Clinical Chemistry Siemens Point of Care - Urinalysis
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Clinical Chemistry 45: 520-525, 1999;
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via ISI Web of Science (17)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Christofides, N. D.
Right arrow Articles by Beckett, G. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Christofides, N. D.
Right arrow Articles by Beckett, G. J.
Related Collections
Right arrow Automation and Analytical Techniques
(Clinical Chemistry. 1999;45:520-525.)
© 1999 American Association for Clinical Chemistry, Inc.

Articles

Assessment of Serum Thyroxine Binding Capacity-dependent Biases in Free Thyroxine Assays

Nicos D. Christofides1,a, Ewan Wilkinson, Mary Stoddart, David C. Ray and Geoff J. Beckett

1 Research and Development, Ortho-Clinical Diagnostics, Cardiff Laboratories, Whitchurch, Forest Farm Estate, Cardiff CF4 7YT, Wales, UK.
a Author for correspondence. Fax 44 (0) 1222 526635; e-mail NCHRISTOFIDES{at}compuserve.com


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Background: Free thyroxine (FT4) assays may exhibit biases that are related to serum T4 binding capacity (sBC). We describe two tests that can be used to assess the presence and magnitude of sBC-dependent biases in FT4 assays.

Methods: We used a direct equilibrium dialysis FT4 assay as the reference method and compared the results obtained with those of the FT4 assays under investigation, in patient sera having a wide range of sBC. We then compared the expected and observed FT4 results for sera diluted with an inert buffer. Because serum dilution causes a predictable decrease in sBC, an increasingly negative bias on progressive dilution is indicative of a sBC-dependent bias.

Results: The automated FT4 assay investigated (Vitros FT4) showed no demonstrable sBC-dependent bias by either test.

Conclusion: These two tests can be used to screen for sBC-dependent biases in FT4 assays.© 1999 American Association for Clinical Chemistry


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
Irrespective of the thyroid function test strategy used, a measure of thyroid hormones is required to confirm the diagnosis of thyroid disease. The most appropriate measure of thyroid hormone is the moiety of thyroxine (T4),1 which is not bound by the thyroid hormone-binding proteins, and it is this free form of T4 (FT4) that is best correlated with thyroid status (1)(2)(3). Because the proportion of total T4 found in the free form is extremely small (~0.02% in euthyroid subjects), its quantification in the presence of the relatively very high concentrations of T4 bound to the binding proteins has proved exceedingly difficult. Indeed, an extensive literature exists highlighting interferences experienced with many commercially available FT4 assays (4)(5)(6)(7)(8)(9)(10). More recently, FT4 assays have become automated, which makes their use even more attractive to laboratories and has fueled the move from total to free thyroid hormone measurement. Although the newer FT4 assays are substantially better than the original assays, few seem to be as analytically accurate as the direct equilibrium dialysis (ED) FT4 method (9). Several studies have highlighted weaknesses in assay designs that produce significant biases (7)(9)(11)(12), the magnitude of which are assay specific and related to the concentration of the protein-bound T4 (PBT4). In particular, Nelson et al. (9) have shown that as the concentration of PBT4 decreased, the magnitude of the negative bias seen was increased. Careful examination of the experimental design adopted and data presented by Nelson et al. (9) suggested that the method-specific biases observed were dependent not only on the concentration of PBT4, but also on the serum binding capacity (sBC; calculated as the concentration x the affinity of free binding proteins). In the present study, we examine the ability of two simple experimental tests in identifying the presence and magnitude of sBC-dependent biases in an automated FT4 assay (Vitros FT4). Both of these tests challenge the FT4 assays in the clinical situations outlined recently by the National Academy of Clinical Biochemistry (13), i.e., determination of the assay-specific biases in sera having a broad spectrum of binding protein abnormalities. For the first test, we used sera from patients having a wide sBC range. To ensure that the widest possible range of sBC was challenged, the test panel included sera from pregnant women in their third trimester, ambulatory subjects, and severely ill hospitalized patients because these groups previously have been shown to have high, normal, and low sBC, respectively. For the second test, variation of sBC was achieved by the serial dilution of third trimester sera in an inert buffer.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
The patient panel examined in the assays described below consisted of 26 ambulatory subjects, 18 women at the third trimester of pregnancy, and 25 severely ill patients who were admitted to an intensive care unit with a variety of illnesses, including sepsis, cardiac arrest, cardiac failure, and respiratory failure. All procedures used were in accordance with the Helsinki Declaration of 1975 (as revised in 1996). The patient sera were collected for routine analyses and were kept frozen (-20 °C) until required for the hormone measurements performed as part of the present study. All assays (with the exception of the ED FT4, which was performed in the Cardiff laboratories of Ortho-Clinical Diagnostics) were carried out at the Clinical Biochemistry Laboratory of the Royal Infirmary, Edinburgh.

