Download New Ultrafiltration Method for Free Thyroxin Compared with

CLIN. CHEM. 36/5, 800-804
(1990)
New Ultrafiltration Method for Free Thyroxin Compared
with Thyroid Dysfunction and Nonthyroidal
Illness
SarI
Dialysis
This new ultrafiltration
method
for free thyroxin in serum
(FT4(U)] is based on radioimmunoassay
of the free hormone
fraction in ultrafiltrates
obtained by centrifuging
serum sampIes in Unisep’
Ultracent-1 0 ultrafiltration
devices. We compared the results obtained
with those by an equilibrium
dialysis method [FT4(D)].
In 36 euthyroid
healthy subjects,
the mean FT4(U)
concentration
was 24.2 pmol/L and the
mean FT4(D) concentration
14.8 pmol/L. In hyperthyroid
and
hypothyroid
patients,
results by the ultrafiltration
method
were also approximately
twice as high as those obtained by
the dialysis method. In 23 patients with various nonthyroidal
illnesses,
mean FT4(U) was 41.2 pmoVL and mean FT4(D)
19.8 pmol/L. The mean FT4(U)/FT4(D)
ratio in patients with
nonthyroidal
illnesses (1.97) was not significantly higher than
in control subjects
(1.68), making it unlikely that the increase
in serum FT4 is caused by weakly protein-bound
and therefore dialyzable
inhibitors of thyroxin binding to carrier proteins. However,
two nonthyroidally
ill patients with a clearly
increased
FT4(U) but a normal FT4(D) concentration
might
have had such inhibitors, whereas
for two other nonthyroidaily ill patients
a high molar ratio of free fatty acids to
albumin is a more likely explanation
for increased FT4(U) and
FT4(D) concentrations.
On theoretical
grounds, we consider
the FT4(U) concentrations
analytically
more nearly accurate
than FT4(D) values for all patient groups studied.
Keyphrases:
variation, source
radioimmunoassay
of
When
one measures
the free (unbound)
concentration
of
an analyte
in serum,
the analytical
conditions
must
not
alter the balance
between
the bound
and the unbound
fraction
of the analyte.
In measuring
free thyroxin
(Fr4) in
serum,
methods
in which the free fraction
is first separated
from the serum
carrier
proteins
by equilibrium
dialysis
or
ultrafiltration
and thereafter
quantified
are therefore
theoretically
sound.5 In assays of FF4 in serum where physiological T4-carrier
proteins
or anti-P4
autoantibodies
are
present along with reagent
anti-T4
antibody,
as in one-step
thyroxin-analog
tracer
assays, accurate
determination
is
possible only if the radioactive
tracer is not significantly
associated
with the endogenous
binders. To date, no such
analog
tracer
has been synthesized;
therefore,
the present
generation
of analog-based
assays of FF,5 give spurious
results, to a variable
extent, in subjects with quantitati
or qualitative
abnormalities
of T4-binding
proteins (1-6)
Two-step
rapid radioiinmunoassays
of VF4 are based o
immunoextraction
of the free fraction
by immobilized
antiantibody
in the first step and back-titration
of unoccupi
hormone-binding
sites in the second step (2, 6, 7). Th
assays, although
less prone than the one-step analog assays
errors induced by altered T4-protein
binding
in serum, still
not fulfill the requirement
that the free hormone
equilibri
must not be changed, because the extracted
hormone
fractio
much exceeds
the true free hormone
fraction.
F’1 assa
involving
the use of T4-immunoglobulin-acridinium
ester
T4-enzyme
conjugates
(8-11)
offer several advantages
ov
radioimmunoassays,
but are also potentially
prone to erro
caused by protein-binding
of conjugates,
particularly
bin
to T4-autoantibodies
(10).
Our objective in the present study was to measure
by
new ultrafiltration
method the Fr4 concentration
[VF4(U
in sera from patients with various
forms of thyroid
dysftm
tion and nonthyroidal
illness (NT!),
and to compare
th
results
with
those obtained
by a well-validated
equilibrium
dialysis
method
[FF4(D)]
(2). We also pa
formed a correlation
analysis between
the concentrations
o
VF4 and free fatty acids (FFA) in sera from NT! patien
FFA have been suggested by some investigators
to functio
as inhibitors
of T4-protein
binding
in NT! (4, 12, 13
although
others have not been able to provide evidence fo
such an effect (14, 15). If, however,
the increase
in Fr
observed in some NT! patients
is attributable
mainly
dialyzable
inhibitors
of T4-protein
binding,
one can expe
a discrepancy
between
the Fr4 concentrations
determine
by equilibrium
dialysis and ultrafiltration.
