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About
®
EliGene
Respiratory
Infections
Sexually
Transmitted
Urinary
Infections
Viral
Infections
Elisabeth Pharmacon
2014
Bacterial
Infections
Human
Genetics
DNA/RNA
Isolation
&
Spectral
Calibration
Instruments
ZEPHYRUS®
elisabeth
pharmacon group
EliGene®
ZEPHYRUS
B I O S A F E T Y B OX
®
ZEPHYRUS®
ZEPHYRUS®
P C R B O X H A R M O NY
M A G N E T O
Content
Pages:
1. Introduction
02 - 06
About
®
EliGene
07 - 10
Respiratory
Infections
11 - 14
Sexually
Transmitted
Urinary
Infections
Product Guarantee Policy
Shipping
EliGene Kits and RealTime PCR Instruments
Sample Processing - Sampling, Transport, Storage, DNA Isolation
2. Diagnostics of Respiratory Infections
Legionella pneumophila (CE)
Mycoplasma pneumoniae (CE)
Chlamydophila pneumoniae (CE)
Influenza A/B/pandemic (CE)
3. Diagnostics of Sexually Transmitted Urinary Infections
Chlamydia trachomatis (CE 1023)
Neisseria gonorrhoeae (CE)
Ureaplasma sp. (CE)
Mycoplasma hominis/genitalium (CE)
15 - 24
4. Viral Diagnostics
Adenovirus (CE)
Norovirus (CE)
Enterovirus (CE)
Herpes simplex virus 1 (CE)
Herpes simplex virus 2 (CE)
Varicella zoster virus (CE)
Hepatitis B virus (CE 1023)
Hepatitis C virus (CE 1023)
5. Bacterial Diagnostics
25 - 27
Bacterial
Infections
28 - 30
Human
Genetics
31 - 33
DNA/RNA
Isolation
&
Spectral
Calibration
34 - 35
Instruments
ZEPHYRUS®
Mycobacterium tuberculosis (CE)
Borrelia (CE)
Anaplasma (CE)
6. Diagnostics for Human Genetics
Coeliac disease (CE)
Lactose intolerance (CE)
Spondylitis HLA-B27 (CE)
7. DNA Isolation and Spectral Calibration Kits
®
EliGene MTB Isolation kit (CE)
EliGene® Urine Isolation kit (CE)
®
EliGene Spectral Calibration kit LC
EliGene® Spectral Calibration kit LC2
EliGene® Spectral Calibration plate
8. Instruments ZEPHYRUS
®
ZEPHYRUS® Magneto and isolation kits MAGNETO
®
ZEPHYRUS Bio Safety Box
ZEPHYRUS® PCR Box Harmony
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Viral
Infections
01
1 - Introduction
1. ELISABETH PHARMACON
About
®
EliGene
ELISABETH PHARMACON was founded in Brno, Czech Republic in 2001 by Czech biologist Omar Šerý and
businessman Petr Čermák. In its early days the company was known for research, development and
producing DNA diagnostics under the brand name EliGene®. ELISABETH PHARMACON cooperates with
many research institutes and clinics. Since its beginnings, the company has cooperated with the Faculty of
Science, Masaryk University. In 2008, it became the first Czech company to produce DNA diagnostics under
EN ISO 13485:2003 certification. Since 2012 company develops and manufactures laboratory instruments,
e.g. biohazard boxes. The quality systems EN ISO 13485:2012 and EN ISO 9001:2008 cover all activities such
as designing, developing, manufacturing, purchasing, storage and selling of in-vitro diagnostic medical
devices and molecular biology testing, measuring and analysis. The aim is to offer the best quality products
at the best available price on the market.
By 2013, the company had invested more than 2,4 million EUR in laboratory instruments and the research.
The company's research is continually supported by grant projects of the Investment and Business
Development Agency (CzechInvest) and the Ministry of Industry and Business of the Czech Republic. In
2012, ELISABETH PHARMACON company was founded also in United Kingdom for the distribution of
products. Both companies are members of ELISABETH PHARMACON Group.
2. Product guarantee policy
Product quality is our highest priority at ELISABETH PHARMACON. Clinical studies are available for all CE
certified kits. As manufacturing, testing, packaging and delivery are carried out under the EN ISO
13485:2003 quality system, we guarantee batch-to-batch consistency. All EliGene products are supplied
with an Certificate of Analysis, summarizing the conditions and results of quality assurance tests.
3. Shipping
EliGene diagnostic kits must be shipped frozen on dry ice, for short journeys, kits can be shipped packed on
blue ice. Freeze-thaw cycles lower the sensitivity of diagnostic kits. Diagnostic kits are packed in polystyrene
boxes to protect them from thawing during shipment and laboratory work.
02
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4. EliGene kits and RealTime PCR instruments
EliGene® kits are divided according to detection chemistry to basic lines RT, LC and UNI, respectively.
About
®
EliGene
In RT line, the TaqMan chemistry using hydrolyzing fluorescence probes is used in the combination with hotstart polymerase and ROX dye as a passive reference (see Figure 1A). The total reaction volume is 25ul when
5ul of isolated DNA is added directly to ready-to use mastermix containing all necessary components for
reaction. In RT kits, the fluorescence dyes with excitation wavelengths of 495, 520 nm and emission
wavelengths of 520, 550 nm together with non-fluorescence quenchers are used. These kits are basically
designed for ABI instruments of 7000 line and they are optimized on LightCycler480 instrument but can be
used with other instruments that meet the spectral characteristics of used fluorescence dyes. The typical
time of the run for RT kits is about 2 hours. The list of compatible instruments you can find in Table 1.
In LC line, the Molecular Beacons chemistry using hybridization fluorescence probes is used in combination
with hot-start polymerase (see Figure 1B). The total reaction volume is 20 ul when 5 ul of isolated DNA is
added to mastermix created by mixing of polymerase with reaction mix containing all necessary
components for the reaction. The fluorescence dyes with excitation wavelengths of 495, 460 and emission
wavelengths of 515, 705 together with non-fluorescence quenchers are used. The LC kits are basically
designed for LightCycler 1.2 and 2.0 instruments but they can be used with other instruments, that meets on
the one hand the spectral characteristics of used fluorescence dyes in individual versions and on the other
hand rapid heating/cooling rate. The typical time of run for LC kits is about 1 hours. The list of compatible
instruments you can find in Table 1.
In UNI line, the Molecular Beacons chemistry using hybridization fluorescence probes is used in
combination with hot-start polymerase (see Figure 1B). The total reaction volume is 20 ul when 5 ul of
isolated DNA is added directly to ready-to use mastermix containing all necessary components for reaction.
The usage of uracil in the reaction mix enables UNG treatment to prevent the re-amplification of carryoverPCR products by removing any uracil incorporated into single- or double-stranded DNA. In UNI kits the
fluorescence dyes with excitation wavelengths of 495, 520 and emission wavelengths of 520, 550 together
with non-fluorescence quenchers are used. These kits are designed for almost all real-time instruments on
the market. The typical time of run for UNI kits is about 50 minutes. The list of compatible instruments you
can find in Table 1.
Figure 1. Basic principle of TaqMan hydrolization probes (A) and Molecular Beacons hybridization probes (B)
action.
Annealing
Annealing
The TaqMan probe hybridizes between the primers in annealing step and during elongation step probe is cleaved by exonuclease
activity of Taq Polymerase. The Molecular Beacon probe forms beacon-like structure that opens to specific sequence during the
hybridization in annealing step (B).
“TaqMan” is registered trademark of Roche Molecular Systems Inc., “Molecular Beacon” is registered trademark of Public Health
Research Institute Properties, Inc. (PHRI)
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03
1 - Introduction
Table 1. Kit / Instrument Compatibility Chart
About
®
EliGene
The gray ticks mean that the compatibility was not tested and it is based only on technical specification
of individual instruments.
Instrument
RT kits Uni kits LC kits
Compatibility tested with individual kits (*not tested)
ABI 7300 (Applied Biosystems)
*
ABI 7500FAST (Applied Biosystems)
LightCycler 2.0 (Roche)
LightCycler 480 (Roche)
LightCycler Nano (Roche)
Smart cycler (Cepheid)
RotorGene 6000 (Qiagen)
RotorGeneQ
RotorGene 3000 (Qiagen)
Compatibility on the basis of filter configuration (not tested)
Illumina Eco (Illumina)
Mastercycler® ep realplex (Eppendorf)
Mx3000P QPCR System (Agilent)
Mx3005P QPCR System (Agilent)
iQ5 & MyiQ Cycler (Bio-Rad)
CFX96 & CFX384 (Bio-Rad)
SLAN® Real-Time PCR (Shanghai Hongshi Medical
Technology Co., Ltd.)
ExiCycler™ 96 (Bioneer)
Advantages of EliGene® isolation kits

04
Isolation on columns enables robust and rapid isolation of DNA
Special composition of provided reagents and columns guarantees high yields and minimize risk of
inhibition during PCR
Isolation procedures tested directly with variety of clinical samples covering blood, plasma, serum,
sputum, urine, sperm and buccal, cervical, eye, rectal and urethral swabs
Kits are CE certified IVD
New fully automatic isolation of DNA/RNA from all kinds of clinical samples by ZEPHYRUS® Magneto
instrument
CE certified kits for automatic isolation fully compatible with EliGene® Real Time PCR kits.
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Advantages of EliGene® detection kits

The usage of the Internal Control allows to monitor the extraction procedure and to check possible PCR
inhibition
Kits are supplied with positive control allowing quality control of the analysis
Simultaneous amplification (multiplex) of DNA/RNA from pathogen and internal control in one PCR
tube
The usage of Hot-Start technology minimize the risk of non-specific reaction and provide ultimate
sensitivity 1-10 copies of nucleic acid/reaction
The usage of Ready to use Mastermixes containing all reaction components for easier reaction setup and
handling
The usage of uracil in the Mastermixes of UNI kits enables UNG treatment to prevent the reamplification of carryover-PCR products by removing any uracil incorporated into single- or doublestranded DNA
To ensure the highest quality of our kits, the row material from the leading providers is used
Quantitative Standards in the most of the kits enables to calculate precisely the NA load from 102 to 106
copies/reaction
High stability of used reagents
The usage of EliGene® UNI kits enables complete detection in 50 minutes
Kits are CE certified IVD
Kits are validated on LightCycler® 1.2, 2.0, Nano and 480, ABI® 7300, 7500 and 7500FAST, RotorGene-Q
(RotorGene 6000)
Compatibility of the kits with broad range of instruments - RotorGene®, SmartCycler®, Mx3000P® and
3005P® QPCR systems or iQ5®, CFX96® and other cyclers (see details in Table 1)
About
®
EliGene
LightCycler is registered trademark of Roche Molecular Systems Inc., "ABI" is a registered trademark of Applied Biosystems. "iQ5",
"MyiQ" and " CFX96 & CFX384 " are registered trademarks of Bio-Rad Laboratories. "SmartCycler" is registered trademark of Cepheid.