The sBC and PBT4 concentrations of three sera were reduced in vitro by serially diluting (2- to 64-fold dilutions) sera in 10 mmol/L HEPES (Sigma Chemical; cat. no. H3375) buffer solution, pH 7.4. The sera chosen were from third-trimester pregnancies because these represented sera with high sBC and PBT4 concentrations. The undiluted and diluted samples were assayed in the two FT4 methods using standard protocols.

The automated FT4 method used was the Vitros Immunodiagnostic Products (Ortho-Clinical Diagnostics, Amersham UK). The manual direct ED method was the Nichols FT4 method (Nichols Institute Diagnostics). The physicochemical principles of the assays used are fundamentally different. The Vitros FT4 assay is a labeled antibody method (2)(14), and the Nichols assay is a direct equilibrium method (15).

The ED method is carried out by dialyzing the FT4 from 200 µL of serum into 2.4 mL of dialysis buffer (at 37 °C, over a 16- to 18-h incubation period). The FT4 in the protein-free dialysate is then quantified by a sensitive solid-phase RIA for T4. To minimize variability (e.g., assay-to-assay variation), all samples studied were analyzed on one occasion, with the samples randomized. The package insert quotes an intraassay CV at doses falling within and above the euthyroid range as being <13%. The euthyroid FT4 range, which was the observed range with one outlier deleted from each end, quoted in the package insert is 10.3–34.7 pmol/L.

In the Vitros FT4 assay, 25 µL of sample is pipetted into microwells coated with a triiodothyronine (T3)-protein conjugate, followed by 100 µL of a sheep anti-T4 antibody labeled with horse-radish peroxidase in a 150 mmol/L phosphate buffer containing 1.0 g/L bovine gelatin and 1.0 g/L bovine {gamma}-globulin. During the 16-min incubation at 37 °C, a proportion of the labeled antibody binds to the serum FT4 and to the well surface, with the amount binding to the well surface being inversely related to the serum FT4 concentration. The well is then washed, and a signal reagent that produces luminescence is then added; the resulting light emitted is measured in a luminometer. All procedures are carried out automatically by the Vitros ECi Immunodiagnostic system. The samples were assayed in a batch mode, with quality-control samples at three concentrations run at the beginning and end of the assay. The within-run imprecision for FT4 values in the euthyroid and hyperthyroid range is quoted in the package insert as being <3%. The assay was calibrated using an in-house ED assay. The euthyroid range (1 and 99 percentiles) quoted in the package insert is 10–28.2 pmol/L.

In addition to the FT4 assays, the patient sera were also analyzed in the Vitros Immunodiagnostic Products Total T4 (TT4) and T3 uptake (T3U) assays. The euthyroid T4 range (2.5 and 97.5 percentiles) quoted in the package insert is 71.2–141 nmol/L. The T3U assay is calibrated in %T3U units, which are inversely related to the serum binding capacity (i.e., a serum with a high %T3U has a low binding capacity). The euthyroid %T3U range (2.5 and 97.5 percentiles) quoted in the package insert is 23.5–40.6% uptake.

All assays were performed following the manufacturers' instructions.

statistics
Analysis of data was carried out by standard methods with Microsoft Excel spreadsheets. The dependence of the FT4 bias observed with any variable was determined by correlating (linear regression analysis performed by the least-squares method) the methodological difference (from ED FT4) of each patient (y-axis) against the variable studied (x-axis). The sBC was calculated by dividing the Vitros TT4 concentration by the ED FT4 concentration (9). The concentration of PBT4 was estimated by subtracting the concentration of FT4, as measured by ED, from the concentration of TT4 (9). Because the FT4 concentration relative to TT4 was small (<0.1% of the TT4), the PBT4 concentration showed little difference from the TT4 concentration. The agreement between the Vitros FT4 and the manual ED FT4 was tested by Deming regression (16). A measure of the sBC was also obtained by the T3U, where the %T3U is inversely related to the binding capacity. The Student unpaired t-test was used to compare the biochemical profiles between the patient groups (within each assay format), and the paired t-test was used when the comparison was across different assay methods.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
The quality-control sera used in each assay were all within expected ranges, and there was no evidence of drift in performance in any of the assays examined.

