solution to avoid using the wrong one.
Washer system had
microbial contamination
or contained an alternate
wash formulation.
Clean out microbial contamination by flushing the system with a dilute solution of
bleach (10% by volume) followed by a large amount of distilled or deionized water,
then prime the system with the appropriate wash solution. Be sure each unique
wash solution is properly labeled. Prime the system thoroughly when switching
between solutions.
Too many wash cycles
were used.
Stay within the recommended range for the number of wash cycles. Try to use the
lowest number of washes recommended for the assay.
Incubation periods were
too short.
Follow protocol for incubation times. Time each plate separately to ensure
accurate incubation periods.
Reagents and plates were
too cold.
Make sure plates and reagents are at room temperature by taking them out of the
refrigerator, and the kit components out of the kit box, at least 2–3 hours before
starting the assay.
Reagents were expired or
intermixed from a different
lot number.
Verify the expiration dates and lot numbers on the reagents.
Wrong conjugate was
used, conjugate was
prepared incorrectly or
has deteriorated.
Be sure that the conjugate used is the one that came with the kit and all
conjugates are kit- and lot-specific. If preparation of a working conjugate is
needed, be sure that the concentrate and diluent are mixed in appropriate
volumes. Do not prepare the working solution too far in advance, and do not save
any unused portion for future use. If no conjugate preparation is necessary, be
sure to pour out only the amount required for immediate use, and do not return
any unused portion to the stock bottle.
Assay plate read was at
wrong wavelength, or reader
was malfunctioning.
Verify the correct wavelength for the assay and read the plate again. Verify reader
calibration and lamp alignment.
Positive control was diluted
(indirect format only).
ELISA Technical Guide
Excessive kit stress
has occurred.
Assay plates were
compromised or
previously used.
7-20
Do not dilute controls unless specified in the package insert.
Check records to see how many times the kit has cycled from the refrigerator.
Check to see if the kit was left out on a loading dock or other area for too long or at extreme temperatures.
Be sure to refrigerate plates in sealed bags with a desiccant to maintain stability.
Prevent condensation from forming on plates by allowing them to equilibrate
to room temperature while in the packaging. If partial plates are used, be sure
to label used wells to prevent reuse; cover them with sealing tape and use the
remaining wells as soon as possible. Do not store partially used plates with other
plates. Include a desiccant in the storage bag.
ELISA
ELISA Troubleshooting continued
Issue:
Replicates within a plate show poor reproducibility
Possible Causes
Recommended Actions
Excessive time was taken
to add samples, controls or
reagents to the assay plate.
Be sure to have all materials set up and ready to use quickly. Use a multichannel
pipette to add reagents to multiple wells simultaneously. Rack controls with
samples and dispense them onto the plate at the same time as the samples.
Multichannel pipette was not
functioning properly.
Verify pipette calibration and check that tips are on tight. Be sure all channels of
the pipette draw and dispense equal volumes.
There was inconsistent
washing or washer
system malfunctioning.
There was poor distribution
of antibody in the sample.
Verify the performance of the washer system. Have the system repaired if any
ports drip or dispense/aspirate poorly.
If the sample was thawed or refrigerated, make sure it was mixed prior to dilution.
Diluted samples also need to be mixed prior to adding them to the plate.
Issue:
No color development
Possible Causes
Reagents were used in the
wrong order or an assay step
was omitted.
Samples were not added to
diluent (indirect format only).
Wrong conjugate was
used, conjugate was
prepared incorrectly or
has deteriorated.
Recommended Actions
Check the package insert for the assay protocol and repeat the assay.
Verify that the samples were added to the diluent.
Be sure that the conjugate used is the one that came with the kit. All conjugates are
kit- and lot-specific. If preparation of a working conjugate is needed, be sure that the
concentrate and diluent are mixed in correct volumes. Do not prepare the working
solution too far in advance and do not save any unused portion for future use. If no
conjugate preparation is necessary, be sure to pour out only the amount required for
immediate use and do not return any unused portion to the stock bottle.
ELISA Technical Guide
7-21
ELISA
ELISA Troubleshooting continued
Issue:
Poor reproducibility plate to plate
Possible Causes
Recommended Actions
Inconsistent incubation times
occurred from plate to plate.
Time each plate separately to ensure that plates have consistent incubation periods.
Inconsistent washing
occurred from plate to plate.
Use the same number of washes for each plate. Verify the performance of the washer
system. Have the system repaired if any ports drip or dispense or aspirate poorly.
Pipette was working
improperly.
Kit controls and samples
were at different
temperatures.
ELISA Technical Guide
Reagents were being used
from different kit lots
7-22
Check the pipette calibration. Verify that pipette tips are on tight before use and
that all channels draw and dispense equal volumes.
Be sure to allow sufficient time for sample diluent, samples and kit controls to
come to room temperature by removing them from the kit box. Larger volumes
will require longer equilibration time. If using a water bath to hasten equilibration,
make sure it is maintained at room temperature; do not use a warm water bath for
controls, samples or diluent.
If running two different kit lots at the same time, make sure to label reagent trays,
etc. so all reagents within a lot are used with the corresponding plates.
ELISA
Appendix A:
Gravimetric Pipette Calibration Procedure
For adjustable-volume pipettes,
calibrations should be performed
at a minimum of two settings—a
low volume and a high volume at
commonly used settings.