"Rotor-Gene" is a trademark of Corbett Research. "MX 3000P" and "MX3000P" are registered trademarks of Stratagene, Inc.
5. Sample processing - sampling, transport, storage,
DNA isolation
The amplification success depends mostly on the quality of the purified NA. The highest sensitivity in PCR
detection can only be achieved when you have first-rate sample preparation. The quality of sample
preparation depends not only on isolation procedure but also on timing of collection (clinical relevancy),
transport and storage conditions. The presence of conservation agents in transport media could lead to PCR
inhibition as well. Taking together all these facts, the isolation procedure must be rigorously tested to avoid
false negative results.
EliGene® and MAGNETO isolation kits offer to customer optimized solution with the high quality reagents
for manual and automatic preparation. EliGene® and MAGNETO isolation kits are rigorously tested in order
to exclude the presence of contaminants and PCR inhibitors. Together with EliGene® detection kits
including Internal Control, EliGene® or MAGNETO isolation kits, we provide optimal solution for accurate
and high sensitive diagnostics.
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05
1 - Introduction
6. Catalog numbers of products
About
®
EliGene
Respiratory
Infections
Sexually
Transmitted
Urinary
Infections
Viral
Infections
Bacterial
Infections
Human
Genetics
DNA/RNA
Isolation
&
Spectral
Calibration
Instruments
®
ZEPHYRUS
Standards
TM
EliDNA
Name of kit
RT line
LC line
UNI line
EliGene® Chlamydophila pneumoniae (CE)
None
None
90057-UNI
EliGene® Legionella pneumophila (CE)
None
None
90056-UNI
EliGene® Mycoplasma pneumoniae (CE)
None
None
90055-UNI
EliGene® Influenza A/B/pandemic (CE)
None
90058-LC*
None
EliGene® Chlamydia trachomatis (CE 1023)
None
90046-LC
90046-UNI
EliGene® Neisseria (CE)
None
90047-LC
90047-UNI
EliGene® Ureaplasma (CE)
None
90049-LC
90049-UNI
EliGene® Mycoplasma hom/gen (CE)
None
90052-LC
90052-UNI
EliGene® Adenovirus (CE)
90036-RT
None
None
EliGene® Enterovirus (CE)
None
90053-LC
None
EliGene® Norovirus (CE)
None
90054-LC
None
EliGene® HSV1 (CE)
90035-RT
None
None
EliGene® HSV2 (CE)
90038-RT
None
None
EliGene® VZV (CE)
90040-RT
None
None
EliGene® HBV (CE 1023)
90037-RT
None
90037-UNI
EliGene® HCV (CE 1023)
None
90059-LC*
None
EliGene® MTB (CE)
90030-RT
90030-LC
90030-UNI
EliGene® Borrelia (CE)
90024-RT
90024-LC
90024-UNI
EliGene® Anaplasma phagocytophilum (CE)
None
None
90061-UNI
EliGene® Coeliac (DQ2, DQ8, DRB4 | CE)
90048-RT
None
None
EliGene® Coeliac (DQ2, DQ8 | CE)
90048-RT28
None
None
EliGene® Lactose Intolerance C-13910T (CE)
None
90044-LC
None
EliGene® Lactose Intolerance G-22018A (CE)
None
90045-LC
None
EliGene® Spondylitis HLA-B27 (CE)
90060-RT
None
None
EliGene® MTB Isolation kit
50 isolations
90043-50
EliGene® MTB Isolation Supplement
50 isolations
1220-MTB-IZO
EliGene® Urine Isolation Kit
50 isolations
90051-50
EliGene® Sperm Isolation Supplement
50 isolations
90051-50SPO
Magneto BodyFluid DNA/RNA isolation kit
180 isolations
M-7001-180
Magneto 1ml Serum/Plasma DNA/RNA isolation kit
180 isolations
M-7002-180
EliGene® Spectral calibration kit
1 run
90099-SC
EliGene® Spectral calibration kit2
1 run
90099-SC2
EliGene® Spectral calibration plate
1 x 96 well plate
90099-SCP
ZEPHYRUS® PCR Box Harmony (Basic)
668 x 670 x 760
Z-PCR-600
ZEPHYRUS® PCR Box Harmony (Eco)
668 x 670 x 760
Z-PCR-600-E
ZEPHYRUS® PCR Box Harmony (Active)
668 x 670 x 760
Z-PCR-600-A
ZEPHYRUS® Bio Safety Box (120)
1207 x 1530 x 855
Z-BSB-120
ZEPHYRUS® Bio Safety Box (150)
1507 x 1530 x 855
Z-BSB-150
ZEPHYRUS® Bio Safety Box (180)
1807 x 1530 x 855
Z-BSB-180
ZEPHYRUS® Magneto
620 x 520 x 550
Z-MAG-12
EliDNA™ Adenovirus
QRT standard
5 reactions
90036-CL.QRT
EliDNA™ HSV1
QRT standard
5 reactions
90035-CL.QRT
EliDNA™ HSV2
QRT standard
5 reactions
90038-CL.QRT
EliDNA™ VZV
QRT standard
5 reactions
90040-CL.QRT
EliDNA™ Lactose Intolerance C-13910T
LC standard
20 reactions
90044-CL.LC
EliDNA™ Lactose Intolerance G-22018A
LC standard
20 reactions
90045-CL.LC
* kit is compatible with LightCycler® 480, LightCycler® Nano, LightCycler® 2.0 and also with ABI 7300, ABI 7500, RotorGene 6000/Q
06
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2.1 Legionella pneumophila
Kit
EliGene® Legionella pneumophila UNI Kit
90056-UNI
Package size:
Analytical specificity:
Specimens:
50 reactions
Legionella pneumophila
sputum, bronchoalveolar lavage, nasopharyngeal swab, nasopharyngeal
aspirate
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480, LightCycler® Nano, LightCycler® 2.0, fully compatible with
RotorGene instruments.
CE certification:
Detection technology:
Clinical study description and results
Yes, CE
Molecular Beacons
new kit, clinical study will be finished in 2014
Respiratory
Infections
Description:
Legionella pneumophila is a thin, aerobic, gram-negative, non-encapsulated, aerobic bacillus of the genus Legionella
containing more than 50 species. It is the best-characterized member of the genus and major causative agent of
Legionnaires' disease, a severe form of acute pneumonia. L. pneumophila is an intracellular pathogen when the bacteria
invades and replicates in alveolar macrophages and epithelial cells. Macrolides (azithromycin) or fluoroquinolones
(moxifloxacin) are the standard treatment for L. pneumophila in humans. About 3 to 8% of all community-acquired
pneumonias are caused by L. pneumophilla when the case-fatality rate for patients with legionellosis is between to 30%
with elderly and immunocompromised patients at greater risk of death.
References:
Heuner K, Swanson M. 2008. Legionella: Molecular Microbiology. Caister Academic Press. ISBN 978-1-904455-26-4.
Newton HJ, Ang DK, van Driel IR, Hartland EL. 2010. Molecular pathogenesis of infections caused by Legionella
pneumophila. Clin Microbiol Rev. 23(2):274-98
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07
2. Diagnostics of Respiratory Infections
2.2 Chlamydophila pneumoniae
Respiratory
Infections
Kit
EliGene® Chlamydophila pneumoniae UNI Kit
90057-UNI
Package size:
Analytical specificity:
Specimens:
50 reactions
Chlamydophila pneumoniae
sputum, bronchoalveolar lavage, nasopharyngeal swab, nasopharyngeal
aspirate
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480, LightCycler® Nano, LightCycler® 2.0, fully compatible with
RotorGene instruments.
CE certification:
Detection technology:
Clinical study description and results
Yes, CE
Molecular Beacons
new kit, clinical study will be finished in 2014
Description:
Chlamydophila pneumoniae is an obligate intracellular pathogen. It is a member of the Chlamydiae, a diverse range of
obligate intracellular bacteria that includes parasites of amoebae, fish, reptiles, mammals, and humans. Today C.
pneumoniae is associated with a remarkably wide range of chronic diseases as meningoencephalitis, arthritis or
myocarditis and it is the second most often cause of community-acquired pneumonia. C. pneumoniae spreads by drop
infection but in the form of infection elementary bodies (EB) resistant to environmental stresses, it can persist on the
surfaces or be spreaded by the aerosol. During the life about 80% of population is infected by C. pneumoniae however in
majority of cases the infection is asymptomatic. Some forms of pneumonia can be very serious but in these cases is not
clear if C. pneumoniae is not only co-pathogen.
References:
Pignanelli S, Shurdhi A, Delucca F, et al. 2009. Simultaneous use of direct and indirect diagnostic techniques in atypical
respiratory infections from Chlamydophila pneumoniae and Mycoplasma pneumoniae. J Clin Lab Anal. 23 (4): 206–9.
Roulis E, Polkinghorne A, Timms P. 2013. Chlamydia pneumoniae: modern insights into an ancient pathogen. Trends
Microbiol. 21(3):120-8.
08
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2.3 Mycoplasma pneumoniae
Kit
EliGene® Mycoplasma pneumoniae UNI
90055-UNI
Package size:
Analytical specificity:
Specimens:
50 reactions
Mycoplasma pneumoniae
sputum, bronchoalveolar lavage, nasopharyngeal swab, nasopharyngeal
aspirate
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480, LightCycler® Nano, LightCycler® 2.0, fully compatible with
RotorGene instruments.
CE certification:
Detection technology:
Clinical study description and results
Yes, CE
Molecular Beacons
new kit, clinical study will be finished in 2014
Respiratory
Infections
Description:
Mycoplasma pneumoniae belonging to class Mollicutes has one of the smallest known genomes. It lacks a
peptidoglycan cell wall and thus it is resistant to the effects of penicillins and other beta-lactam antibiotics. In humans it
causes atypical pneumonia. M. pneumoniae infects the upper and lower respiratory tracts, leading to upper respiratory
tract infection, tracheobronchitis, bronchitis and community-acquired pneumonia. Infections of the lower respiratory
tract generally manifest with a cough, sometimes with dyspnoea, adenopathy, wheezing and, rarely, with respiratory
failure. Although M. pneumoniae infections are usually mild, and many are asymptomatic, they are not always selflimiting. M. pneumoniae causes up to 40% of cases of community-acquired pneumonia, but M. pneumoniae respiratory
tract infections are also associated with a wide range of extra-pulmonary manifestations, including neurological,
cardiac, dermatological, musculoskeletal, haematological and gastrointestinal symptoms.