The relationship between the Vitros FT4 and ED FT4 assays in all patients studied is shown in Fig. 1 . A significant correlation (r = 0.96; P <0.001) between the two assays was observed. The equation describing the relationship was:



View larger version (13K):
[in this window]
[in a new window]
  		Figure 1. Relationship between the Vitros FT4 and ED FT4 in ambulatory ({diamondsuit}), hospitalized ({blacksquare}), and pregnant ({triangleup}) subjects.

Vitros FT4 = 0.85 (± 0.03; P <0.001) ED FT4 + 3.67 (± 0.61; P <0.001).

Good agreement between the FT4 methods was achieved in all patient groups.

The relationship between the Vitros FT4 to ED FT4 in (a) the ambulatory and pregnant subjects (groups combined, n = 44), and (b) the hospitalized patients (n = 25) is described by the following equations:

(a) Vitros FT4 = 0.9 (± 0.09) ED FT4 + 3.09 (± 1.19); r = 0.83;P <0.001

(b) Vitros FT4 = 0.89 (± 0.07) ED FT4 + 2.33 (±1.9); r = 0.94; P <0.001

Table 1 shows the mean (± SD) and observed ranges for the two FT4 methods under investigation (FT4 in pmol/L), the TT4 (in nmol/L), %T3U, and sBC (in nmol/pmol) in the ambulatory, pregnant, and hospitalized patients. The FT4 concentration, as measured by both FT4 methods, in the hospitalized group was significantly (P <0.001) higher than the FT4 concentration in the ambulatory group. None of the patients in the hospitalized group had FT4 concentrations below the corresponding euthyroid range of the two FT4 assays examined. The calculated sBC in the hospitalized group was significantly lower (P <0.001) than that found in the ambulatory group, whereas the corresponding %T3U value was significantly higher (P <0.001). Thus, both measures of serum T4 binding support the view that hospitalized patients have decreased serum T4 binding capacities. The TT4 concentration in the hospitalized group was significantly lower (P <0.001) than that obtained in the ambulatory group, and 12 of the 25 hospitalized patients had TT4 concentrations below the euthyroid range. The FT4 concentration in the pregnancy group was significantly lower (P <0.001) than in the ambulatory group with both FT4 methods, whereas the sBC and TT4 concentrations were increased (P <0.001). As expected, the %T3U was significantly (P <0.001) reduced.


View this table:
[in this window]
[in a new window]
  		Table 1. Measured and calculated values in the three patient groups studied.

The FT4 concentrations obtained in the pregnancy group by the Vitros method were higher than the corresponding concentration obtained with the ED method. These differences, although small (mean difference, 2.6 pmol/L; range of difference in 95% confidence intervals, 2.1–3.2 pmol/L), were statistically significant (P <0.001). Similar differences from ED FT4 were seen in the ambulatory group (mean difference, 1.6 pmol/L; range of differences in 95% confidence intervals, 0.7 to 2.6 pmol/L; both P <0.001). The Vitros FT4 concentration in the hospitalized group was not significantly different (P >0.05) from the mean concentration obtained in the ED method.

Fig. 2 depicts the relationship between the Vitros FT4 bias and sBC in all the patient samples. The regression equations derived in each individual category of subjects are shown below:



View larger version (11K):
[in this window]
[in a new window]
  		Figure 2. Relationship between the Vitros FT4 bias (determined by ED FT4) and sBC in ambulatory ({diamondsuit}), hospitalized ({blacksquare}), and pregnant ({triangleup}) subjects.

Ambulatory group: Vitros FT4 bias = 0.77 (± 0.43) sBC - 3.01 (± 2.62); r = 0.34; P >0.05

Pregnant group: Vitros FT4 bias = 0.167 (± 0.077) sBC - 0.079 (± 0.167); r = 0.475; P <0.05

Hospitalized group: Vitros FT4bias = -0.31 (± 0.665) sBC + 0.29 (± 0.182); r = 0.098; P >0.05

The slopes and intercepts of all the relationships did not differ significantly from each other (P >0.05). The regression equation describing the relationship of the Vitros FT4 bias and sBC (the only individual patient category that yielded a significant slope and r value, both at P <0.05, was the pregnancy group) indicates that at the highest observed sBC (23.9 nmol/pmol), the Vitros FT4 will be positively biased by 4 pmol/L (95% confidence interval, 2.6–5.4 pmol/L).