Materials
•Pipette
•Analytical Balance
•Glass Beaker
•Deionized or Distilled Water
•Weighing Vessel
•Thermometer
Procedure
Automated Calibration
System
The ARTEL PCS® Pipette
Calibration System is an
automated instrument/reagent
system designed to facilitate
regular and frequent verification
of pipette performance and
operator technique. For more
information call ARTEL,
at 1-888-406-3463 or
1-207-854-0860, or visit their
Web site at artel-usa.com.
1. To avoid erroneous results due to evaporation, we recommend
humidifying the analytical balance chamber at least two hours prior to
calibration. This can be achieved by placing a small, half-filled beaker of
water into the chamber with all its doors closed.
2. Follow the manufacturer’s directions for cleaning and lubrication of
pipettes prior to calibration.
3. The room temperature should remain constant within ±0.5°C, preferably
between 18°–25°C.
4. Allow a sufficient volume of deionized or distilled water to come to room
temperature and take a temperature reading.
5. Record the beginning weight of the weighing vessel or zero the balance.
6. Using a new pipette tip with each delivery, pipette water into the weighing
vessel and record the weight. Repeat this step 10 times.
7. Calculate the volume dispensed for each delivery.
Calculations
1. Calculate the actual volume delivered as follows:
Volume = Weight of Water
Density of water at 16°–21°C = 0.998 mg/µL
Density of water at 22°–25°C = 0.997 mg/µL
Density of Water
2. Calculate the mean (M), standard deviation (SD) and coefficient of
variance (CV) of the 10 volumes to determine precision of the pipette.
3. Determine the accuracy of the pipette as follows: (1-[Difference between stated and actual volume/stated volume]) x100 = % accuracy
Recommended specifications
1. Precision: CV ≤5.0%
2. Accuracy: ≥95%
ELISA Technical Guide
Labeling
Label the pipette with the calibration date, the technician’s initials, the
precision and the accuracy. Also, record these data in a laboratory
notebook or log for long-term storage.
7-23
Lot Number
Number of Kits Received
Date Received
Expiration Date
Comments
7-24
Appendix B:
Inventory Control Tracking Chart
Kit Type
ELISA Technical Guide
ELISA
te
7-25
Ki
N t Lo
u
m t
be
r
Te
c
ELISA Technical Guide
re
Ro
Te om
m
p
In
Co tern
a
n
tro l
l
In
Co tern
a
n
tro l
l
In
C tern
o
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In
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o
nt al
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P
o
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tro e
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Ne
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o
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Pl
Nu ate
m
be
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s
*FlockChek, HerdChek and CHEKIT are trademarks or registered trademarks of IDEXX Laboratories, Inc. in the United States and/or other countries.
Da
n
cia
hn
i
Assay Name:
tu
er
a
Appendix C:
FlockChek*, HerdChek* and CHEKIT* Laboratory Tracking Chart
Comments
ELISA
ELISA
Appendix D:
Maintenance and Calibration Schedule
Procedure
Daily
Weekly
Monthly
Quarterly
Yearly
Pipettes
Clean exterior
X
Check calibration
X
Clean interior and O-rings
X
Dilutor
Flush system with DI water
X
Purge system
X
Check calibration*
X
Soak syringes
X
Check/Change tubings
Washer
Flush system with DI water when
using wash solutions other than DI
water and when changing between
wash solution types Change
Check
Change
X
Check traps, filters and foaming
X
Check aspiration and dispensing
needles for drips and debris
X
Check tubes and bottles for microbial growth
X
Decontamination—flush system with
bleach or alcohol solution*
X
Calibration check—Purge system
X
Clean exterior
X
Run through “clean cycle” with DI water*
X
Check/Change tubing
ELISA Technical Guide
Check
Reader
Calibration plate**
Lamp alignment
X
After bulb replacement
Clean optics
Clean exterior
X
X
*Refer to your manufacturer’s guide for specific instructions for your make and model.
**Call IDEXX Technical Services or your reader manufacturer for recommendations on calibration plates.
7-26
ELISA
Appendix E:
Quality Control Quick Check
Monitoring and tracking assay performance on quality-control charts
provides insight as to when it is necessary to troubleshoot problems. Below is a checklist to review if you are having problems with your ELISA.
If you have followed all the steps below and are still having problems with
your assay, contact IDEXX Production Animal Services Technical Services at 1-207-856-0895, option 2; or 1-800-548-9997, option 2.
✔ Equipment
mKeep preventive maintenance up-to-date
mCalibrate and clean pipettes
mCalibrate reader
mSanitize and maintain wash system
✔ Reagents
mMaintain inventory control—FIFO
mInspect components
mBring reagents to room temperature
mAvoid contamination
mVerify proper storage
✔ Technique
mMonitor sample quality
mVerify reagent preparation
mVerify appropriate sample mixing
mVerify proper pipetting
mCheck timing—multiple plate runs
mCheck washing of assay plates
mUse in-house controls—track results
✔ Other
ELISA Technical Guide
mMonitor laboratory temperature
mUse sterile disposables and reservoirs
mMonitor assay performance; record in a log
7-27
ELISA Technical Guide
ELISA
Notes
7-28