I
-
MaterIals
and Methods
Subjects.
The control
group consisted of 36 euthyroic
healthy
subjects, 18 men and 18 women (ages 19-87 years
mean 44.5). We also studied 20 women with hyperthyroidisir
(ages 19-79 years,
mean 43.7) and 18 patients with hypothyroidism,
three men and 15 women (ages 26-73 years, mear
53.8). Their
diagnoses
were based on clinical
findings
and
standard
laboratory
tests as reported
previously
in detail (16)
concentrations
ofT4 (RIA), triiodothyronine
(RIA), and thy.
rotropin
(immunoradiometric
assay) in serum,
and a trilodot
hyronine-uptake
test for calculation
of an Fr4 index. Jr
addition
we studied
23 patients, 12 men and 11 women (ages
17-90 years, mean 53.6), with various
severe NTIs: diabeth
ketoacidosis
(n = 8), cancer
(n
8), heart disease
(n = 4).
bacterial
infection
(n = 2), or anorexia
nervosa (n = 1). Nc
patient was given heparin
or other drugs known
to influenc
the concentration
of VF4 in serum.
Ultrafiltration
method.
The essential
features
of thE
method are as follows:
because
of the known
importance
ol
pH for the Fr4 concentration,
even within the physiological
pH range, one must adjust the serum pH to 7.4 by adding
50 L of 1 molJL 4-(2-hydroxyethyl)-1-piperazineethane
sulfonic
acid (HEPES) buffer to 1 mL of serum,
then incubatE
the sample for 20 mm at 37 #{176}C
to achieve equilibrium
ol
=
1
Kuopio University
2Minerva
Foundation
Central
Hospital,
Institute,
Kuopio;
Helsinki;
3Department
of Clinical
Chemistry,
University
of Helsinki,
Helsinki,
Finland.
4Address correspondence
to this author at: Medix Biochemica
Oy, Asematie
13, 02700 Kauniainen,
Finland.
6Nonstandard
abbreviations:
T4, thyroxin;
FF4, free thyroxin;
FF4(U), free thyroxin
by ultrafiltration
assay; Fr4(D), free thyroxin by equilibrium
dialysis assay; FFA, free fatty acids; and NT!,
nonthyroidal
illness.
Received November
27, 1989; accepted March 1, 1990.
800
CLINICAL
in Patients
and B. Krlstlan LiewendahIaS
H. Tlkanoja”24
AddItIonal
with Equilibrium
CHEMISTRY,
Vol. 36, No. 5, 1990
inding
at this temperature.
Wash the Unisep”
UltracentLO ultrafiltration
devices
(Bio-Rad
Laboratories,
Richnond, CA) by passing 1.5 mL (maximal
volume allowed) of
50 mmoIJL
phosphate
buffer,
pH 7.4, and 1.5 mL of
listilled
water through
the membrane.
[We selected this
Liltrafiltration
device
for use in our assay after testing
ilbumin
leakage
in four different
devices,
as reported
lsewhere
(17).] Apply 1-1.5 mL of serum for ultrafiltration
i.r 30 mm at 37#{176}Cand 2000
x g (fixed-angle
rotor),
liscarding
the ultrafiltrate
formed
during
the first 5 mm.
Afterwards,
without
delay, analyze the ultrafiltrate for T4.
To quantify the Fr4, we use 100 pL of ultraflltrate
in a
adioimmunoassay
involving
sheep anti-T4 antiserum
(Inernational
Laboratory
Services,
London, U.K.), in a final
lilution
of 1:1.5 x iO’, and [1I]T4
(specific
activity
4000
i/g;
Cambridge
Medical
Diagnostics,
Billerica,
MA). An;iserum
and tracer
are diluted
in 50 mmoIJL
phosphate
)uffer
(pH 7.4) containing
2 g of gelatin
and 200 mg of
odium
azide per liter. The tubes are incubated
at 4#{176}C
)vermght
before separation
of free and bound radioactivity
y adding 1 mL of 250 g/L polyethylene
glycol reagent
:Carbowax
6000; Fluka,
Buchs, Switzerland)
and 100 pL of
15 g/L bovine gamma-globulin
solution
(Sigma Chemical
jo., St. Louis, MO). The lowest concentration
of standard
ve include
in the assay is 6.6 pmol/L;
the highest,
211
molJL.