There was no apparent relationship (P >0.05) between the PBT4 concentrations and the Vitros FT4 concentrations in any of the patient groups studies.

Fig. 3 depicts the relationship between the Vitros FT4 bias and %T3U in all patients studied. The equations derived in each of the individual patient categories are summarized below:



View larger version (12K):
[in this window]
[in a new window]
  		Figure 3. Relationship between the Vitros FT4 bias (determined by ED FT4) and %T3U in ambulatory ({diamondsuit}), hospitalized ({blacksquare}), and pregnant ({triangleup}) subjects.

Ambulatory group: Vitros FT4 bias = -0.04 (± 0.22) %T3U + 2.75 (± 6.46); r = 0.035; P >0.05

Pregnant group: Vitros FT4 bias = 0.003 (± 0.199) %T3U + 2.61 (± 3.78); r = 0.003; P >0.05

Hospitalized group: Vitros FT4 bias = 0.06 (± 0.066) %T3U - 3.37 (± 3.18); r = 0.19; P >0.05

None of these relationships reached statistical significance.

The results of the serum dilution experiment performed on the pregnancy sera are shown in Fig. 4 . The theoretically derived FT4 concentrations suggest that the decrease after a 32-fold dilution will be <1%. Similarly, the FT4 concentration as measured by ED was only slightly affected by serum dilution. At dilutions of 1:16 and 1:32 (i.e., dilutions that are expected to decrease the sBC to levels usually seen in some hospitalized patients) the ED FT4 was within 5% of the concentration obtained in the undiluted sample. The Vitros FT4 concentrations at these dilutions did not vary significantly (<5%) from the FT4 concentration obtained in the undiluted sample.



View larger version (10K):
[in this window]
[in a new window]
  		Figure 4. Mean percentage of change (from the undiluted sample) of ED ({diamondsuit}) and Vitros FT4 ({blacksquare}) after serial dilution of pregnancy sera with 10 mmol/L HEPES, pH 7.4.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
It is now accepted that FT4 shows a better correlation to the thyroid status than total T4 (1)(2)(3); thus it is the free hormone fraction that should be used to confirm the diagnosis of thyroid disease (17)(18)(19). The direct measurement of FT4 has, however, proved to be exceedingly difficult. Over the last two decades, numerous methodologies have been developed, and their limitations have been the subject of many publications and heated discussions in the literature (3)(7)(11)(20)(21)(22)(23)(24)(25)(26)(27)(28). Several limitations (e.g., "albumin" effects) of the early FT4 assays have been eliminated or minimized, but as shown recently, large methodological differences are still present (9). In their study, Nelson et al (9) measured FT4 (using different FT4 RIAs) in serum preparations in which the concentration of PBT4 (and sBC) was altered, whereas the ED FT4 concentration was kept constant. The results showed that the three FT4 methods studied were significantly, but to varying degrees, influenced by the serum PBT4. This method-specific dependency on PBT4 was proposed as a possible explanation for the discordant FT4 measurements seen in nonthyroidal illness. In an additional study, this group of investigators (10) suggested that the method-specific PBT4 dependency was a direct result of increased sequestration of T4 by the assay reagents.