References:
Sánchez-Vargas FM, Gómez-Duarte OG. 2008. Mycoplasma pneumoniae-an emerging extra-pulmonary pathogen. Clin
Microbiol Infect. 14(2):105-17.
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09
2. Diagnostics of Respiratory Infections
2.4 Influenza virus
Respiratory
Infections
Kit
EliGene® Influenza A/B/pandemic LC
90058-LC
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
YES
Influenza virus A, B, A/H1N1
1 – 10 viruses in the amplified sample
serum, plasma, sputum, bronchoalveolar lavage, nasopharyngeal swab,
nasopharyngeal aspirate, tissue
Compatible instruments:
ABI 7000, 7300, 7500 (Applied Biosystems), LightCycler® 480 and
LightCycler® Nano, LightCycler® 2.0 (Roche), RotorGene 6000 or
RotorGene Q (Qiagen)
CE certification:
Detected DNA region:
Detection technology:
Clinical study description and results
Yes, CE
genome
Molecular Beacons probes
new kit, clinical study will be finished in 2013
Detection technology description:
The EliGene® Influenzavirus A/B/H1N1 kit is intended for detection and quantification of Influenza A, B and H1N1 virus
RNA. Kit contains reversion transcription in one mix with RealTime detection. Results of analyzes distinguishes Influenza
A, B and H1N1 virus types.
Pathogen description:
Influenza is an infectious disease of birds and mammals caused by RNA viruses of the family Orthomyxoviridae, the
influenza viruses. The most common symptoms are chills, fever, runny nose, sore throat, muscle pains, headache,
coughing, weakness and general discomfort. Approximately 33% of people with influenza are asymptomatic. Flu can
occasionally lead to pneumonia, either direct viral pneumonia or secondary bacterial pneumonia, even for persons who
are usually very healthy. Typically, influenza is transmitted through the air by coughs or sneezes, creating aerosols
containing the virus. Influenza can also be transmitted by direct contact with bird droppings or nasal secretions, or
through contact with contaminated surfaces. Influenza viruses can be inactivated by sunlight, disinfectants and
detergents. Often, new influenza strains appear when an existing flu virus spreads to humans from another animal
species, or when an existing human strain picks up new genes from a virus that usually infects birds or pigs. An avian
strain named H5N1 raised the concern of a new influenza pandemic after it emerged in Asia in the 1990s, but it has not
evolved to a form that spreads easily between people. In April 2009 a novel flu strain evolved that combined genes from
human, pig, and bird flu. Initially dubbed "swine flu" and also known as influenza A/H1N1. Influenza B almost exclusively
infects humans. This type of influenza mutates at a rate 2–3 times slower than type A.
References:
Rose N, Hervé S, Eveno E, Barbier N, Eono F, Dorenlor V, Andraud M, Camsusou C, Madec F, Simon G. 2013. Dynamics of
influenza A virus infections in permanently infected pig farms: evidence of recurrent infections, circulation of several
swine influenza viruses and reassortment events. Vet Res. 44(1):72.
Labella AM, Merel SE. 2013. Influenza. Med Clin North Am. 97(4):621-45.
Cha RM, Smith D, Shepherd E, Davis CT, Donis R, Nguyen T, Nguyen HD, Do HT, Inui K, Suarez DL, Swayne DE, PantinJackwood M. 2013. Suboptimal protection against highly pathogenic avian influenza (H5N1) viruses from Vietnam in
ducks vaccinated with commercial poultry vaccines. Vaccine. S0264-410X(13)01137-7.
10
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M A G N E T O
3.1 Chlamydia trachomatis
Kits
EliGene® Chlamydia trachomatis LC Kit
90046-LC
EliGene® Chlamydia trachomatis UNI Kit
90046-UNI
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
On demand
Chlamydia trachomatis, including “Sweden variant“
1 – 10 copies of bacterial DNA in the amplified sample
urine, swabs (cervical, urethral, conjunctival, rectal), sperm
Compatible instruments:
LC Kit is intended for LightCycler® 2.0, LightCycler® 480 instruments
UNI Kit is intended for ABI7500FAST, SmartCycler, RotorGene 6000 and
for all LightCycler® instruments
CE certification:
Detected DNA region:
Detection technology:
yes, certified by Notified Body No. 1023, ITC Zlín
Chlamydia cryptic plasmid and genome (ompA gene)
Molecular Beacons
Sexually
Transmitted
Urinary
Infections
Clinical study description and results:
Within the frame of testing the functional characteristics of EliGene® Chlamydia trachomatis LC kit overall 120 clinical
specimens were analyzed and results were verified by Cobas® Amplicor® Chlamydia trachomatis Test and by DNA
sequencing of 16S ribosomal RNA gene. From 120 clinical samples, 81 samples were C. trachomatis positive and 36
specimens were right determined by EliGene® Chlamydia trachomatis LC kit as C. trachomatis negative.
Sensitivity: 98,78%
Specificity: 100%
Pathogen description:
Chlamydias are gram-negative bacteria that parasitize inside the susceptible cells due their disability to synthesise ATP.
Genus Chlamydia covers four known species: C. trachomatis, C. psittaci, C. pecorum and C. pneumoniae when C. psittaci
and C. pecorum primary belongs to animal parasites. Chlamydia trachomatis is the most frequently reported sexually
transmitted pathogen when up to 10% of population is infected.
References:
Jurstrand M, Christerson L, Klint M, Fredlund H, Unemo M, Herrmann B. 2010. Characterisation of Chlamydia
trachomatis by ompA sequencing and multilocus sequence typing in a Swedish county before and after identification of
the new variant. Sex Transm Infect. 86(1): 56-60.
Kalwij S, Macintosh M, Baraitser P. 2010. Screening and treatment of Chlamydia trachomatis infections. BMJ. 21:
340:c1915.
Figure 2. Analysis of Positive sample by EliGene® Chlamydia trachomatis LC kit on
LightCycler® 2.0 instrument showing results in 530 channel (Chlamydia detection)
and 705 channel (Internal Control detection)
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11
3. Diagnostics of Sexually Transmited Urinal Infections
3.2 Neisseria gonorrhoeae
Sexually
Transmitted
Urinary
Infections
Kits
EliGene® Neisseria LC Kit
90047-LC
EliGene® Neisseria UNI Kit
90047-UNI
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
On demand
Neisseria gonorrhoeae
1 – 10 copies of bacterial DNA in the amplified sample
urine, swabs (cervical, urethral, conjunctival, rectal), sperm
Compatible instruments:
LC Kit is intended for LightCycler® 2.0, LightCycler® 480 instruments
UNI Kit is intended for ABI7500FAST, SmartCycler, RotorGene 6000 and
for all LightCycler® instruments
CE certification:
Detected DNA region:
Detection technology:
yes
genome (porA gene)
Molecular Beacons
Clinical study description and results:
Within the frame of testing the functional characteristics of EliGene® Neisseria LC Kit, totally 88 clinical specimens were
analyzed and the results were determined by Cobas® Amplicor® CT/NG Test. The EliGene® Neisseria LC Kit diagnosed as
Neisseria gonorrhoeae positive all 35 specimens. Totally 53 specimens were rightly determined by EliGene® Neisseria LC
kit as Neisseria gonorrhoeae negative.
Sensitivity: 100%
Specificity: 100%
Pathogen description:
Neisseria gonorrhoeae (gonococci) are gram-negative, non-motile and non-sporulated diplococci. Of the eleven
species of Neisseria that colonize humans, only two are pathogens, N. gonorrhoeae and N. meningitis. N. gonorrhoeae is
the causative agent of sexually transmitted disease gonorrhoea that is the third most common sexual disease.
Worldwide, about 62,2 millions cases of gonorrhoea are registered. The simultaneous infection by Chlamydia
trachomatis is common.
References:
Whiley DM, Buda PP, Freeman K, Pattle NI, Bates J, Sloots TP. 2005. A real-time PCR assay for the detection of Neisseria
gonorrhoeae in genital and extragenital specimens. Diagn Microbiol Infect Dis. 52(1):1-5.
Whiley DM, Garland SM, Harnett G, Lum G, Smith DW, Tabrizi SN, Sloots TP, Tapsall JW. 2008. Exploring 'best practice'
for nucleic acid detection of Neisseria gonorrhoeae. Sex Health. 5(1):17-23.
12
elisabeth
pharmacon group
EliGene®
ZEPHYRUS
®
B I O S A F E T Y B OX
ZEPHYRUS®
ZEPHYRUS®
P C R B O X H A R M O NY
M A G N E T O
3.3 Ureaplasma sp.
Kits
EliGene® Ureaplasma LC Kit
90049-LC
EliGene® Ureaplasma UNI Kit
90049-UNI
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
On demand
Ureaplasma urealyticum, Ureaplasma parvum
1 – 10 copies of bacterial DNA in the amplified sample
urine, swabs (cervical, urethral, conjunctival, rectal), sperm
Compatible instruments:
LC Kit is intended for LightCycler® 2.0, LightCycler® 480 instruments
UNI Kit is intended for ABI7500FAST, SmartCycler, RotorGene 6000 and
for all LightCycler® instruments
CE certification:
Detected DNA region:
Detection technology:
yes
genome (ureA gene)
Molecular Beacons
Sexually
Transmitted
Urinary
Infections
Clinical study description and results:
Within the frame of testing the functional characteristics of EliGene® Ureaplasma LC Kit, totally 57 clinical specimens
(urine, swabs) were analyzed. The EliGene® Ureaplasma LC Kit diagnosed correctly 9 specimens as Ureaplasma
urealyticum or Ureaplasma parvum positive. Totally 48 specimens were rightly determined by EliGene® Ureaplasma LC
kit as Ureaplasma urealyticum or Ureaplasma parvum negative.
Sensitivity: 100%
Specificity: 100%
Pathogen description:
Ureaplasma urealyticum is bacterium belonging to the family Mycoplasmataceae. U. urealyticum consists of 14
serovars that can be divided into two biovars, biovars 1 and biovars 2, on the basis of genotypic characteristics. Evidence
has been presented that the species currently known as U. urealyticum should be separated into two species, namely, U.
parvum (previously, U. urealyticum biovar 1) and U. urealyticum (previously, U. urealyticum biovar 2). The majority of
human Ureaplasma isolates belong to the new species U. parvum. Ureaplasma sp. is the main cause of non-gonococcal,
non-chlamydial urethritis, acute prostatitis and acquired arthritis in men. In pregnant and non-pregnant women,
Ureaplasma can cause chorioamnionitis and pre-term delivery, abortion, pre-term birth, vaginitis and cervicitis. Clinical
studies have demonstrated that infants born to infected mothers become infected with these bacteria, and colonization
of the respiratory tract of infants has been associated with pneumonia, respiratory distress and meningitis.
