1. science
2. chemistry

serum digoxinthat allows the complete automation of this
analysis. Up to now, the application of the EMIT method to
automated instruments, including the Roche Cobas analyzers, necessitates a preliminary manual step consisting of
pre-treatment
of the samples with the sodium hydroxide
reagent. In the proposed protocol, every step of the analysis
is taken in charge by the instrument, including the pretreatment step. The reagents are reconstituted exactly as in
the original EMIT procedure, except for the buffer, which is
prepared by diluting the concentrate to 160 mL instead of
225 mL; this higher concentration is necessary to counteract
the dilution effect introduced by the no-default water addition used in the system for flushing the delivery needles
after addition of the start reagent and the sample. There is
no need to prepare, by dilution, a working B reagent, and
this ensures better reagent stability. The reaction conditions
were kept as close as possible to the ones of the original EMIT
protocol. The instrumental conditions are as follows:
Sample: 20 4, water 7 4
Incubation: 5 s
Start reagent 1 (pre-treatment reagent): 8 4, water 5
Incubation: 300 s
Reagent (working A): 125 4
Incubation: 900 s
Start reagent 2 (B reagent): 5 4, water 38 4
Incubation:5 s, Wait time: 5 8
Readings: first 120 s, interval180 s, number 10
The method shows very good inter-run
6.6% for a 1.6 ng/mL concentration,
4
precision (CV of
seven single determina-
tions). When various digoxin-free sera were supplemented
with known amounts of digoxin and analyzed with the new
protocol and the results correlated with the true values (the
amount added), the following regression equation was obtained: y = 1.07x
0.061, r = 0.99, n = 28, where the y
values are the measured ones. The Cobas FARA values are
significantly higher than the true values (standard error on
the slope, 0.041) and this difference is of unknown origin.
When patients’ samples submitted for digoxin determination were analyzed simultaneously by various methods and
the results compared with those obtained with the Cobas
FARA, the following lines of regression were obtained:
-
Reference methods
manual (Syva)
FPI (Abbott)
Enzymun (BMC)
EMIT
Equation
y
y
y
=
=
=
1.08x
1.09x
1.09x
-
+
+
0.104
0.179
0.102
r
n
0.96
0.87
0.86
35
39
28
Major advantages of this new protocol are improved
precision, reduced reagent consumption (#{188}o
that in the
original procedure), and simplicity of operation.
Microcomputer Program for Internal Quality Control
with Use of Westgard’s Multi-Rule Shewhart Chart,
Mario Pandin (Laboratorio di analisi chimico-climche,
Ospedale Generale Provinciale, U. S. S. L. n. 19 del
“Mediobrenta,” 35013 Cittadella (PD), Italy)
As reported (1), I developed a microcomputer program,
written in BASIC, for internal quality control in a laboratory
that is not equipped with a main computer with a built-in
automatic quality-control facility.
This new version of that program presents new features
and more flexibility.
The hardware consists of an HP-86 microcomputer with a
built-in user memory of 128K bytes and with the option of a
12-in. monitor; two disc-drives HP-9130A, storing 270K
bytes on one disc (5#{188}
in.); a bidirectional thermal printer
HP-2671A; a graphic plotter HP-7470A with two built-in
pen stalls for two-color plotting (all from Hewlett-Packard
Co., Personal Computer Div., Corvallis, OR 97330).
In addition to the previous features, the software program, which uses two disc-drives, now has these characteristics:
1. The use of the Westgard algorithm (2) as a criterion
used in deciding whether the values indicate the analytical
run is in or out of control.
2. Display on the screen and printing of error messagesof
the control rule that causes the rejection of the run.
3. Simultaneous error messages storage on floppy-disc,
with the indication if the error is probably a systematic
error or a casual error.
4. Cumulative reports of error messages, selected by
periods (day, month, year) and by single or all tests.
5. Possibilityto modify any parameter used in the quality-control program.
6. A new graphic presentation: the cusum chart in addition to Shewhart plot and Youden plot already available.
7. Visualization on the Youden plot, using two keys, the
normal and pathological value of the control serum and the
date, for an easy inspection of the plot.
8. The program is not copyrighted, so the user may
modify or add other features as needed.
This program is available, preferably on floppy disc, upon
request from the author or from the Editorial Office of this
journal.
References
1. Pandin M. Computer program for quality control, especially
suited for the small laboratory. Clin Chem 1985;31:166-7.
2. Westgard JO, Barry FL, Hunt MR. A multi-rule Shewhart chart
for quality control in clinical chemistry. Clin Chem 1981;27:493501.
Reference Intervals for Alkaline Phosphatase Activity
Determined by the IFCC and AACC Reference Methods,
Norbert W. Tietz and Denise F. Shuey (University
Kentucky, College of Medicine, Department of
Pathology, Lexington, KY 40536-0084)
of
We determined the reference interval for alkaline phosphatase (EC 3.1.3.1) activity in serum for populations between 20 and 50 years old and above 60 years old. Enzyme
activity was determined at 30 and 37#{176}C
by the method as
recommended by the International
Federation of Clinical
Chemistry (IFCC) (1) and the American Association for
Clinical Chemistry
(AACC) (2). We used two spectrophotometers (Model 25 and Acta CIII; Beckman Instruments,
Inc., Fullerton, CA 92634). The individuals between 20 and
50 years of age were selected from apparently healthy
hospital employees; those older than 60 years were part of a
group of healthy volunteers who participated
in an extensive study of reference ranges for individuals above 60 years
of age (3). All participants studied had fasted for at least
10 hand had sat for 20-30 mm before the blood collection, to
CLINICALCHEMISTRY, Vol. 32, No. 8, 1986 1593
the effect of hemoconcentration on alkaline phosphatase activity. All assays were performed within 6 h after
collection.
Table 1 lists the total ranges observed, the central 95th
percentile reference intervals (4), and the mean results for
each group and assay temperature, The distribution
of
values is shown in histogram form in Figure 1.
minimize
Table 1. Observed
Values for Alkaline Phosphatase
ActivIty in Serum
Activity, U/L
Temp.,
Sex/age(yr)
2/20-50
t3120-50
Reference
‘c
n
Range
Interval’
Mean
30
37
79
79
24-84
35-104
28-78
42-98
50
67
30
90
90
74
31-98
46-132
36-124
38-94
53-128
40-111
64
85
63
74
48
48
50-162
37-89
46-122
53-141
43-88
56-119
82
62
81
37
9/60
30
37
30
f/60
37
‘Central 95 percentIles.
-Soy
F,20-50y
30
20
l0
-v
20
60
30
M, 2O-5Oy
30
1
20
30
70
110
20
I
lUl
#{149}
60
Influence of Sample Processing on Urine Phosphorus
Results wIth the Ektachem#{174}
400 and 700 Analyzers,
Felix G. Soloni, Roberto Bandin, and Kip Amazon (Dept.
of Pathol. and Lab. Med., Mount Sinai Medical Center,
.4300 Alton Rd., Miami Beach, FL 33140)
Many laboratories are using Ektachem analyzers (Eastman Kodak, Rochester, NY) to determine phosphorus in
urine after dilution
of the urine sample. Since there currently is no written procedure available for urine phosphorus
determination in the Ektachem 400 and 700 analyzers, and
consideringthe viscosity and density differences of serum
and urine, we decided to dilute urine samples 10-fold with a
serum control of known phosphorus concentration, instead of
water. We then assayed all samples as if they were serum,
with the instruments calibrated for serum phosphorus, and
calculated results as follows:
P
I
I
100
=
1OR
9C
-
where P is the urine phosphorus content, R is the result
obtained from the analyzer, and C is the phosphorus content
of the serum control used to dilute the urine samples and
also assayed in the same run with them (all units mg/dL).
The phosphorus results we reported for the CAP urine
survey were consistently higher than those from other
10
i
48.
2. Enzyme working group of the Subcommittee on Standards,
AACC, Study Groupon Alkaline Phoaphatase. A reference method
for measurement of alkaline phosphatase activity in human serum.
Clin Chem 198329:751-61.
3. Tietz NW, Wekstein DR. Shuey DF, Brauer GA. A two-year
longitudinal reference range study for selected serum enzymes in a
population more than 60 years of age. J Am Geriatr Soc
1984;32:563-70.
4. Solberg HE. Establishmentand use of referencevalues.In: Tietz
NW, ed.Textbookofclinical chemistry, Philadelphia: WB Saunders
Co., 1986:371-4.
‘M,20-50y
20
10
References
1. Tietz NW, Rinker AD, Shaw LM. IFCC methods for the measurement of catalytic concentration of enzymes.Part 5. IFCC method
for alkaline phosphatase. J Clin Chem Clin Biochem1983;21:731-
URINE PI-IOSPHORUS
30
70
110
F,? 60y
i
o diluted serum
S
0’;
#{149}
,
20
60 I0#{149}0’
30
U
70
U
a
110 150
V
-z
20
20
M,
M1?60y
r=099
n=21
?60y
a
Lu
.
IC
10
O.826x+2.433
=0.99
n =21
IIUU
30
70 110
FIg. 1. HIstograms indicating the distributionof alkalinephosphatase
values(In U/L) observed in men (M) and women(F) in populationsat
20
60
ages 20-50 and >60 years
Leftxiumn shows values at 30 ‘C; nght cokimn shows values at 37 ‘C
1594 CLINICALCHEMISTRY, Vol. 32, No. 8, 1986
theoreticol,
mgML