We correct the measured
concentration
of FT4 for a
10.3% (average)
adsorption
of T4 to the ultrafiltration
nembrane.
We determined
this adsorption
value by adding
lIT4 tracer to pooled ultrafiltrates.
The within-assay
coefficient of variation
(CV) for assay of
‘F4 was 10.9% and the between-assay
CV 9.7%, as calcu.ated from duplicate
determinations
of an Fr4 concentraion of 27 pmoLIL.
Other
methods.
We measured
serum Fr4 by an equilibium dialysis-based
method
as described
earlier
(2). The
220
concentration
of total nonesterified
FFA was determined
enzymatically
(NEFA
C-Test;
Wako
Chemicals,
Tokyo,
Japan)
in sera from the NT! patients
and controls.
The
bromcresol
purple method (Orion Ltd., Espoo, Finland)
was
used to quantify
albumin
in sera from NT! patients.
Statistics.
Group differences
were tested for significance
by the nonparametric
Wilcoxon’s
rank sum test. Correlation coefficients
were calculated
by Spearman’s
method,
and linear-regression
analysis was performed
by the leastsquares method.
Results
Concentrations
of Fr4 in serum
measured
by ultrafiltration and equilibrium
dialysis
methods
in patients
with
thyroidal
dysfunction
or NT! are summarized
in Table 1.
The distribution
of Fr4(U)
concentrations
in the various
groups of patients is presented
in Figure 1. Figure
2 shows
the correlation
between
the FT4(U)
and FT4(D) concentrations for the subjects studied.
Values
measured
by the ultrafiltration
method
were
higher than by equilibrium
dialysis
(Table 1). The mean
FF4(U)
in the euthyroid
control group was 24.2 (SD 6.9)
pmol/L,
whereas in NT! patients
the mean value was 41.2
(SD 31.5) pmoIJL.
The corresponding
values obtained
by
equilibrium
dialysis were 14.8 (SD 3.5) and 19.8 (SD 5.3)
pmol/L,
respectively.
The mean ratio of FT4(U) to Fr4(D)
in
the group of control
subjects
was 1.68 (SD 0.41) and in the
NT! group 1.97 (SD 1.00). The diagnoses
[and FF4(U)/
Fr4(D)
ratios] in the four NT! patients
with the highest
Fr4(U)
values were as follows: bacterial
endocarditis
and
heart failure
(2.3), severe heart failure
caused by persistent
truncus
arteriosus
(5.1), metastatic
melanoma
(3.3), and
metastatic
ovarian
cancer
(4.1). The FT4(U)IF’I’4(D)
ratios
for hypothyroid
and hyperthyroid
patients
were 2.16 (SD
0.83) and 1.91 (SD 0.50), respectively.
There was no overlapping
of Fr4(U)
or Fr4(D)
results
in
the euthyroid
healthy
and hyperthyroid
groups,
whereas
several of the hypothyroid
patients
with mild disease
had
concentrations
in the euthyroid
range.
The mean FFA concentrations
were 0.94 and 0.59 mmol/
200#{149}
300
180
0
,
#{163}
leo
250
200
140
f
6.
120
6
150
100
2#{149}
80
too
V
II’.
80
60
40
‘
20
A
25
(C)
75
F14101
8
C
0
ig. 1. Concentrations
of FT4 in serum
as measured by the
iltrafiltration
technique in euthyroid healthy subjects (A), and in
atlents with nonthyroidal illnesses (,
hyperthyroidism
(C), or
ypothyroidism
80
125
100
l5O
lp.,,oILl
Correlation
between
FT4 concentrations
as measured
by
ultrafiltration [FT4(U)] and by equilibrium dialysis [FT4(D)) in subjects
with various thyroid states: euthyroid healthy (A), euthyroid sick (Lx),
hyperthyroid (#{149}),
or hypothyroid (0)
The overall regression equation for all the sublects studied is y 1 .78x + 1.72
Fig. 2.
=
(n
=
97,
r
=
0.93)
CLINICAL
CHEMISTRY,
Vol. 36, No. 5, 1990
801
Table
1. ConcentratIons
of FT,8 In Serum
as Measured by Ultrafiltration
Several Groups of Subjects
M.an (and rang.),
P
<0.01,
36
20
18
23
8P <0.001, as compared
24.2
(14-48)
l38.O
(61-265)
14.8a (6.9-31)
41.2a (21-138)
groups,
respectively.
In the NT!
group, the correlation
coefficient for FFA vs Fr4(D)
was not
significant
(r = 0.27), nor was that for FFA vs FT4UJ) (r =
0.07). When the FF4 concentrations
in NT! patients
were
correlated
with the FFA to albumin
molar
ratio,
we obtained
the following
FFA/albumin
vs FF4(D), r = 0.59 (P
<0.01);
FFA/albumin
vs Fr4(U),
r = 0.39 (P <0.05).
Discussion
Comparison
of the FT4 concentrations
in serum reported
by investigators
who used either equilibrium
dialysis
(2,
18-22) or ultrafiltration
(23-29) reveals that ultrafiltration
methods
usually
gave higher
estimates,
although
to a
variable
extent;
with the ultrafiltration
and equilibrium
dialysis
methods
used by Surks
et al. (30), the dialysis
method yielded the higher Fr4 estimates.
In our euthyroid
control subjects, the Fr4 concentration
determined
by ultrafiltration
was 64% higher
than that measured
by equilibrium
dialysis.
In the various
groups
of patients
we
studied, the relative
differences
between
the FF4 concentrations
determined
by ultrafiltration
and dialysis
were
even greater
(84-108%),
but for no patient group was the
mean Fr4(U)/FF4(D)
ratio significantly
higher than that
for the control
group. The finding that Fr4(D)
concentrations are lower than corresponding
Fr4(U)
concentrations
is explained,
at least partly,
by our observation
that a
10-fold dilution
of normal
serum by buffer caused a decrease of about 30% in the measured
Fr4(U)
concentration,
whereas
further
dilution
had no additional
effect (unpublished data). Interestingly,
a similar
effect of dilution
on
Fr4(D) has been reported
by some earlier
investigators
(see
references
cited in 22). This phenomenon
may be due to a
decreased
affinity of carrier proteins for T4, attributable
to
the components
of the buffers, or may indicate the existence
of physiological
inhibitors
of T4-protein
binding
in serum.
According
to a hypothesis
by Chopra et al. (12, 31), an
increased
concentration
of FFA, particularly
oleic acid, is
responsible
for the increase
in the Fr4 concentration
in
NT!.
Observations
in our laboratory
provide
further
evidence for the validity
of this hypothesis,
because
NT!
patients with high ratios for unsaturated
FFAialbumin
had
a supranormal
Fr4 concentration
measured
(13, 32). In
vitro studies
conducted
by others also have demonstrated
the necessity
of a very high FFA/albumin
ratio before the
concentration
of Fr4 serum can be expected
to increase
(14,
33,34).
According to our experience,
only in patients
with
a total FFA/albumin
ratio >2 can the Fr4 concentration
be
expected
to exceed the mean + 2 SD limit of the control
population
(13). Our conclusion
that FFA-induced
increases
of serum
FT4 are infrequent
in NT! patients is
supported
somewhat
by a recent
report
in which no NT!
patient studied had an FFA/albumin
ratio (>1.7) that could
CLINICAL
CHEMISTRY,
(D) Methods
ii
FT4D)
14.8
(8-23)
75.1#{176}(29.-135)
7.8#{176}(3-14)
l9.8t(l534)
FT4(U)/FT4(D)
1.68 (0.9-2.5)
1.91 (1.1-3.3)
2.16
(1.0-3.8)
1.97(1.1-5.1)
with the euthyroidhealthy group.
L in the NT! and control
802
Dialysis
pmol/L
FT4(U)
n
Euthyroid
Hyperthyroid
Hypothyroid
NTI
(U) and Equilibrium
Vol. 36, No. 5, 1990
be expected
to increase the Fr4 concentration
(35).
In the NT! group, we observed
a stronger
associatioi
between
serum
FFA/albumin
ratio and Fr4 as measured
b:
dialysis
than with
Fr4 by the ultrafiltration
method.
I
seems unlikely
that
FFA are significantly diluted
durin
dialysis, because they are tightly bound to albumin
and t
thyroxin-binding
globulin
(36). Elsewhere
(37) we hay
observed some generation
of FFA from triglycerides
durin
overnight
dialysis
of serum at 37 #{176}C,
the FFA concentratioi
being somewhat
higher after than before dialysis.
For th
time being,
we conclude
that the stronger
correlatioi
between
FFA/albumin
ratio and Fr4(D)
than betweei
FFAIalbumin
ratio and Fr4(U)
is possibly due to this small
in vitro and therefore
artifactual
increase
in the FFA
concentration
during
dialysis.
Notably,
Wang et al. (38)
observed that heparin
therapy
caused a larger increase in
the Fr4 fraction
when measured
by equilibrium
dialysis
than when measured
by ultraffitration,
again probably
because during dialysis there is more time for liberation
FFA, thereby
displacing
T4 from proteins,
than
during
ultrafiltration.
If, in NT!, unsaturated
FFA displace T4 from its protein
binding sites, one would expect total T4 to correlate
negatively with total FFA or with the unsaturated
FFAJalbumm ratios, in particular.
However,
neither
we (32) nor
Haynes
et al. (36) have observed
such correlations,
although
Csako et al. (4) did. The absence of an inverse
correlation
between
T4 and FFA does not exclude
the
possibility
that in NT! the increase
in serum Fr4 might be
induced
by the displacement
of T4 by unsaturated
FFA: the
correlation
between FFA and total T4 may be obscured by
other factors affecting
the total T4 concentration,
e.g., the
marked
decrease
in the concentrations
of the T4-carrier
proteins
(13, 15).
According
to observations
by Nelson
and Weiss (22),
dilution
of sera from NT! patients
results in a progressive
fall in the FF4 concentration,
a phenomenon
not found in
sera from healthy
subjects.
In our equilibrium
dialysis
system, the final serum dilution
is 1:55, which could result
in a spuriously
low concentration
of Fr4 in NT! sera if
inhibitors
of T4-protein
binding are diluted.
The risk for a
dilution
effect on serum
Fr4 is particularly
apparent
in
sera from patients
taking
drugs such as phenytoin
(39),
salicylate
(40), or furosemide
(41), which are known
to
displace
T4 from its protein binding
sites. Indeed,
several
of
the earlier
studies
involving
NTI patients
cannot
be considered to have adequately
excluded
the possibility
that the
increase
in serum Fr4 was attributable
to the drugs given
the patients.
One risks reaching
erroneous
conclusions
concerning
the mechanism
of the increased concentrations
of Fr4 in serum
when patients
treated
in intensive-care
units
are studied,
because
these patients
often receive
drugs that interfere
with thyroid-hormone
binding
to se-
ofi
um proteins
or with the peripheral
metabolism
of thyroid
ormones.
NT! sera could also contain unknown
endogenous
dialyzLble inhibitors.
However,
our results
provide
no clear
vidence
for the existence
of as-yet-unidentified
dialyzable
ithibitors
of T4-protein
binding
in NT! sera, the Fr4(U)/
T4(D) ratio in NT! patients being not significantly
higher
han in the control
subjects. This does not exclude
the
ossibility
that sera from the four NT! patients
with the
dghest Fr4(U)
concentrations
might contain such inhibiors, because
the three highest Fr4(U)/FI’4(D)
ratios
were
Lctually observed in this group of cases. However,
two of
hese NT! patients
also had high (>3) FFA/albumin
ratios,
hich
could have caused the observed
increase
in the
T4(U) and FT4(D) concentrations.
On the other hand, the
wo other NT! patients had low (<1.5) FFAialbumin
ratios;
herefore,
their
increased
Fr4(U)
concentrations
might
tave been caused by dialyzable
inhibitors,
especially
given
hat their Fr4(D)
concentrations
were normal.
Obviously,
ne must account for the possibility
that changes in Fr4
oncentrations
in NT! patients
can be caused by both FFA
nd dialyzable
inhibitors.
Interestingly,
Mendel and Cavaieri (42), according
to a preliminary
communication,
were
Lnable to demonstrate
the presence of binding inhibitors
on
he basis of expected
changes in FF4 fractions
in mixtures
f NT! and normal pool sera; their normal poo1 had a low
oncentration
of triglyceride,
to minimize
the possible in
itro generation
of FFA.
In conclusion,
the ultrafiltration
method we developed
ives a significantly higher and possibly more nearly accuate estimate
of Fr4 in serum than does the equilibrium
ialysis method
in healthy
subjects and in patients
with
hyroid dysfunction
and nonthyroidal
illnesses.
The ultraItration
method, being more practicable
and significantly
ss time consuming
than the equilibrium
dialysis method,
an therefore
be used in clinical service laboratories
with
adioimmunoassay
experience
and appropriate
equipment.
We are indebted to Drs. Margaretha
Turula, Matti VAlim#{228}ki,
nd Gustav WAgar for providing
us with sera from patients with
iyroidal and nonthyroidal
diseases. This study was supported by
rants from the Finnish
Medical Society (Finska LakaresAllskaet) and the Finnish
Cultural
Foundation,
to which
we are
rateful.
eferences
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on the assay
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).
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(1990)
Therapeutic
Monitoring of Cyclosporine:
Impact of a Change in Standards on
1251-Monoclonal RIA Performance
in Comparison with Liquid Chromatography
Paul A. Keown,12 Jane Glenn,1 Jorge Denegri,1
Ursula Maclejewska,’
David Seccombe,’
Marilyn Stawecki,4
David
Calvin Stiller,4 Christopher
Shackleton,3
Eugene
Cameron,2
This study examines
the measurement
of cyclosporine
(CsA)
by 1251-monoclonal
AlA, and describes
the impact of the recent
change
in the standard curve provided.
GsA concentrations in
serum and whole-blood
control samples measured
by 1251-RIA
were initially 8-18% higher than those by H PLC. During the first
two months of 1989, a significant and sustained deviation in the
125i..RIA produced results that exceeded
the HPLC results by
21-28%
(P <0.001).
Introduction of the new standard curve in
March 1989 returned
the concentration
of the whole-blood
controls to the previous
range (11-12%
above
HPLC,
P
<0.001).
Measurement
of clinical samples from heart, liver, and
bone-marrow
graft recipients by 1251-RIA by both old and new
kit standards produced a close linear correlation (y = 0.89 x
19.02; r
0.99; n = 75, range
40-850 ig/L),
with use of the
new standards yielding results 82 (SD 8)% of those with the
preceding
assay. However, even with the new standard curve,
CsA concentrations
by 1251-RIA in the clinical
samples
exceeded those by HPLC by a factor of 1.37 (SD 0.18) to 1.52 (SD
0.19). Segregation
for transplant type did not affect the RIA’
HPLC ratio. The results suggest cross-reactivity
of the 1251-RIA
with material present in vivo.
-
=
Additional
Keyphrases:
British
Room
Columbia
19, 855 West
control
materials
variation,
source of
Transplant
Society,
Heather Pavilion,
D-10,
12th Avenue,
Vancouver,
British
Columbia,
V5Z 1M9, Canada;
and the Departments
of ‘Pathology,
2Methcine,
and
Surgery, Vancouver
General Hospital and the
University
of British Columbia, Vancouver, and the Departments
of 4Medicine
and 5Pharmacology,
University
of Western Ontario, London, Ontario,
Canada.
Received December
11, 1989; accepted
March 9, 1990.
804
CLINICAL
CHEMISTRY,
Vol. 36, No. 5, 1990
Freeman,4’5
and G. Phillips2
One of the most important
developments
in the thera.
peutic monitoring
of cyclosporine
(CsA) is the recent intro
duction
of a monoclonal
antibody
that is selective
for thE
parent
CsA molecule
(1).6 This
provides
the theoretica]
advantages
of consistency
and specificity
in a rapid radio.
immunoassay
(RIA),
which
correlates
reasonably
closel3
with measurement
by HPLC.
Two commercial
RIA kits currently
use this antibody
with either
a 3H-labeled
tracer
and charcoal
separation
(Sandinmiune-SP;
Sandoz Ltd., Basel, Switzerland),
or 1251.
labeled
tracer with double-antibody
precipitation
(CYCLO.
Trac-SP;
INCStar
Ltd., Stillwater,
MN). Because
gamma
counting
offers advantages
of availability
and speed, ani
avoids
the need for quench
correction
with whole-blooc
samples,
the latter
RIA has become
the predominani
method
for therapeutic
monitoring
of CsA in Canada.
Both
kits have
comparable
operating
characteristics
although
the ‘251-RIA
appears
to record
CsA concentra
tions greater
than those measured
by the 3H-RIA
or
HPLC
(2-6).
This difference
in concentrations
has beer
attributed
to an inaccuracy
in the standard
curve, and th
manufacturers
have recently
changed
the standards
sup
plied to bring the results
for these techniques
into close
alignment.
Here, we chronicle the impact of this change
or
the concentrations
of CsA measured
in both control
anc
clinical
samples.
6Nonsdard
abbreviations:
CsA, cyclosporine;
VGH, Vancou
General Hospital; SDI, standard deviation
index; and UHL
University
Hospital,
London.
7Canadian
Working Group on Cyclosponine
Monitoring.
Unpub
lished data.
yen