We have used two simple tests to examine the bias of an automated (Vitros) FT4 method. The first test involved the comparison of the FT4 results obtained in the Vitros method in patients having a wide range of binding capacities against those obtained in a commercial direct ED system. The main reason for choosing ED as the reference method is that it is one of the methods, in addition to ultrafiltration, generally considered as the "gold standard" method for free thyroid hormone measurement. However, even these gold standards have their own inconsistencies and technical weaknesses (15)(29)(30)(31)(32). Thus, the elevation of this particular ED method as the definitive gold standard method is arguable. Nonetheless, the validity of the ED method chosen has been well documented (15), although even this ED may be biased in some nonthyroidal illness sera because of dilution effects during the dialysis step (33). The ED method we used produces a 13-fold dilution of dialyzable substances in sera. The patient categories included in the study have been documented to have high (i.e., third- trimester pregnancy) and low (i.e., hospitalized patients) binding capacities (13). This binding capacity profile has been confirmed in the present study by two independent methods (the sBC, derived by dividing the TT4 result by the ED FT4 result, and the %T3U). The second test used examined the effect of serum dilution on the FT4 concentration obtained by the methods under investigation (Vitros and ED FT4). The sample dilution test was chosen for several reasons: (a) one can alter the sBC in a predictable fashion (e.g., a twofold dilution will reduce the sBC by ~50%); (b) one can readily predict the ideal performance of an assay and thus eliminate the need to compare results with those obtained by a reference method; (c) one can "manufacture" serum dilution pools whose sBCs mimic the range of sBCs found in patients [e.g., the sBC concentration range in patients undergoing thyroid testing has been shown to be ~30-fold (13); thus dilution of a pregnant serum by this factor will encompass the sBC likely to be experienced by a laboratory]; and (d) this test can easily be performed by any user.

The law of mass action (2)(3) dictates that the concentration of FT4 in serum depends on the equilibrium that exists between the PBT4 and the concentration and affinity (i.e., the sBC) of the free binding sites:

In estimating the sBC (i.e., PBT4/FT4) we made the same assumption as Nelson and co-workers (9)(13), which is that the FT4 as measured by ED gives an unbiased estimate of the true FT4. As discussed previously, this assumption is arguable. The highest and lowest individual sBC values in our study population were 23.9 and 1.2 nmol/pmol, which represent a range of 19.9-fold. Thus, it is clear from this and other studies (9)(13) that patients undergoing thyroid function tests possess a very wide range of serum T4 binding (and consequently wide ranges of PBT4). The results presented here show that the FT4 concentrations obtained by the Vitros assay (relative to ED), were not dependent on T4 binding (sBC or T3U) or PBT4. The small positive bias (from ED) seen in pregnancy is likely because of calibration differences. This is supported by the fact that the corresponding FT4 concentrations in the ambulatory group were also positively biased to a similar degree.

In the second test, we examined the FT4 biases of the two methods (Vitros and ED FT4) by analyzing sera whose protein concentrations were decreased in vitro. Lowering of the sBC by decreasing the concentration of the binding proteins was accomplished by diluting sera in an inert buffer (HEPES). The FT4 concentrations as measured by both the ED and Vitros methods were not reduced by the serum dilution.

The estimation of serum FT4 by immunoassay, irrespective of the methodology used will invariably disturb the normal equilibrium between PBT4 and sBC (2)(3). This will come about as a result of the addition of the antibody and the dilution of the sample in assay reagents that may also contain T4 binders such as albumin or animal sera, which are often added to protect assays from interferences from heterophilic antibodies. These additions will lead to the establishment of a new equilibrium and a new "in vitro" FT4 concentration. The concentration of this in vitro FT4 will be dictated by the in vitro bound T4 and unbound binding site concentrations. The in vitro bound T4 will be the net sum of PBT4 and T4 bound by the immunoassay reagents (iPBT4), and the in vitro unbound binding sites will include, in addition to sBC, the free antibody-binding sites and the free binding sites of other binders included in the reagents [i.e., immunoassay binding capacity (iBC), which will be equal to the affinity x concentration of the free immunoassay binding sites]. Thus, the in vitro FT4 will be equal to (PBT4 + iPBT4)/(sBC + iBC). From this formula, one can predict that a high iBC will cause biases that will be dependent on the magnitude of the endogenous sBC. Thus, as the sBC decreases (as seen in hospitalized patients and mimicked by serum dilution), the assay will yield increasingly negative results. The results presented show a lack of sBC-dependent biases in both the Vitros and the ED FT4 assays, suggesting that the in vitro disturbance of the T4/protein equilibrium induced by both assays is negligible.

The tests described in the present study can be used to assess the presence and magnitude of sBC-dependent biases of other commercial FT4 methods.


   Footnotes
 
University Departments of 2 Clinical Biochemistry and 3 Anesthetics, The University of Edinburgh, Royal Infirmary, Lauriston Place, Edinburgh, Scotland, UK.