References:
Allam AB, Alvarez S, Brown MB, Reyes L. 2011. Ureaplasma parvum infection alters filamin A dynamics in host cells.
BMC Infect Dis. 11:101.
Pinna GS, Skevaki CL, Kafetzis DA. 2006. The significance of Ureaplasma urealyticum as a pathogenic agent in the
paediatric population. Curr Opin Infect Dis. 19(3):283-9.
Figure 3. Analysis of Clinical specimens by EliGene®
Ureaplasm LC kit on LightCycler® 2.0 instrument including
Positive control (PC)
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13
3. Diagnostics of Sexually Transmited Urinal Infections
3.4 Mycoplasma hominis / genitalium
Sexually
Transmitted
Urinary
Infections
Kits
EliGene® Mycoplasma hom/gen LC Kit
90052-LC
EliGene® Mycoplasma hom/gen UNI Kit
90052-UNI
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
On demand
Mycoplasma hominis, Mycoplasma genitalium
1 – 10 copies of bacterial DNA in the amplified sample
urine, swabs (cervical, urethral, conjunctival, rectal), sperm
Compatible instruments:
LC Kit is intended for LightCycler® 2.0, LightCycler® 480 instruments
UNI Kit is intended for ABI7500FAST, SmartCycler, RotorGene 6000 and
for all LightCycler® instruments
CE certification:
Detected DNA region:
Detection technology:
yes
genome (gap gene – M. hominis; mgpB gene – M. genitalium)
Molecular Beacons, multiplex PCR distinguishes between M. hominis and
M. genitalium in LC kit
Clinical study description and results:
Within the frame of testing the functional characteristics of EliGene® Mycoplasma hom/gen LC Kit, totally 60 clinical
specimens (urine, swabs) were analyzed. The EliGene® Ureaplasma LC Kit diagnosed correctly 8 specimens as
Mycoplasma hominis and 5 specimens as Mycoplasma genitalium positive. Totally 47 specimens were rightly
determined by EliGene® Mycoplasma hom/gen LC Kit as Mycoplasma hominis or Mycoplasma genitalium negative.
Sensitivity: 100%
Specificity: 100%
Pathogen description:
Mycoplasma sp. are bacteria belonging to the family Mycoplasmataceae that are a group of microorganisms that can
involve the respiratory and genital tracts in man to produce clinical disease. They are found in about 70% of sexually
active humans.
Mycoplasma hominis is associated with pelvic inflammatory disease, bacterial vaginitis, post partum fever, sepsis and
infections of the central nervous system often leading to serious conditions.
In case of Mycoplasma genitalium there is strong evidence of having a causative role in non-chlamydia, non-gonococcal
urethritis (NCNGU) in men and cervicitis in women. M. genitalium infection has also been associated with pelvic
inflammatory disease but the exact role has not yet been determined.
References:
Edberg A, Jurstrand M, Johansson E, Wikander E,Höög A, Ahlqvist T, Falk L, Skov J, Fredlund H. 2008. A comparative
study of three different PCR assays for detection of Mycoplasma genitalium in urogenital specimens from men and
women. Journal of Medical Microbiology,57,304–309.
Baczynska A, Svenstrup HF, Fedder J, Birkelund S, Christiansen G. 2004. Development of real-time PCR for detection of
Mycoplasma hominis. BMC Microbiology, 4:35.
Simpson T, Oh MK. 2004. Urethritis and cervicitis in adolescents. Adolesc Med Clin. 15(2):253-71.
14
elisabeth
pharmacon group
EliGene®
ZEPHYRUS
®
B I O S A F E T Y B OX
ZEPHYRUS®
ZEPHYRUS®
P C R B O X H A R M O NY
M A G N E T O
4.1 Adenovirus
Kit
EliGene® Adenovirus RT Kit
90036-RT
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
EliDNA ADV QRT standard
all 51 types of Adenoviruses
1 – 10 copies of viral DNA in the amplified sample
urine, CSF, feces, swabs, serum, plasma
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480 and NANO fully compatible with RotorGene instruments.
CE certification:
Detected DNA region:
Detection technology:
yes
genome (37 hexon gene)
TaqMan probes
Clinical study description and results:
Within the frame of testing the functional characteristics of EliGene® Adenovirus RT kit overall 350 clinical specimens
were analyzed. From these 350 clinical specimens were 20 positive. EliGene® Adenovirus RT kit analyzed 20 samples as
positive. Totally 330 clinical specimens were properly by EliGene® Adenovirus RT kit analyzed as negative.
Sensitivity: 100%
Specificity: 100%
Viral
Infections
Pathogen description:
Adenoviruses represent the largest nonenveloped viruses. They are able to be transported through the endosome. They
are about 70 to 90 nm in size, nonenveloped icosahedral viruses that are made from up to 252 capsomeres containing
double-stranded DNA. Infection is usually transmitted by droplets of respiratory or ocular secretions, by alimentary
transfer or sexual intercourse and by the contact with contaminated things or water (virus is able to live out of body at
temperature 20°C and lower for a few weeks). The places of the first reproduction are the most often cells of epithelial
conjunctiva, nasopharinx and intestine. Adenovirus infection is clinically manifested by the fever, upper respiratory tract
infections, tonsillitis, laryngitis, bronchitis and pneumonia.
References:
Bannister BA, Begg NT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
Jothikumar N, Cromeans TL, Hill VR, Lu X, Sobsey MD, Erdman DD. 2005. Quantitative real-time PCR assays for
detection of human adenoviruses and identification of serotypes 40 and 41. Appl Environ Microbiol. 71(6):3131-6.
Hlinomazová Z, Loukotová V, Horáčková M, Serý O. 2010. The treatment of HSV1 ocular infections using quantitative
real-time PCR results. Acta Ophthalmol. 10: 1755-3768.
Figure 4: Analysis of EliGene® ADV QRT Standards of 102 to 106 cop/reaction on
ABI®7300 Real-time PCR cycler with resulted Standard curve
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15
4. Viral Diagnostics
4.2 Norovirus
Kit
EliGene® Norovirus LC Kit
90054-LC
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
YES
Norovirus genogroup 1 and 2
1 – 10 viruses in the amplified sample
serum, plasma, feces, water, CSF
Compatible instruments:
ABI 7500 (Applied Biosystems), LightCycler® 480 and LightCycler® Nano,
LightCycler® 2.0 (Roche), RotorGene 6000 or RotorGene Q (Qiagen).
CE certification:
Detected DNA region:
Detection technology:
yes
genome
Molecular Beacons probes
Detection technology description:
The EliGene® Norovirus LC kit is intended for detection and quantification of Norovirus RNA. Kit contains reversion
transcription in one mix with RealTime detection.
Viral
Infections
Clinical study description and results:
Within the frame of testing functional characteristics of the EliGene® Norovirus LC kit overall 100 clinical specimens
were analyzed. From these specimens 100 blind specimens were verified by reference method (La Rosa et al. 2010).
From these 100 samples 56 samples were Norovirus RNA positive. The EliGene® Norovirus LC kit diagnosed as Norovirus
RNA positive all 56 specimens. There were no discrepant results. Totally 44 specimens were right determined by the
EliGene® Norovirus LC kit as Norovirus RNA negative.
Sensitivity: 100%
Specificity: 100%
Pathogen description:
Noroviruses are a genetically diverse group of single-stranded RNA, non enveloped viruses in the Caliciviridae family.
The viruses are transmitted by fecally contaminated food or water, by person-to-person contact, and via aerosolization
of the virus and subsequent contamination of surfaces. Noroviruses are the most common cause of viral gastroenteritis
in humans. Norovirus affects people of all ages. Outbreaks of norovirus infection often occur in closed or semiclosed
communities, such as long-term care facilities, overnight camps, hospitals, prisons, dormitories, and cruise ships, where
the infection spreads very rapidly either by person-to-person transmission or through contaminated food. Norovirus is
rapidly inactivated by either sufficient heating or by chlorine-based disinfectants, but the virus is less susceptible to
alcohols and detergents, as it does not have a lipid envelope.
References:
BannisterBA, BeggNT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
Oberste MS, Maher K, Kilpatrick DR, Pallansch MA. 1999. Molecular evolution of the human Noroviruses: correlation
of serotype with VP1 sequence and application to picornavirus classification". J. Virol. 73 (3):1941–8
16
elisabeth
pharmacon group
EliGene®
ZEPHYRUS
®
B I O S A F E T Y B OX
ZEPHYRUS®
ZEPHYRUS®
P C R B O X H A R M O NY
M A G N E T O
4.3 Enterovirus
Kit
EliGene® Enterovirus LC Kit
90053-LC
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
Yes
Enteroviruses (Coxsackie A, Coxsackie B and Echovirus)
1 – 10 copies of viral RNA in the amplified sample
CSF, feces, respiratory samples, pericardial fluid, blood
Compatible instruments:
LightCycler® 2.0, LightCycler® 480, LightCycler® NANO, ABI7500 and
RotorGene 6000
CE certification:
Detected DNA region:
Detection technology:
yes
genome
Molecular Beacon
Clinical study description and results:
Clinical study will be finished in 2013
Pathogen description:
Human Enteroviruses are a genus of (+)ssRNA viruses associated with several human and mammalian diseases. Until
now, 66 serotypes are known of which there are 62 non-polio enteroviruses that can cause disease in humans: 23
Coxsackie A viruses, 6 Coxsackie B viruses, 28 echoviruses, and 5 other enteroviruses. Enteroviruses are ubiquitous
pathogens with a high incidence worldwide (ca. 500 million infections/year) and are often found in the respiratory
secretions (e.g., saliva, sputum, or nasal mucus) and stool of an infected person. Enteroviruses may cause lifethreatening infections, especially among children when diseases such as myocarditis, paralysis, multiple organ failure,
meningitis and encephalitis may be associated with infections.
Viral
Infections
References:
Bannister BA, Begg NT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
Oberste MS, Maher K, Kilpatrick DR, Pallansch MA. 1999. Molecular evolution of the human enteroviruses: correlation
of serotype with VP1 sequence and application to picornavirus classification". J. Virol. 73 (3): 1941–8.
Santti, Juhana; Heli Harvala, Leena Kinnunen, and Timo Hyypia. 2000. "Molecular epidemiology and evolution of
coxsackievirus A9. Journal of General Virology 81 (5): 1361–1372.
Figure 5: Analysis of Positive sample by EliGene® Enterovirus LC kit on LightCycler® 2.0
instrument showing results in 530 channel (Enterovirus detection)
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17
4. Viral Diagnostics
4.4 Herpes simplex virus 1
Kit
EliGene® HSV1 RT Kit
90035-RT
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
EliDNA HSV1 QRT standard
Herpes simplex virus 1
1 – 10 copies of viral DNA in the amplified sample
CSF, biopsy, swabs (skin, mucosa), blood
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480 and NANO, fully compatible with RotorGene instruments.
CE certification:
Detected DNA region:
Detection technology:
yes
genome (US4 gene)
TaqMan probes
Clinical study description and results:
Viral
Infections
Within the frame of testing the functional characteristics of EliGene® HSV1 RT kit overall 636 clinical specimens were
analyzed. From these 636 clinical specimens 201 were positive. EliGene® HSV1 RT kit analyzed 197 samples as positive.
Totally 435 clinical specimens were properly by EliGene® HSV1 RT kit analyzed as negative.
Sensitivity: 98,04%
Specificity: 100%
Pathogen description:
HSV1 is enveloped DNA virus. To date, at least eight different herpesvirus species are known to infect man. A primary
infection typically occurs during the childhood. The primary infection goes often through symptomless. The HSV1
infection can manifest as an undifferentiational catarrh of respiratory system of adults or older children. The infection of
the conjunctiva and cornea manifests as keratoconjunctivitis. HSV1 is a neurotropic virus.
References:
Bannister BA, Begg NT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
Hlinomazová Z, Loukotová V, Horáčková M, Serý O. 2010. The treatment of HSV1 ocular infections using quantitative
real-time PCR results. Acta Ophthalmol. 10: 1755-3768.
18
elisabeth
pharmacon group
EliGene®
ZEPHYRUS
®
B I O S A F E T Y B OX
ZEPHYRUS®
ZEPHYRUS®
P C R B O X H A R M O NY
M A G N E T O
4.5 Herpes simplex virus 2
Kit
EliGene® HSV2 RT Kit
90038-RT
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
EliDNA HSV2 QRT standard
Herpes virus 2
1 – 10 copies of viral DNA in the amplified sample
CSF, biopsy, swabs (skin, mucosa), blood
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480 and NANO, fully compatible with RotorGene instruments.
CE certification:
Detected DNA region:
Detection technology:
yes
genome (US4 gene)
TaqMan probes
Clinical study description and results:
Within the frame of testing the functional characteristics of EliGene® HSV2 RT kit overall 211 clinical specimens were
analyzed. From these 211 clinical specimens 24 were positive. EliGene® HSV2 RT kit analyzed all 24 samples as positive.
Totally 187 clinical specimens were properly by EliGene® HSV2 RT kit analyzed as negative.
Sensitivity: 100%
Specificity: 100%
Viral
Infections
Pathogen description:
Herpesvirus is enveloped DNA virus. To date, at least eight different herpesvirus species are known to infect man. HSV2
causes genital herpes, characterized by sores in the area of the genitalia. Genital herpes is a sexually transmitted
disease. The main symptoms are herpes ulcers in glans penis in men. Lesions in woman are localized in cervix, vagina or
perianal area. The activation of the latent infection can produce asymptomatic releasing of virus in cervical secret. HSV2
can cause infection of the brain (encephalitis) in the case of severely defective or compromised immune system.
References:
Bannister BA, Begg NT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
Hlinomazová Z, Loukotová V, Horáčková M, Serý O. 2010. The treatment of HSV1 ocular infections using quantitative
real-time PCR results. Acta Ophthalmol. 10: 1755-3768.
www.elisabethgroup.com | www.elisabeth.cz | www.zephyrusbsb.com
19
4. Viral Diagnostics
4.6 Varicella zoster virus
Kit
EliGene® VZV RT Kit
90040-RT
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
EliDNA VZV QRT standard
Varicella zoster virus
1 – 10 copies of viral DNA in the amplified sample
CSF, biopsy (skin, liver), swabs
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480 and NANO, fully compatible with RotorGene instruments.
CE certification:
Detected DNA region:
Detection technology:
yes
genome
TaqMan probes
Clinical study description and results:
Viral
Infections
Within the frame of testing the functional characteristics of EliGene® VZV RT kit overall 481 clinical specimens were
analyzed. From these 481 clinical specimens 51 were positive. EliGene® VZV RT kit analyzed all 51 samples as positive.
Totally 430 clinical specimens were properly by EliGene® VZV RT kit analyzed as negative
Sensitivity: 100%
Specificity: 100%
Pathogen description:
VZV is a double stranded DNA virus of the Herpes group. VZV infection can have two different manifestations of the
disease - chickenpox or shingles. The manifestations include rash, low grade fever and malaise. Varicella may rarely
result in complications including VZV encephalitis or pneumonia. VZV is certainly transmissible through the airborne
route and does not require close personal contact. The skin lesions are certainly full of infectious virus particles. After an
incubation period of 14 days, the virus arrives at its main target organ, the skin. The virus can survive in the human body
in the latent form for a long time.
References:
Bannister BA, Begg NT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
Dwyer DE, Cunningham AL. 2002. Herpes simplex and varicella-zoster virus infections. Med J Aust. 177(5):267-73.
Hlinomazová Z, Loukotová V, Horáčková M, Serý O. 2010. The treatment of HSV1 ocular infections using quantitative
real-time PCR results. Acta Ophthalmol. 10: 1755-3768.
20
elisabeth
pharmacon group
EliGene®
ZEPHYRUS
B I O S A F E T Y B OX
®
ZEPHYRUS®
ZEPHYRUS®
P C R B O X H A R M O NY
M A G N E T O
4.7 Hepatitis B virus
Kits
EliGene® HBV RT Kit
90037-RT
EliGene® HBV UNI kit
90037-UNI
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
Yes
Hepatitis B virus
0.1 IU/ml of isolated specimen
serum, plasma
Compatible instruments:
RT Kit intended for ABI 7000, 7300, 7500 (Applied Biosystems),
LightCycler® 480 and LightCycler® Nano (Roche), RotorGene 6000 or
RotorGene Q (Qiagen).
UNI Kit intended for LightCycler® 2.0
CE certification:
Detected DNA region:
Detection technology:
Yes, CE 1023
genome (core protein)
TaqMan probes (RT version), Moleular Beacons (UNI version)
Clinical study description and results:
Viral
Infections
Within the frame of testing the functional characteristics of EliGene® HBV RT Kit and EliGene® HBV UNI Kit overall 500
clinical specimens were analyzed. From these specimens, 300 HBV positive specimens and 200 HBV negative specimens
were confirmed by artus® HBV TM PCR Kit. The EliGene® HBV RT Kit and EliGene® HBV UNI Kit diagnosed as HBV positive
299 specimens. 1 specimen from Nigeria was determined by the EliGene® HBV RT Kit and EliGene® HBV UNI Kit as
negative. The sensitivity and specificity of EliGene® HBV RT Kit and the EliGene® HBV UNI Kit is 98.78% and 100%,
respectively. EliGene® HBV RT Kit and the EliGene® HBV UNI Kit shows 99,8% (499 from 500 specimens) match in
detection of HBV DNA compared to results determined by artus® HBV TM PCR Kit.
100% sensitivity can be declared for both kits for the concentration of 5 IU/ml of serum corresponding to concentration
of 0.1 IU/ml of isolated specimen by Magneto 1ml Serum/Plasma DNA/RNA isolation kit (see page 34) - extraction
volume: 1.0 ml, elution volume: 50 μl. The linear range of the EliGene® HBV RT Kit and EliGene® HBV UNI Kit has been
determined to cover concentrations from 0.1 IU/μl to at least 1 x 105 IU/μl.
Precision data of the EliGene® HBV RT Kit were calculated on basis of the Cp values of the amplification curves using the
Quantification standard 4 (10 000 IU/ml) and Internal Control (Figure 7).
Pathogen description:
Hepatitis B virus is a small DNA virus divided into Hepadnaviridae family. Eight different genotypes signed by
alphabetical letters A to H, have been previously described. Since the proposal of the four described genotypes (A–D),
four others (E–H) have been characterized during the two last decades. Recently, a ninth “genotype” evidenced in
North-West China, India, Lao and Vietnam and tentatively termed “I” was suggested, although it is still subject to debate
and as being a recombinant strain with a genotype C backbone. Finally, very recently, a tenth genotype provisionally
assigned to genotype “J” was proposed for a Japanese patient's HBV isolate.
Hepatitis B (HB) is transmitted by the exchange of body fluids e.g. blood, semen, breast milk and saliva. The virus spread
between people who have unprotected sexual intercourse, drug users who share needles and syringes and health care
workers in contact with potentially contaminated blood or body fluids.
It is estimated that one third of the world's population has serological evidence of past or present infection with HBV and
that 350 to 400 million people are still chronically infected, of whom 78 % lived in Asia, 16 % in Africa, 3 % in South
America and the remaining 3% in Europe, North America and Oceania.
HBV infection has a broad spectrum of clinical diseases, ranging from acute hepatitis (including fulminant hepatic
failure) to a low viraemic asymptomatic “inactive” carrier state or to progressive chronic hepatitis that may lead to
cirrhosis with an annual rate of 2 to 5 % in HBe-positive patients and hepatocellular carcinoma (HCC) with a cumulative
5-year incidence of 15 to 20 %. Both HBV-related end-stage liver disease and HCC are responsible for around 1 million
deaths per year.
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21
4. Viral Diagnostics
4.7 Hepatitis B virus (continued)
References:
Bannister BA, Begg NT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
Ho SKN, Yam W-C, Leung ETK, Wong L-P, Leung JKH, Lai K-N, Chan TM. 2003. Rapid quantification of hepatitis B virus
DNA by real-time PCR using fluorescent hybridization probes. J Med Microbiol. 52(5): 397-402.
Viral
Infections
Figure 6. Calculation of the linear range. The straight was determined by linear regression
of the log10 calculated concentrations with the log10 nominal concentrations. The equation
of the regression line in included in the figure.
Figure 7: Precision data on the basis of the Cp values.
Figure 8: Analysis of specimens with different concentration of HBV from 0.5 IU/ml
to 5000 IU/ml with EliGene® HBV RT kit.
A – amplification curves for FAM channel (HBV detection),
B - amplification curves for HEX channel (Internal control)
22
elisabeth
pharmacon group
EliGene®
ZEPHYRUS
®
B I O S A F E T Y B OX
ZEPHYRUS®
ZEPHYRUS®
P C R B O X H A R M O NY
M A G N E T O
4.8 Hepatitis C virus
Kit
EliGene® HCV LC Kit
90059-LC
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
Yes
Hepatitis C virus
1 – 10 viruses in the amplified sample
serum, plasma
Compatible instruments:
ABI 7000, 7300, 7500 (Applied Biosystems), LightCycler® 480 and
LightCycler® Nano, LightCycler® 2.0 (Roche), RotorGene 6000 or
RotorGene Q (Qiagen).
CE certification:
Detected DNA region:
Detection technology:
Yes, CE 1023
genome (core protein)
Molecular Beacons probes
Clinical study description and results:
The EliGene® HCV LC kit is intended for detection and quantification of hepatitis C virus RNA. Kit contains reversion
transcription in one mix with RealTime detection.
Clinical study description and results:
Viral
Infections
Within the frame of testing the functional characteristics of EliGene® HCV LC Kit overall 500 clinical specimens were
analyzed. From these specimens, 300 HCV positive specimens and 200 HCV negative specimens were verified by COBAS
AMPLICOR HCV MONITOR Test, version 2.0 (v2.0). The EliGene® HCV LC Kit diagnosed as HCV positive 299 specimens. 1
specimen was determined by the EliGene® HCV LC Kit as negative. The sensitivity and specificity of EliGene® HCV LC Kit is
98,78% and 100%, respectively. The correlation of the quantitative results of EliGene® HCV LC Kit and COBAS AMPLICOR
HCV MONITOR Test, version 2.0 (v2.0) was analysed by linear regression, when the logarithmised results of both kits
were plotted as scatterplots against each other (Figure 9). Correlation coefficient was calculated to R2=0,81.
100% sensitivity can be declared for the concentration of 50 IU/ml of serum isolated by Magneto 1ml Serum/Plasma
DNA/RNA isolation kit (see page 34) - extraction volume: 1.0 ml, elution volume: 50 μl. The linear range of the
EliGene® HCV LC Kit has been determined to cover concentrations from 50 IU/μl to at least 1 x 105 IU/ml (Figure 10).
Pathogen description:
Hepatitis C virus is enveloped RNA virus classified to Flaviviridae family. Virus genome consists of 9379 nucleotides. HCV
represents heterogeneous group of viruses divided into 6 genotypes (groups) and more than 40 sub-types. EliGene®
HCV LC Kit gives the positive results with genotypes 1a, 1b, 2a, 2b, 3a, 4a, 6 and 6b.
Today, the word prevalence about 175 milions persons infected by hepatitis C virus is assessed. Virus is disseminating
mainly by parenteral way, the sexual transfer is observed rare. In our conditions, significantly risk group is represented
by the injection drug consumers.
References:
Bannister BA, Begg NT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
www.elisabethgroup.com | www.elisabeth.cz | www.zephyrusbsb.com
23
4. Viral Diagnostics
4.8 Hepatitis C virus (continued)
Figure 9. Comparison of the COBAS AMPLICOR HCV MONITOR Test (v.2.0) against the EliGene® HCV LC Kit
Viral
Infections
Figure 10. Calculation of the linear range. The straight was determined by linear regression of the log10
calculated concentrations with the log10 nominal concentrations. The equation of the regression line
is included in the figure.
Figure 11. Analysis of specimens with different concentration of HCV from 126 IU/ml
to 21000 IU/ml with EliGene® HCV LC kit.
A – amplification curves for five HCV standards,
B - amplification curves for HCV positive specimens.
24
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5.1 Mycobaterium tuberculosis
Kits
EliGene® MTB LC Kit
90030-LC
EliGene® MTB RT Kit
90030-RT
EliGene® MTB UNI Kit
90030-UNI
Package size:
Possibility of quantification:
Analytical specificity:
50 reactions
On demand
Mycobacterium tuberculosis, Mycobacterium bovis, negative result for
Mycobacterium kansasii, Mycobacterium xenopi, Mycobacterium avium
and Mycobacterium marinum
1 – 10 copies of bacterial DNA in the amplified sample
urine, sputum, BAL, exudates, tissue, paraffin embedded tissue
Analytical sensitivity:
Specimens:
Compatible instruments:
LC Kit intended for LightCycler® 2.0, LightCycler 480 instruments
RT Kit intended for ABI7000, ABI7300, ABI7500 instruments, fully
compatible with RotorGene instruments
UNI Kit is intended for ABI7500FAST, SmartCycler, RotorGene 6000 and
for all LightCycler® instruments
CE certification:
Detected DNA region:
Detection technology:
yes
genome (insertion sequence IS6110)
Molecular Beacons (LC, UNI versions), TaqMan probes (RT version)
Clinical study description and results:
Totally 559 clinical samples were tested by EliGene® MTB RT Kit, 518 of them were blind samples with unknown status
and 41 MTB positive samples verified by cultivation from Brno region. EliGene® MTB RT Kit detected correctly all 52
positive samples. Totally 481 samples were correctly diagnosed by EliGene® MTB RT Kit as MTB negative.
Sensitivity: 100%
Specificity: 100%
Bacterial
Infections
Pathogen description:
Genus Mycobacterium comprises more than 50 species when among them obligatory pathogenic etiological agents of
tuberculosis and leprosy and on the other hand also facultative pathogenic and non-pathogenic species representing
important part of natural biotopes can be found. M. tuberculosis comprises to the one complex with genetically close
related M. bovis and it is etiological agents of human tuberculosis. M. bovis is etiological agent of cattle tuberculosis and
some others pets and wild beast and it is rarely detected in humans.
References:
Kriz P, Kralik P, Slany M, Slana I, Svobodova J, Parmova I, Barnet V, Jurek V, Pavlik I. 2011. Mycobacterium pinnipedii in a
captive Southern sea lion (Otaria flavescens): a case report. Veterinarni Medicina, 56:307-313.
Mendes A.C., Fernandes S.J., Ferreira L.C., Pereira M., Ramos H., Cabeda J.M. 2009. Evaluation of MTB Q Alert kit for
detection of Mycobacterium tuberculosis directly on patient samples
Clinical Microbiology and Infection, Suppl. 19, P1321.
Figure 12. Sensitivity test of EliGene® MTB
LC kit on LightCycler® 2.0 instrument
shows the detection of 2 DNA copies in
reaction mix.
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25
5. Bacterial Diagnostics
5.2 Borrelia
Kits
EliGene® Borrelia LC Kit
90024-LC
EliGene® Borrelia RT Kit
90024-RT
EliGene® Borrelia UNI Kit
90024-UNI
Package size:
Possibility of quantification:
Analytical specificity:
Analytical sensitivity:
Specimens:
50 reactions
On demand
Borrelia garinii, Borrelia afzelii and Borrelia burgdorferi sensu stricto
1 – 10 copies of genomic DNA in the amplified sample
blood, urine, CSF, synovial fluid, tissue
Compatible instruments:
LC Kit intended for LightCycler® 2.0, LightCycler 480 instruments
RT Kit intended for ABI7000, ABI7300, ABI7500 instruments, fully
compatible with RotorGene instruments
UNI Kit is intended for ABI7500FAST, SmartCycler, RotorGene 6000 and
for all LightCycler® instruments
CE certification:
Detected DNA region:
Detection technology:
yes
genome (fla gene)
Molecular Beacons (LC, UNI versions), TaqMan probes (RT version)
Clinical study description and results:
Totally 486 clinical samples were tested by EliGene® Borrelia RT Kit. EliGene® Borrelia RT Kit detected correctly all 9
positive samples. Totally 477 samples were correctly diagnosed by EliGene® Borrelia RT Kit as Borrelia negative.
Sensitivity: 100%
Specificity: 100%
Bacterial
Infections
Pathogen description:
Lyme borreliosis is a disease caused by spirochete Borrelia burgdorferi. The detection of this bacterium could be made
from any clinical material using the DNA diagnostics in Europaen Union. There can be found three pathogenic species of
Borrelia genus: B. b. sensu stricto, B. afzelii and B. garinii. EliGene® Borrelia Kits contain mix of primers and probes that
are specific to all three mentioned Borrelia species. DNA diagnostics is useful in cases of Borrelia suspicion (anamnesis of
tick or insect that suck the blood).
References:
Hytönen J, Hartiala P, Oksi J, Viljanen MK. 2008. Borreliosis: recent research, diagnosis, and management. Scand J
Rheumatol. 37(3):161-172.
Priem S, Rittig MG, Kamradt T, Burmester GR, Krause A. 1997. An optimized PCR leads to rapid and highly sensitive
detection of Borrelia burgdorferi in patients with Lyme borreliosis. J Clin Microbiol. 35(3): 685–690.
Figure 13. Detection of pathogenic Borrelia sp. in
DNA samples (isolated from ticks) by EliGene®
Borelia RT kit on ABI®7500FAST instrument.
26
Figure 14. Detection of pathogenic Borrelia sp. in DNA
samples (isolated from ticks) by EliGene® Borelia RT
kit on ABI®7500FAST instrument.
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5.3 Anaplasma phagocytophilum
Kit
EliGene® Anaplasma phagocytophilum UNI Kit
90061-UNI
Package size:
Analytical specificity:
Specimens:
50 reactions
Anaplasma phagocytophilum
blood, urine, CSF, synovial fluid, tissue
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480, LightCycler® Nano, Light Cycler 2.0, fully compatible with RotorGene
instruments.
CE certification:
Detection technology:
Clinical study description and results:
yes
Molecular Beacons
new kit, clinical study will be finished in 2013
Description:
Anaplasma phagocytophilum is obligate intracellular, gram-negative rickettsial organisms infecting human leukocytes.
Anaplasma is transmitted to humans by tick bites (Ixodes scapularis, Ixodes ricinus and Ixodes pacificus). It causes
human granulocytic anaplasmosis (HGA) previously known as human granulocytic ehrlichiosis (HGE). Typical symptoms
include fever, headache, chills, and muscle aches and they usually occur within 1-2 weeks of a tick bite. The
epidemiology of this infection is very much like that of Lyme disease (caused by Borrelia burgdorferi) and babesiosis
(caused primarily by Babesia microti), which all have the same tick vector.
References:
Gary P. Wormser et al. 2006. The Clinical Assessment, Treatment, and Prevention of Lyme Disease, Human Granulocytic
Anaplasmosis, and Babesiosis: Clinical Practice Guidelines by the Infectious Diseases Society of America. Clin Infect Dis.
43 (9): 1089-1134.
Johan S. Bakken, MD, Stephen Dumler. 2008. Human Granulocytic Anaplasmosis. Clin Infect Dis. 22 (3): 433–448
www.elisabethgroup.com | www.elisabeth.cz | www.zephyrusbsb.com
Bacterial
Infections
27
6. Diagnostics for Human Genetics
6.2 Lactose Intolerance
Kits
EliGene® Lactose Intolerance C-13910T LC Kit
90044-LC
EliGene® Lactose Intolerance G-22018A LC Kit
90045-LC
Package size:
50 reactions
Standards:
EliDNA Lactose Intolerance C-13910T Standard (Ref. 90044-CL.LC)
TM
EliDNA Lactose Intolerance G-22018A Standard (Ref. 90045-CL.LC)
Analytical specificity:
Specimens:
C-13910T polymorphism (EliGene® Lactose Intolerance C-13910T LC Kit)
G-22018A polymorphism (EliGene® Lactose Intolerance G-22018A LC Kit)
blood, buccal swabs
Compatible instruments:
LightCycler® 2.0, LightCycler 480, RotorGene 6000
CE certification:
Detected DNA region:
Detection technology:
yes
genome (core protein)
Molecular Beacons
TM
Clinical study description and results:
EliGene® Lactose Intolerance C-13910T LC Kit and EliGene® Lactose Intolerance G-22018A LC Kit specificity were tested
on 50 samples of human DNA with genotypes determined by DNA sequencing. The specificity of the method was
validated by searching in the DNA databases also.
Specificity: 100%
Description:
Patients presenting with symptoms of lactose intolerance are in some laboratories routinely tested for a singlenucleotide polymorphism C-13910T and G-22018A that are located upstream of the lactase gene (LCT) and are tightly
associated with genetically determined lactase persistence/non-persistence. The genotyping of these polymorphisms
enables differential diagnosis of genetic versus secondary causes of lactose intolerance. Several PCR-based methods
have been established as tests for this SNP. In particular, automated genotyping assays conducted on LightCycler
platforms provide a rapid method for routine high-throughput analysis of this variant.
Human
Genetics
References:
Ridefelt P, Håkansson LD. 2005. Lactose intolerance: lactose tolerance test versus genotyping. Scand J Gastroenterol.
40(7):822-6.
Bulhões AC, Goldani HA, Oliveira FS, Matte US, Mazzuca RB, Silveira TR. 2007. Correlation between lactose absorption
and the C/T-13910 and G/A-22018 mutations of the lactase-phlorizin hydrolase (LCT) gene in adult-type hypolactasia.
Braz J Med Biol Res. 40(11):1441-6.
Figure 15. Analysis of samples with -13910 CC and TT genotypes by EliGene®
Lactose Intolerance C-13910T LC kit on LightCycler® 2.0 instrument
28
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6.1 Coeliac disease
Kits
EliGene® Coeliac RT Kit
90048-RT
EliGene® Coeliac RT28 Kit
90048-RT28
Package size:
Analytical specificity:
50 reactions
EliGene® Coeliac RT Kit - HLA-DQ2, DQ8 and DRB1*04
EliGene® Coeliac RT28 Kit - HLA-DQ2 and DQ8
blood, buccal swab
Specimens:
Compatible instruments:
Real-Time Systems ABI 7000, 7300, 7500 (Applied Biosystems),
LightCycler® 480 and Nano (Roche), fully compatible with RotorGene
and Q- instruments.
CE certification:
Detection technology:
yes
TaqMan probes
Clinical study description and results:
EliGene® Coeliac RT kit specificity was tested on 50 samples of human DNA with genotypes determined by DNA
sequencing. The specificity of the method was validated by searching in the DNA databases also. It is necessary to use
recommended concentration of human DNA for the genotypization.
Specificity: 100%
Pathogen description:
Coeliac disease, also called coeliac sprue is one of the most common enteropathogenic disorders and is characterized by
a lifelong hypersensitivity to gluten proteins found in wheat, rye, oat and barley. Until recently, coeliac disease was
considered relatively uncommon with an estimated prevalence rate ranging from 1 in 1000 to 1:4000. However, the
availability of new serologic tests have led to the observation that coeliac disease is much more common, affecting
about 1 of 100-400 persons in Europe, the majority of patients showing little clinical symptoms. Considerable evidence
now indicates that coeliac disease has a strong genetic component. Coeliac disease is a multifactorial disorder
associated with HLA-DQ2 (DQA1*05/DQB1*02) or DQ8 (DQA1*0301/DQB1*0302) or HLA-DRB1*04. HLA DQ2 is
expressed in the majority (>90%) of those with coeliac disease and DQ8 in about 8%. The expression of these HLA-DQ2
or DQ8 molecules is necessary but not sufficient to develop coeliac disease and accounts for only about 50% of the
genetic component of the disease. Studies in sibling (sib recurrence risk for coeliac disease of 10%) and of identical twins
(concordance of 70%) suggest that the contribution of HLA genes in coeliac disease is less than 50%. About 95% of all
coeliac patients possess this particular genotype compared to approximately 20% of the normal population. From this
point of view absence of these alleles is useful in excluding coeliac disease. However, at least one other non-HLA gene
and some environmental factors are also likely to be involved in the disease. On the basis of recent results the analysis of
HLA-DQ2 (DQA1*05/DQB1*02) and DQ8 (DQA1*0301/DQB1*0302) alleles is sufficient to exclude coeliac disease.
Human
Genetics
References:
Troncone R, Jabri B. 2011. Coeliac disease and gluten sensitivity. J Intern Med. 269(6):582-90.
Trynka G, Wijmenga C, van Heel DA. 2010. A genetic perspective on coeliac disease. Trends Mol Med. 6(11):537-50.
Bourgey M. et al. 2007. HLA related genetic risk for coeliac disease. Gut 2007;56:1054–1059.
Margaritte-Jeannin P. et al. 2004. HLA-DQ relative risks for coeliac disease in European populations: a study of the
European Genetics Cluster on Coeliac Disease. Tissue Antigens 63: 562–567.
Figure 16. Tables used for evaluation of results obtained by EliGene®Coeliac RT kit (A) and EliGene®Coeliac RT28 kit (B).
www.elisabethgroup.com | www.elisabeth.cz | www.zephyrusbsb.com
29
6. Diagnostics for Human Genetics
6.3 Spondylitis HLA-B27
Kit
EliGene® Spondylitis HLA-B27 RT Kit
90060-RT
Package size:
Analytical specificity:
Specimens:
50 reactions
allele HLA-B27
blood, buccal swabs
Compatible instruments:
Real-Time Systems 7000, 7300, 7500 Applied Biosystems, LightCycler®
480, LightCycler® Nano, fully compatible with RotorGene instruments.
CE certification:
Detection technology:
Yes, CE
TaqMan probes
Clinical study description and results:
EliGene® Spondylitis HLA-B27 RT Kit specificity were tested on 100 samples of human DNA with genotypes determined
by DNA sequencing. The specificity of the method was validated by searching in the DNA databases as well.
Specificity: 100%
Description:
Human
Genetics
Human Leukocyte Antigen (HLA) B27 (subtypes B*2701-2759) is an HLA class I surface antigen that is encoded in the B
References:
locus
in the major histocompatibility complex (MHC) on the short arm of chromosome 6. In HLA-B27 antigen a strong
association with ankylosing spondylitis (AS) and spondyloarthropathies (SpA) was found. Moreover the increased
incidence of this antigen in other diseases as Reiter´s syndrome or uveitis was found. Association studies found an
association between HLA-B27 antigen and AS in all ethnic and racial groups worldwide however the prevalence of HLAB27 antigen and the strength of its association with AS does vary. For example, the prevalence of HLA-B27 antigen is
about 8% in Caucasians, 4% in North Africans, 2-9% in Chinese, and 0.1-0.5% in Japanese. Further, among northern
Europeans only 8% of the general population possesses HLA-B27, but more than 90% of the patients with AS possess
this gene. In contrast, among African Americans 2% to 4% of the general population and only 50% to 60% of patients
with AS possess this gene. From this reasons HLA-B27 test cannot be used to screen an asymptomatic population to
detect AS but the test provides a statement of increased probability of the existence of AS in the symptomatic patient.
Also, the presence of HLA-B27 antigen influences the clinical manifestations of AS disesase, because HLA-B27–positive
patients have a significantly younger age at onset of their disease and a higher prevalence of episodes of eye
inflammation (acute anterior uveitis) and hip joint involvement.
References:
Khan MA. Remarkable Polymorphism of HLA-B27: An Ongoing Saga. Curr Rheumatol Report. 2010; 12: 337-41
Robinson PC, Brown MA. 2012. The genetics of ankylosing spondylitis and axial spondyloarthritis. Rheum Dis Clin North
Am. 38(3):539-53
Thomas GP, Brown MA. Genetics and Genomics of Ankylosing Spondylitis. Immunol Rev. 2010; 233:162-180.
30
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7.1 Isolation of DNA from MTB and other bacteria
Kits
EliGene® MTB Isolation Kit
90043-50
EliGene® MTB Isolation Supplement
12200-MTB-IZO
Package size:
Specimens:
50 isolations
sputum, decontaminated sputum, cultures, BAL, exudates, urine, swabs
Intended use:
EliGene® MTB Isolation Kit is intended for the isolation of bacterial DNA
from clinical material
EliGene® MTB Isolation Supplement is intended for DNA isolation from M.
tuberculosis and eventually futher microorganisms (G+) from sputum or
cultures by using Maxwell®* 16 FFPE Tissue LEV DNA Purification Kit
(Promega).
CE certification:
Isolation technology:
yes
silica columns
Clinical study description and results:
Within the frame of testing the functional characteristics of EliGene® MTB Isolation Kit overall 559 clinical specimens
were analyzed. From these specimens, 518 blind specimens from Prague region and 41 MTB positive specimens verified
by cultivation form Brno region were analyzed. The isolation kit showed 100% sensitivity in isolation of DNA from clinical
decontaminated material which means that this procedure for isolation of MTB DNA gives in 100% sufficient amount of
MTB DNA for the next DNA diagnostic method. The effectiveness of these isolation procedures is 95,34% which means
that from total amount of isolated specimens were isolated 95,34% of specimens without inhibition.
Description:
Many problems like inhibitions or low sensitivity occur when one make the isolation of DNA from Mycobacterium
species. EliGene® MTB Isolation Kit is designed to avoid all problems with PCR inhibitors and low yields of DNA. The total
time of isolation process is about 2 hours including two incubations, each for 30 minutes. Kit contains all components
necessary for the DNA isolation including microtubes. No addition of ethanol or other chemicals is needed.
References:
Bannister BA, Begg NT, Gillespie SH. 2000. Infectious Disease. Blackwell Science, 2th Ed.
DNA/RNA
Isolation
&
Spectral
Calibration
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31
7. DNA Isolation and Spectral Clibration Kits
7.2 Isolation of total DNA (including bacterial) from clinical specimens
Kits
EliGene® Urine Isolation Kit
90051-50
EliGene® Sperm Isolation Supplement
90051-50SPO
Package size:
Specimens:
50 isolations
Intended use:
EliGene® Urine Isolation Kit is intended for the isolation of DNA from
clinical material.
EliGene® Sperm Isolation Supplement is intended for DNA isolation from
the sperm.
CE certification:
Isolation technology:
yes
columns
blood, buccal tissue samples, sperm, urine, swabs (cervical, urethral)
Clinical study description and results:
Within the frame of testing, the functional characteristics of EliGene® Chlamydia trachomatis LC kit overall 120 clinical
specimens of urine were analyzed. Before the detection by EliGene® Chlamydia trachomatis LC, DNA from all samples
was isolated by the EliGene® Urine Isolation Kit. No inhibition was observed. The effectiveness of this isolation
procedure was 100%. It means that from total amount of isolated specimens were isolated 100% of specimens without
inhibition.
Description:
Many problems like inhibitions, low sensitivity etc. occur when one make the isolation of DNA from clinical samples.
EliGene® Urine Isolation Kit is designed to avoid all problems with PCR inhibitors and low yields of DNA. The total time of
isolation process is about 1 – 2 hour including incubation. Kit contains all components necessary for the DNA isolation
including microtubes. No addition of ethanol or other chemicals is needed.
DNA/RNA
Isolation
&
Spectral
Calibration
32
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7.3 Spectral Calibration Kit
Kits
EliGene® LC Spectral Calibration Kit
90099-SC
EliGene® LC Spectral Calibration Kit 2
90099-SC2
Package size:
For one Spectral calibration run on LightCycler® 1.2 (ref: 90099-SC) and
LightCycler® 2.0 instruments (ref: 90099-SC2)
Description:
EliGene® LC Spectral Calibration Kit is intended for spectral calibration of instruments LightCycler® 1.2 and 2.0 for the
EliGene® LC kits using 530 and 705 channels for detection. EliGene® LC Spectral Calibration Kit 2 is intended for spectral
calibration of instruments LightCycler® 2.0 for the EliGene® LC kits using 530, 550 and 705 channels for detection. Before
the usage of EliGene® LC kits, it is necessary to perform spectral calibration with EliGene® LC Spectral Calibration Kit or
EliGene® LC Spectral Calibration Kit 2. Once the spectral calibration was done; it is valid until the instrument is upgraded
or optics is changed.
Kit
EliGene® LC Spectral Calibration Plate
90099-SCP
Package size:
96-well plate for Spectral calibration run on LightCycler® 480
Description:
EliGene® LC Spectral Calibration Plate is intended for spectral calibration of instruments LightCycler® 480 for the kits
EliGene® LC and UNI using 530, 560 and 705 channels for detection. Before the using of EliGene® LC kits it is necessary to
perform spectral calibration with EliGene® LC Spectral Calibration Plate. Once the spectral calibration was done, it is
valid until the instrument is upgrading or optics is changed.
DNA/RNA
Isolation
&
Spectral
Calibration
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33
8. Instruments ZEPHYRUS
8.1 ZEPHYRUS® Magneto (CE IVD )
ZEPHYRUS®
M A G N E T O
ZEPHYRUS® Magneto represents
high quality sample preparation
instrument based on proven
m a g n et i c p a r t i c l e s e p a rat i o n
technology. It utilizes many years
experience in automated nucleic acid
isolation and in combination with
EliGene CE IVD kits it provides optimal
solution for sensitive, accurate and
cost effective diagnostic applications.
Reagents are delivered by automated
dispensing system instead of using
expensive prefilled cartridges. The
total hand on time is minimized with
processing time about 50 minutes for
1 or up to 12 samples per batch.
ZEPHYRUS® Magneto use metal rods for magnetic separation that are magnetised by an external magnet.
To enable the often complicated re-suspension step of once separated magnetic particle pellets e.g. in wash
or elution buffers, the magnet can be switched off. Re-suspension is then realized by rotation of the rods
guaranteeing the efficient, complete and smooth re-suspension of the bead pellet. Due to this solution this
step becomes quick and thorough, resulting in isolation products with both high yields and purities
respectively.
Parameters:
DNA/RNA
Isolation
&
Spectral
Calibration
Instruments
ZEPHYRUS®
34
1 – 12 samples per run
sample sizes up to 1000 µl
Magneto kits and protocols for a huge variety of sample materials (Body Fluid, Blood, Tissue,
Serum/Plasma, Swabs)
certified according to Annex 3 of the EU IVD Directive 98/79/EC
integrated buffer dispensing revolutionary resuspension technology
standard plastic devices
touch panel software
USB slot for data transfer
Ref.
Name / description
Z-MAG-12
ZEPHYRUS® Magneto
Available isolation kits for ZPEHYRUS® Magneto
Ref.
Name / description
EZ-7001-50
M-7001-180
Magneto BodyFluid DNA/RNA isolation kit / for DNA/RNA isolation from 200
microliters of blood, serum, plasma, buccal swabs, tissues, sperm, urine, synovial
fluid, CSF etc.
EZ-7001-50
M-7002-180
Magneto 1ml Serum/Plasma DNA/RNA isolation kit / for DNA/RNA isolation
from HIV, HCV, HBV and other viruses from 1000 microliters of serum or plasma
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8.2 ZEPHYRUS® PCR Box Harmony (PCR workstation)
400.0µm
First PCR workstation with
ZEPHYRUS®
P C R B O X H A R M O NY
nano-filtration
ZEPHYRUS® PCR Box Harmony (PCR workstation) brings several advantages:
Greater mobility of the arms in the workzone; the samples are better protected against DNA contamination; automatic
and programmable UVC light decontamination; replaceable plexiglass box at the end of its life; unique filtration of air
by nanofilter, security sensors for turn off the UVC light when you open the cover; enough space for pipettes and plastic
in the PCR workstation etc.
Equipment of individual variants
BASICsize
External
ECO size
External
ACTIVEsize
External
Ref.:
x Z-PCR-600
x Z-PCR-600-E
x Z-PCR-600-A
Upper part with buttons and electronics
x
x
x
x
x
x
x
x
x
x
x
x
x
x
x
x
x
x
Replaceable plexiglass part (580 x 580 x 520 mm)
1
Manual start of immediate decontamination
Manual start of scheduled decontamination
2
3
Automatically started scheduled decontamination
Electronics for power-saving operation
4
Electronically monitored function of UV lamps
Active decontamination
x
5
x
6
Special air nano-filtration
x
x
7,8
x
-
x
x
-
x
x
OPTION
Stainless base
USB interface for export of operation log to PC
9
Accessories
x
x
x
x
x
x
Notes:
Ref.
Description
Z-PCR-601
ZEPHYRUS® Plexiglass
Z-PCR-602
ZEPHYRUS® Stainless base
Z-PCR-603
ZEPHYRUS® UVC lamp 15 W
Z-PCR-604
ZEPHYRUS® Y light 15 W
Z-PCR-605
ZEPHYRUS® W Light 15 W
Z-PCR-606
ZEPHYRUS® UVC AD
Z-PCR-607
ZEPHYRUS® Ventilator AD
Z-PCR-608
ZEPHYRUS® Nanofilters AD (10 pcs)
Z-PCR-609
ZEPHYRUS® Nanofilters AD (50 pcs)
Z-PCR-610
ZEPHYRUS® USB interface
1) UV decontamination starts immediately after a longer press of the UV button.
2) Scheduled UV decontamination in preset time starts after a short press of the UV
button.
3) Scheduled UV decontamination starts automatically in preset time after closing
cover.
4) Usage of special electronics helps to decrease power consumption by more than
30 % in comparison with the basic version. Furthermore, the service life of the lamp is
extended.
5) UV lamp malfunction can be detected by the electronics, information of
malfunction is showed to the operator.
6) Active decontamination of air in the workplace is achieved by a germicide UV lamp
placed in a closed decontamination tunnel. Thus decontamination can be running
even while working inside the PCR Box. Air circulation through decontamination
tunnel is provided by two ventilators.
7) At the output of the decontamination tunnel there is a mounted filter made of
specially designed nanomaterial, which is capable of catching DNA fragments longer
than 100bp, bacteria, viruses and all particles larger than 25nm.
8) A variant available only in connection with active decontamination.
9) Logged operation data can be exported via USB cable into PC. It is necessary to
install a USB interface driver and communication SW, which can organize received
data for example into an excel table or create regular decontamination protocol.
www.elisabethgroup.com | www.elisabeth.cz | www.zephyrusbsb.com
Instruments
ZEPHYRUS®
35
8. Instruments ZEPHYRUS
8.3 ZEPHYRUS® Bio Safety Box
ZEPHYRUS®
B I O S A F E T Y B OX
Biohazard box
new generation...
Class II Type A2
Bio safety cabinet 30% / 70%
Zephyrus® bio safety box (Class II Type A2) can be
either exhausted back into the room or connected
to a facility exhaust air duct system. The cabinet's
airflow is 30% exhausted / 70% recirculated to
minimize cross-contamination of low to moderate
risk biological materials in the absence of volatile
toxic chemicals.
Standard Features

Germicidal UV light
Touch screen
Automatic correction of airflow
Air pressure control on both HEPA filters
Emblem diagnostic signalization
One eletrical outlet and gas valve
Economical Features
Common steel body of cabinet
Box without emblem diagnostics signalization
Optional Features and Accessories

Remote control of cabinet's performance - remote service support
Control of light intensity
Stainless steel stand
Simple carbon steel stand
Different size of box on request available
Up to 4 eletrical outlets
Up to 4 gas valves
LCD screen in work zone
Technical specifications:
Instruments
ZEPHYRUS®
Style of cabinet:
Cabinet construction:
Eletrical:
Viewing window:
Standard inflow velocity:
Bench top / base stand
Pressure tight welded stainless steel 304
230 V / 50 Hz, 10 A
Fully closed to fully open 480 mm
0,5 m/s
Ref.
External size (mm)
Work zone (mm)
Weight
Z-BSB-120
1207 x 1530 x 855*
920 x 660 x 620
175 kg
Z-BSB-150
1507 x 1530 x 855*
1220 x 660 x 620
210 kg
Z-BSB-180
1807 x 1530 x 855*
1520 x 660 x 620
245 kg
* Transport through 800mm doors possible after front cover removal.
36
elisabeth
pharmacon group
EliGene®
Manufacturer:
ELISABETH PHARMACON, spol. s r.o.
Namesti Svobody 18, 602 00 Brno
phone: +420 542 213 851
e-mail: [email protected]
www.elisabeth.cz
®
ZEPHYRUS®
ZEPHYRUS®
ZEPHYRUS
B I O S A F E T Y B OX
P C R B O X H A R M O NY
M A G N E T O
Distributor: