Enzyme immunoassay of free thyroxin in serum.

1984 ◽  
Vol 30 (10) ◽  
pp. 1682-1685 ◽  
Author(s):  
M Ito ◽  
K Miyai ◽  
K Doi ◽  
H Mizuta ◽  
N Amino
Keyword(s):  
Pregnant Women ◽  
Enzyme Immunoassay ◽  
Equilibrium Dialysis ◽  
Normal Adult ◽  
Double Antibody ◽  
Low Concentrations ◽  
The Mean ◽  
Measurable Range

Abstract We describe a double-antibody enzyme immunoassay for free thyroxin (FT4) in serum with use of beta-D-galactosidase conjugated to thyroxin. The method is uninfluenced by thyroxin-binding globulin or albumin. Values for FT4 so determined correlated well with those determined by radioimmunoassay (r = 0.98) and equilibrium dialysis (r = 0.89). The mean variability (CV) within and between assays was 7.4% and 7.6%, respectively. The measurable range of FT4 in serum was 2.8 to 109 ng/L. The FT4 concentrations in serum as determined by this method were 8.4 to 15.5 ng/L for 26 normal adult subjects; 26 to greater than 109 ng/L for 10 patients with hyperthyroidism; less than 2.8 to 8.0 ng/L for seven patients with hypothyroidism; 7.3 to 15.8 ng/L for eight pregnant women; and 12.2 and 13.5 ng/L for two patients with low concentrations of thyroxin-binding globulin.

Enzyme immunoassay of free triiodothyronine in serum.

1987 ◽  
Vol 33 (1) ◽  
pp. 172-176 ◽  
Author(s):  
N Hata ◽  
K Miyai ◽  
Y Endo ◽  
Y Iijima ◽  
Y Doi ◽  
...  
Keyword(s):  
Pregnant Women ◽  
Enzyme Immunoassay ◽  
Solid Phase ◽  
Equilibrium Dialysis ◽  
Reference Interval ◽  
Normal Range ◽  
Free Triiodothyronine ◽  
Normal Limits ◽  
The Mean ◽  
Measurable Range

Abstract In this new solid-phase antibody enzyme immunoassay for free triiodothyronine (FT3) in serum, beta-D-galactosidase conjugated to triiodothyronine is used. Results are uninfluenced by physiological concentrations of thyroxin-binding globulin or albumin. Results correlate well with those determined by equilibrium dialysis (r = 0.95). The mean CVs within and between assays were 6.1 and 9.5%, respectively. The measurable range of FT3 in serum is 0.7 to 26 ng/L; the normal reference interval is 1.9 to 8.9 ng/L. Concentrations of FT3 in serum of patients with hyperthyroidism were high; those of patients with hypothyroidism were within normal limits or low, and those of patients with congenitally decreased or increased TBG were within the normal range. In normal pregnant women, concentrations of FT3 as determined by radioimmunoassay correlated with those of albumin, declining as pregnancy progressed, but FT3 values determined by the proposed method or equilibrium dialysis were within the normal range and did not change during pregnancy.

Enzyme immunoassay of free thyroxin in dried blood samples on filter paper.

1985 ◽  
Vol 31 (5) ◽  
pp. 750-753 ◽  
Author(s):  
N Hata ◽  
K Miyai ◽  
M Ito ◽  
Y Endo ◽  
Y Iijimi ◽  
...  
Keyword(s):  
Enzyme Immunoassay ◽  
Filter Paper ◽  
Room Temperature ◽  
Normal Subjects ◽  
Blood Samples ◽  
Double Antibody ◽  
Coefficients Of Variation ◽  
The Mean ◽  
Measurable Range

Abstract We describe a double-antibody enzyme immunoassay for determination of free thyroxin (FT4) in dried blood samples on filter paper, with use of a T4-beta-D-galactosidase complex. The measurable range of FT4 concentration in two 3-mm blood discs, each of which contained about 2.7 microL of blood, was 1.9 to 93 ng/L, as determined by comparison with concentrations of FT4 in known serum standards. FT4 in blood samples dried on filter paper was stable for at least four weeks when kept dry at -20 degrees C, room temperature, or 37 degrees C. The mean coefficients of variation were 7.6% (within assay) and 6.4% (between assays). Results for FT4 by this method correlated well with those for serum determined by radioimmunoassay (r = 0.98). The proposed method can be used to differentiate persons with hyper- and hypothyroidism from normal subjects and those with abnormal concentrations of thyroxin-binding globulin. The procedure seems suited for screening studies.

Enzyme immunoassay of thyroxin-binding globulin.

1982 ◽  
Vol 28 (12) ◽  
pp. 2408-2411 ◽  
Author(s):  
K Miyai ◽  
M Ito ◽  
N Hata
Keyword(s):  
Pregnant Women ◽  
Enzyme Immunoassay ◽  
Normal Subjects ◽  
Sample Volume ◽  
Coefficients Of Variation ◽  
Analytical Recovery ◽  
The Mean ◽  
Free Serum ◽  
Test Serum ◽  
Measurable Range

Abstract We describe a new double-antibody enzyme radioimmunoassay for thyroxin-binding globulin (TBG) in serum. A TBG-beta-D-galactosidase complex was prepared with m-maleimidobenzoyl N-hydroxysuccinimide ester. The mean analytical recovery of TBG added to serum was 99%, sample volume and TBG value were linearly related, and values for TBG determined by this method and those determined by radioimmunoassay correlated well (r = 0.98). Coefficients of variation averaged 7.6% within assay, 6.6% between assay. The measurable range of TBG in the serum was 3.3 to 52 mg/L, but higher TBG concentrations could be measured by diluting the test serum with TBG-free serum. Mean (and SD) serum TBG concentrations as determined by this method were as follows: for 20 normal subjects, 21.2 (3.7) mg/L; for 10 pregnant women, 50.1 (9.5 mg/L; and for 10 patients with TBG deficiency, not detectable.

Biotin radioligand assay with an 125I-labeled biotin derivative, avidin, and avidin double-antibody reagents.

1987 ◽  
Vol 33 (11) ◽  
pp. 1983-1988 ◽  
Author(s):  
E Livaniou ◽  
G P Evangelatos ◽  
D S Ithakissios
Keyword(s):  
Pregnant Women ◽  
Binding Protein ◽  
Biological Fluids ◽  
Normal Subjects ◽  
Chronic Hemodialysis ◽  
Serum Samples ◽  
Radioligand Assay ◽  
Double Antibody ◽  
Low Concentrations ◽  
High Concentrations

Abstract We describe a new radioligand assay for determining biotin in biological fluids by using a mixture of N-[beta-(4-OH-3-125I-phenyl)ethyl]- and N-[beta-(4-OH-3,5-di-125I-phenyl)ethyl]biotinamides as radiotracer, avidin as a binding protein, and an avidin double-antibody as a separation reagent. The radiotracer is synthesized by coupling (at pH 8.5, 20-22 degrees C, 90 min) N-hydroxysuccinimidobiotin to radioiodinated tyramine. The assay curve is linear and the assay itself is sensitive (less than 10 ng/L), reproducible (intra- and interassay CVs 4.1% and 7.0%, respectively), and allows the simultaneous handling of more than 100 samples in less than 4 h. Serum samples from apparently normal subjects contained 100-840 ng of biotin per liter (mean 340 ng/L). Pregnant women had low concentrations of biotin (100-300 ng/L) in their serum. Patients undergoing chronic hemodialysis treatment showed high concentrations (0.5-3.0 micrograms/L), which may be ascribable to the inability of avidin, which was used as the assay binding protein, to distinguish biotin from biotinyl derivatives with an intact ureido ring.

PROTEIN C/PROTEIN S IN THE FOETAL BLOOD. ABSENCE OF BOUND PROTEINS AND C4 BINDING PROTEIN

1987 ◽  
Author(s):  
E Melissari ◽  
M F Scully ◽  
C Parker ◽  
K H Nicolaides ◽  
V V Kakkar
Keyword(s):  
Pregnant Women ◽  
Protein C ◽  
Binding Protein ◽  
Protein S ◽  
First Trimester ◽  
Normal Adult ◽  
Blood Protein ◽  
Free Protein ◽  
The Mean ◽  
Foetal Blood

Protein C, free and bound protein S and C4 binding protein levels (C4bp), were measured by electroimmunoassay in 7 pregnant women aged 22-29 years at 16-18 weeks of gestation, immediately prior to termination of pregnancy for social reasons. Protein C and protein S levels were also measured in their foetuses from blood taken through the umbilical cord. In this group of pregnant women the mean levels for protein C were 104% of normal adult mean (range 80-128%), for C4bp 100% (52-150%), for free protein S 66% (43-89%). In the foetuses the mean value for protein C was 15.3% (10.5-21%) and for free protein S 36.85% (27-47%) of the normal adult mean. Bound protein S and C4bp levels were zero. Conclusions: (1) free protein S is significantly decreased (< 2SD below the normal adult mean) in women after the first trimester of gestation whereas no change is seen in protein C concentration; (2) C4bp levels are at zero in the foetus as also are the levels of bound protein S; (3) foetal blood protein S level is approximately 2.5 times higher than protein C. Since all other vitamin K-dependent factors have been observed to be in the range of 10-20% of normal at this stage of gestation, our findings may be further proof of a non hepatic (endothelial) source of plasma protein S.

Double-antibody enzyme immunoassay applied to human alpha 1-fetoprotein.

1976 ◽  
Vol 22 (2) ◽  
pp. 198-204 ◽  
Author(s):  
L Belanger ◽  
D Hamel ◽  
D Dufour ◽  
M Pouliot
Keyword(s):  
Pregnant Women ◽  
Enzyme Immunoassay ◽  
Dose Response ◽  
Present Method ◽  
Response Curve ◽  
Dose Response Curve ◽  
Clinical Samples ◽  
Double Antibody ◽  
Coefficients Of Variation ◽  
Analytical Recovery

Abstract We report the development of a double-antibody system for enzyme immunoassay of human alpha 1-fetoprotein. Equilibrium and sequential saturation procedures were evaluated and compared with a similar double-antibody radioimmunoassay. With our method, 3 mug of alpha 1-fetoprotein per liter can be detected. The dose-response curve covers a 100-fold range of analyte concentrations. Within-run and between-run coefficients of variation are respectively 6.9% and 10.0%. Analytical recovery of added antigen is 99.5 +/- 10.6%. Results are routinely obtained in 24 h but can be obtained in less than 8 h. Alpha 1-fetoprotein was measured in a variety of clinical samples, including sera from 488 pregnant women. The advantages of the present method over radioimmunoassay and other enzyme immunoassay procedures are discussed.

Effect of stripping thyroxin from thyroxin-binding globulin on the measurement of free thyroxin in serum by equilibrium dialysis and by radioimmunoassay

1990 ◽  
Vol 36 (2) ◽  
pp. 313-318 ◽  
Author(s):  
T Nakagawa ◽  
K Matsumura ◽  
K Takeda ◽  
N Shinoda ◽  
A Matsuda ◽  
...  
Keyword(s):  
Pregnant Women ◽  
Equilibrium Dialysis ◽  
Normal Subjects ◽  
Sample Volume ◽  
Dilution Factor ◽  
Free T4 ◽  
Mean Values ◽  
Patient Groups ◽  
The Mean ◽  
Influence Measurement

Abstract In considering factors that might influence measurement of free thyroxin (T4), we evaluated the proportion (%) of T4 that could be stripped from thyroxin-binding globulin (TBG). The percentage of free T4 was measured in serially diluted sera from four normal subjects, four patients with hyperthyroidism or hypothyroidism, four pregnant women, and four malnourished subjects with low TBG. The critical percentage of stripping was determined by the product of the percentage free T4 and the critical dilution factor (the point where the percentage free T4 began to decrease). The mean values obtained for the respective patient groups--6.38%, 2.76%, 16.73%, 9.75%, and 4.28%--were proved to be related to the rate of saturation of TBG with T4. Values for percentage stripping determined with the "GammaCoat two-step RIA" and the "LiquiSol RIA" were well within the critical percentage stripping by equilibrium dialysis, except in the case of low-TBG serum as measured by LiquiSol RIA. Free T4 concentration as measured by LiquiSol RIA decreased as sample volume decreased. These findings were ascribed to the relatively high values for percentage stripping in the LiquiSol RIA, which led to erroneously low values for free T4.

A two-step radioimmunoassay for free triiodothyronine in serum.

1987 ◽  
Vol 33 (3) ◽  
pp. 372-376 ◽  
Author(s):  
M G Rajan ◽  
A M Samuel
Keyword(s):  
Pregnant Women ◽  
Solid Phase ◽  
Equilibrium Dialysis ◽  
Normal Subjects ◽  
Free Triiodothyronine ◽  
High Affinity ◽  
The Mean ◽  
Hyperthyroid Patients ◽  
Normal Adults ◽  
Good Agreement

Abstract Using a high-affinity solid-phase-bound antibody (Ka = 1.2 X 10(11) L/mol), we have standardized a two-step radioimmunoassay for free triiodothyronine (FT3) in serum, based on immunoextraction. The method was validated by comparison with an equilibrium-dialysis procedure (r = 0.96) involving RIA of T3 in the dialysate standardized with the same antibody and by a commercial (Liso-Phase, International-CIS) method. The two-step RIA could detect as little as 0.2 pg per milliliter. The mean CVs within and between assays were 9% and 12%, respectively. FT3 values measured in 30 normal adults ranged from 1.77 to 4.77 ng/L. Comparison with ratios of total T3 to thyroxin-binding globulin showed good agreement in normal subjects, pregnant women, and hypothyroid and hyperthyroid patients.

Autoantibodies to Thrombomodulin: Development of an Enzyme Immunoassay and a Survey of their Frequency in Patients with the Lupus Anticoagulant

1992 ◽  
Vol 67 (05) ◽  
pp. 507-509 ◽  
Author(s):  
John Gibson ◽  
Margaret Nelson ◽  
Ross Brown ◽  
Hatem Salem ◽  
Harry Kronenberg
Keyword(s):  
Enzyme Immunoassay ◽  
Lupus Anticoagulant ◽  
Specific Binding ◽  
Technical Problem ◽  
Serum Samples ◽  
Citrate Buffer ◽  
The Mean ◽  
Sample Dilution ◽  
Negative Population ◽  
Serum Components

SummaryIn order to investigate the possibility that autoantibodies to thrombomodulin (TM) may exist in patients with the lupus anticoagulant (LA) and perhaps be implicated in the pathogenesis of recurrent thrombosis seen in such patients, we developed an enzyme-immunoassay to screen serum samples for anti-human TM activity. The major technical problem encountered in developing this assay was to reduce the non-specific binding of serum components from both the LA positive and the negative population. Considerable reduction of non-specific binding was achieved by use of a phosphate/citrate buffer at pH 8.0 and the use of an optimal sample dilution of 1/40. In addition, samples were always tested in parallel in blank wells and results are expressed as an OD ratio. Samples from 113 patients with the LA were assayed and compared to 78 patients referred for LA testing but found to be negative. The mean OD values for the LA positive patients (± SD) was 1.36 (0.44) with a range of 0.78-2.57. This was virtually identical to the values for the LA negative population (1.38 ± 0.40, range 0.76-2.77). The results of this study indicate that there is no evidence for the presence of a significant autoantibody activity to TM in patients with the LA when compared to LA negative patients. If such autoantibodies do exist their frequency must be quite low.

HAPTEN-RADIOIMMUNOASSAY OF PLASMA OESTRADIOL

1973 ◽  
Vol 72 (2) ◽  
pp. 330-344 ◽  
Author(s):  
Peter Doerr
Keyword(s):  
Adult Male ◽  
Normal Values ◽  
Normal Adult ◽  
Adult Males ◽  
Reagent Blank ◽  
Keyhole Limpet Haemocyanin ◽  
Ether Extraction ◽  
The Mean ◽  
Assay Precision ◽  
Plasma Oestradiol

ABSTRACT A hapten-radioimmunoassay for plasma oestradiol is described and information about the reliability of the method is given in detail. Oestradiol-3-hemisuccinate coupled to keyhole limpet haemocyanin is used for immunization of rabbits. The antiserum utilized for the assay is characterized by its titer, affinity and specificity. Following ether extraction and NaOH-light petroleum partition oestradiol is separated from crossreacting oestrogens by TLC. Oxidation of oestradiol on the plate is prevented by mercaptoethanol. To separate free and antibody bound ligand 250 μg dextran-coated charcoal per tube is used in the presence of bovine serum gammaglobulin (1 mg/ml). The between-assay precision based on 15 different determinations of control samples from normal adult male plasma was 9.4% (C. V.). The mean reagent blank value of 31 determinations was equivalent to 0.3 pg oestradiol and the detection limit in terms of the 99% confidence limit for a single blank value, was equivalent to 4.3 pg oestradiol. A procedure for detecting plasma blanks is described. Plasma oestradiol is separated from approximately all concomitant substances originally present in the sample by enzymatic conversion into oestrone and a second TLC. No plasma blanks could be detected with respect to normal adult male plasma. Normal values for adult males based on 51 subjects were characterized by a median of 17.2 pg/ml and the 95 percentiles of 9.5–27.6.

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