1 Nonstandard abbreviations: T4, thyroxine; FT4, free thyroxine; ED, equilibrium dialysis; PBT4, protein-bound thyroxine; sBC, serum binding capacity; T3, triiodothyronine; TT4, total thyroxine; T3U, triiodothyronine uptake; iPBT4, protein-bound thyroxine in the immunoassay; and iBC, immunoassay binding capacity.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Robbins J, Rall JE. The interaction of thyroid hormones and protein in biological fluids. Recent Prog Horm Res 1957;13:161-208.
  2. Ekins R. Measurement of free hormones in blood. Endocrinol Rev 1990;11:5-46. [Abstract/Free Full Text]
  3. Ekins R. The free hormone hypothesis and measurement of free hormones. Clin Chem 1992;38:1289-1293. [Free Full Text]
  4. Gow SM, Kellett HA, Toft AD, Beckett GJ. Accuracy and precision of five analog radioimmunoassays for free thyroxin compared. Clin Chem 1985;31:1888-1892. [Abstract/Free Full Text]
  5. Csako G, Zweig MH, Benson C, Ruddel M. On the albumin dependence of measurements of thyroxin. I. Technical performance of seven methods. Clin Chem 1986;32:108-115. [Abstract/Free Full Text]
  6. Wilke TJ. Five kits for estimating free thyroxine concentration in serum evaluated, and correlated with other indices to thyroid status. Clin Chem 1982;28:2051-2056. [Abstract/Free Full Text]
  7. Helenius T, Liewendahl K. Improved dialysis method for free thyroxin in serum compared with five commercial radioimmunoassays in nonthyroidal illness and subjects with abnormal concentrations of thyroxin-binding globulin. Clin Chem 1983;29:816-822. [Abstract/Free Full Text]
  8. Perdrisot R, Bounaud MP, Bounaud JY, Jallet P. Four nonisotopic immunoassays of free thyroxin evaluated. Clin Chem 1989;35:115-120. [Abstract/Free Full Text]
  9. Nelson JC, Weiss RM, Wilcox RB. Underestimates of serum free thyroxine (T4) concentrations by free T4 immunoassays. J Clin Endocrinol Metab 1994;79:76-79. [Abstract]
  10. Nelson JC, Nayak SS, Wilcox RB. Variable underestimates by serum free thyroxine (T4) immunoassays of free T4 concentrations in simple solutions. J Clin Endocrinol Metab 1994;79:1373-1375. [Abstract]
  11. Czako G, Zweig MH, Benson C, Ruddel M. On the albumin-dependence of measurements of free thyroxin. II. Patients with nonthyroidal illnesses. Clin Chem 1987;33:87-92. [Abstract/Free Full Text]
  12. Wong TK, Pekary AE, Hoo GS, Bradley ME, Hershman JM. Comparison of methods for measuring free thyroxine in nonthyroidal illness. Clin Chem 1992;38:720-724. [Abstract/Free Full Text]
  13. Nelson JC, Wilcox RB. Analytical performance of free and total thyroxine assays. Clin Chem 1996;42:146-154. [Abstract/Free Full Text]
  14. Christofides ND, Sheehan CP. Enhanced chemiluminescence labeled-antibody immunoassay (Amerlite MAB) for free thyroxine: design, development, and technical validation. Clin Chem 1995;41:17-23. [Abstract/Free Full Text]
  15. Nelson JC, Tomei RT. Direct determination of free thyroxine in undiluted serum by equilibrium dialysis/radioimmunoassay. Clin Chem 1988;34:1737-1744. [Abstract/Free Full Text]
  16. Cornbleet PJ, Gochman N. Incorrect least squares regression coefficients in method-comparison analysis. Clin Chem 1979;25:432-438. [Abstract/Free Full Text]
  17. Stockigt JR. Guidelines for diagnosis and monitoring of thyroid disease: nonthyroidal illness. Clin Chem 1996;42:188-192. [Abstract/Free Full Text]
  18. Becker DV, Bigos ST, Gaitan E, Morris JC, Rallison ML, Spencer CA, et al. Optimal use of blood tests for assessment of thyroid function [Letter]. JAMA 1993;269:2736.[Abstract/Free Full Text]
  19. Braverman LE. Evaluation of thyroid status in patients with thyrotoxicosis. Clin Chem 1996;42:174-178. [Abstract/Free Full Text]
  20. Bayer MF. Free thyroxin results are affected by albumin concentration and nonthyroidal illness. Clin Chim Acta 1983;130:391-396. [Web of Science][Medline] [Order article via Infotrieve]
  21. Midgley JEM, Moon CR, Wilkins TA. Validity of analog free thyroxin immunoassays. Part II [Opinion]. Clin Chem 1987;33:2145-2152. [Free Full Text]
  22. Ekins RP. Validity of analog free thyroxin immunoassays [Opinion]. Clin Chem 1987;33:2137-2152. [Abstract/Free Full Text]
  23. Chan DW. Nonthyroidal illness: performance of free T4 methods [Review]. J Clin Immunoassay 1984;7:206-211.
  24. Liewendahl K, Tikanoja S, Helenius T, Valimaki M. Discrepancies between serum free triiodothyronine and free thyroxin as measured by equilibrium dialysis and analog radioimmunoassay in nonthyroidal illnesses. Clin Chem 1984;30:760-762. [Abstract/Free Full Text]
  25. Stockigt JR, deGaris M, Csicsmann J, Barlow JW, White EL, Hurley DM. Limitations of a new free thyroxine assay (Amerlex FT4). Clin Endocrinol (Oxf) 1981;15:313-318. [Medline] [Order article via Infotrieve]
  26. Ooi G, Mahadevan MS, Greenway DC, Gertler SZ. Evaluation of four commercially available assays for free thyroxin. Clin Chem 1988;34:2302-2307. [Abstract/Free Full Text]
  27. Czako G, Zweig MH, Glickman J, Ruddel M, Kestner J. Direct and indirect techniques for free thyroxin compared in patients with nonthyroidal illness. II. Effect of prealbumin, albumin and thyroxin-binding globulin. Clin Chem 1989;35:1655-1662. [Abstract/Free Full Text]
  28. Wilkins TA. Albumin in analog FT4 assay reagents: the facts [Letter]. Clin Chem 1987;33:1293.[Free Full Text]
  29. Tikanoja SH, Liewendahl K. New ultrafiltration method for free thyroxin compared with equilibrium dialysis in patients with thyroid dysfunction and nonthyroidal illness. Clin Chem 1990;36:800-804. [Abstract/Free Full Text]
  30. Docter R, van Toor H, Krenning EP, de Jong M, Hennemann G. Free thyroxine assessed with three assays in sera of patients with nonthyroidal illness and of subjects with abnormal concentrations of thyroxine-binding proteins. Clin Chem 1993;39:2805-2811.
  31. Shannon N, Woolf PD. Determination of free thyroxin in serum by ultrafiltration: validation of a method and preliminary results. Clin Chem 1984;30:1770-1773. [Abstract/Free Full Text]
  32. Wang Y-S, Hershman JM, Pekary AE. Improved ultrafiltration method for simultaneous measurement of free thyroxine and free triiodothyronine in serum. Clin Chem 1985;31:517-522. [Abstract/Free Full Text]
  33. Nelson JC, Weiss RM. The effect of serum dilution on free thyroxine (T4) concentration in the low T4 syndrome of nonthyroidal illness. J Clin Endocrinol Metab 1985;61:239-246. [Abstract/Free Full Text]



The following articles in journals at HighWire Press have cited this article:


Home page
 Ann Clin BiochemHome page
A. Waise and H. C Price
The upper limit of the reference range for thyroid-stimulating hormone should not be confused with a cut-off to define subclinical hypothyroidism
Ann Clin Biochem, March 1, 2009; 46(2): 93 - 98.
[Abstract] [Full Text] [PDF]

Home page
 Clin. Chem.Home page
R. Sapin and M. d'Herbomez
Free Thyroxine Measured by Equilibrium Dialysis and Nine Immunoassays in Sera with Various Serum Thyroxine-binding Capacities
Clin. Chem., September 1, 2003; 49(9): 1531 - 1535.
[Full Text] [PDF]

Home page
 Clin. Chem.Home page
R. Sapin, J.-L. Schlienger, F. Gasser, E. Noel, B. Lioure, F. Grunenberger, B. Goichot, and D. Grucker
Intermethod Discordant Free Thyroxine Measurements in Bone Marrow-transplanted Patients
Clin. Chem., March 1, 2000; 46(3): 418 - 422.
[Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via ISI Web of Science (17)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Christofides, N. D.
Right arrow Articles by Beckett, G. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Christofides, N. D.
Right arrow Articles by Beckett, G. J.
Related Collections
Right arrow Automation and Analytical Techniques

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS