Standardisation of a two-site PTH immunoradiometric assay using various solid phase formats.

BACKGROUND & OBJECTIVES: Estimation of parathyroid hormone (PTH) levels is important in the management of metabolic bone disorders. Here we describe a simple, sensitive and specific second generation immunoradiometric assay (IRMA) to detect intact PTH levels using different solid phase matrices. Different methods for immobilization of antibodies have also been evaluated. METHODS: Experiments were carried out with physical adsorption of antibodies, covalent coupling using 2 per cent glutaraldehyde and N,N`carbonyldiimidazole. In all cases, antibodies raised against C-terminal were used as solid phase agent. Detector antibodies were N terminal antibodies that were radio-iodinated with [125] I followed by gel purification. Several of the antibodies coupled to various solid phase matrices were incubated with PTH standards and the detector antibody as well as the commercially available tracer from DiaSorin kit to identify a suitable match pair. RESULTS: The best pair was polyclonal C-terminal PTH antibody along with the kit tracer from DiaSorin with regards to antibody coated to magnetic cellulose particles. Among the various antibodies and the solid phases evaluated, the best assay was obtained with the matched pair of antibodies (70×G67 and 70×G68) from Fitzgerald immobilized on polystyrene tubes. The polyclonal antibody against C-terminal PTH was chosen as the capture antibody and [125] I labelled polyclonal antibody against N-terminal PTH as the tracer. The sample values obtained in the antibody coated tubes were comparable to those obtained using a commercial kit. INTERPRETATION & CONCLUSIONS: The results indicated the feasibility of adopting this system for further development into a PTH IRMA for regular production as there is no indigenous kit available for intact PTH.

[Inappropriately undetectable thyroglobulin after exclusion of interference from thyroglobulin antibodies]. / Discordances entre différents immunodosages de la thyroglobuline en l'absence d'anticorps anti-thyroglobuline.

Thyroglobuline (Tg) is a large molecule of high molecular weight mainly indicated in monitoring differentiated thyroid cancer (DTC), its measurement remains difficult. We report the case of a patient who underwent total thyroidectomy for a poorly differentiated thyroid insular carcinoma. Despite several (131)I treatments, a progressive elevation of serum Tg is observed. A control performed in another laboratory using an immunoradiometric assay (IRMA Cisbio) returns an undetectable value (< 0,2 mg/L). A new sample was simultaneously sent to different laboratories. Three nonisotopic immunometric assays showed high values of Tg while the IRMA assay, considered the gold standard, gave a result below the detection threshold. The absence of Tg antibodies and of anti-mouse antibodies was confirmed. The IRMA Kit manufacturer agreed to carry out an expertise. After changing their detection antibody, the presence of a high Tg was demonstrated, in agreement with non-isotopic techniques. The expertise conclusion was a lack of detection by the IRMA Tg assay. This incident was notified to AFSSAPS by the manufacturer.

[Influence of temperature on the calibration curves in IRMA for neuron-specific enolase and its physicochemical interpretation]. / Influencia de la temperatura en las curvas de calibración en IRMA de enolasa neuronal específica y su interpretación fisicoquímica.

BACKGROUND: immunoradiometric assay (IRMA) is one of the principal methods used for the analytical determination of neuron specific enolase (NSE) concentration. We studied the influence of temperature on the calibration curves obtained by this method, and a physicochemical justification based on two theoretical models is proposed. MATERIAL AND METHODS: we used a commercially available RIA kit for NSE and a gamma counter. Data was analysed using Statistical software. RESULTS AND DISCUSSION: activity bound to the antibody increases with temperature, producing results that are consistent with two modifications to the four parameter and Langmuir equations. CONCLUSIONS: the two models used successfully reproduce the results, with the adsorption model being preferable due to its greater simplicity and clearer physical significance.

Selection of calibration standard concentrations for determination of intact-PTH by immunoradiometric assay.

The routine determination of parathyroid hormone (PTH) by immunoradiometric assay (IRMA) has been studied. Concentrations of standards have been adequate to the clinical range in order to reduce errors. Proposed standards were tested by the calculation of different quality parameters. Recoveries of sample concentrations were estimated for different experimental alterations (methodological errors, reagent degradation, or changes in background response). Finally, inter-assays demonstrated that reproducibility of samples with concentrations in the critical clinical limits was improved. The results confirmed that the proposed selection provided a more robust method and it is possible to extrapolate to other clinical immunoassays.

Serum levels of insulin-like growth factor-I and insulin-like growth factor-I binding protein-3: quality control for studies of stored serum.

The insulin-like growth factor (IGF) axis, particularly IGF-I and IGF binding protein-3 (IGFBP-3), has been the subject of much attention because of its role in juvenile growth and their association with cancers at several sites. However, epidemiologic studies of IGF-I and IGFBP-3 have had mixed results and several authors have speculated that quality control (QC), sample storage history, and other methodologic concerns could play a role in this heterogeneity. This article documents the results of storage history and QC efforts for a study of IGF-I and IGFBP-3 in 6,226 serum samples from the National Health and Nutrition Examination Survey III (NHANES III). The study was carried out on site at Diagnostic Systems Laboratories in Webster, Texas, using the IGF-I ELISA (DSL 10-5600) and the IGFBP-3 immunoradiometric assay (DSL 6600). A run-in study of assay performance suggested that plates, days, and weeks significantly affected the variance of both assays. Analysis of samples with different storage histories also indicated strong effects of storage history. Serum samples disbursed to laboratories for measurement of diverse analytes and then returned for storage showed reductions in serum IGF-I level averaging 43% and reductions in IGFBP-3 of 25% compared with samples shipped immediately to the repository for long-term storage at -80 degrees C. Therefore, the main study was carried out using samples that had been shipped directly to the National Center for Health Statistics/NHANES collection center for storage. Laboratory analyses of NHANES III and QC samples were carried out over approximately 10 months. QC was monitored through repeated testing of blood samples from six individuals, with two individuals tested twice on each plate. Assay performance was stable over the entire study and coefficients of variation averaged 2% to 3% within plates and approximately 14% for IGF-I and approximately 11.5% for IGFBP-3 over the entire study. Coefficients of variation varied significantly among individual QC subjects, ranging from 12.3% to 17.6% for IGF-I and 8.9% to 12.8% for IGFBP-3. Based on Levy-Jennings plots, approximately 5% of the plates used for IGF-I in the main study were out of compliance. Finally, location on a plate had small but significant effects on IGF-I level. Together, these results highlight the need for care in large studies of putative biomarkers for cancer risk and illustrate some probable sources of heterogeneity in past epidemiologic studies of the IGF axis and cancer.

Conformational changes in prorenin during renin inhibition in vitro and in vivo.

BACKGROUND: Some renin inhibitors induce changes in the conformation of prorenin in vitro and influence the quantification of active renin by immunoradiometric assays. Whether such changes in renin recognition by monoclonal antibodies exist after oral administration of aliskiren, the first orally available renin inhibitor, is not known. METHODS: Two commercially available immunoradiometric assays (Cisbio and Nichols) were compared to determine immunoreactive active renin concentrations in plasma samples collected in a single oral dose crossover study comparing the renin inhibitor, aliskiren (300 mg), with the angiotensin II antagonist, valsartan (160 mg), in healthy male subjects. RESULTS: The addition of aliskiren to plasma samples in vitro, at concentrations of 1-100 micromol/l, increased active renin immunoreactivity in both the Cisbio and Nichols assays. In the crossover study, the two assays gave similar values for the plasma immunoreactive active renin concentration before treatment and following valsartan administration (intraclass coefficient for agreement between the two assays = 0.92). However, a Bland-Altman plot showed a systematic bias towards higher values (1.75-fold higher; 95% confidence interval = 1.02-3.01) in the Nichols than in the Cisbio assay following aliskiren administration. The difference between the results obtained with the two assays depended on incubation time. CONCLUSIONS: Depending on incubation conditions, circulating renin inhibitors interfere with the recognition of active renin molecules by the monoclonal antibodies used in commercially available assays. Careful consideration must therefore be given to the methodology used for quantifying immunoreactive plasma active renin when patients are treated with renin inhibitors, to avoid an overestimation of the magnitude of active renin release attributable to conformational changes in plasma prorenin.

Comparison between two methods in the determination of circulating chromogranin A in neuroendocrine tumors (NETs): results of a prospective multicenter observational study.

Several methods for analyzing CgA using either monoclonal or polyclonal antibodies have been developed, which differ in their diagnostic performance. The present paper describes the results of a prospective multicenter study aimed at comparing the clinical value of the two most widely used commercially available CgA assay kits in patients affected by neuroendocrine tumors (NETs). Two hundred sixty-one patients from 40 different centers and 99 healthy subjects were evaluated. CgA levels were measured with two different methods, a two-step immunoradiometric assay (IRMA) and an enzyme-linked immunosorbent assay (ELISA). CgA was measured centrally by two reference laboratories, one of which used IRMA and the other ELISA, and it was measured by the participating institutions with the method routinely used by each of them. The major findings of the present study were: (i) the two assays for the determination of CgA present good diagnostic performance; (ii) both assays are robust and guarantee comparable results when applied in different settings (central vs local laboratory); (iii) the negative/positive cutoff points (87 ng/mL for IRMA and 21.3 U/L for ELISA) were established according to standardized criteria; (iv) the results obtained with the two assays in basal clinical samples of patients affected by NETs show an apparently satisfactory correlation (rs = 0.843, p < 0.0001). However, a possibly clinically meaningful 36% discordance rate was found. These findings support the hypothesis that the two CgA kits might provide partially different information.

Development of a novel immunoradiometric assay exclusively for biologically active whole parathyroid hormone 1-84: implications for improvement of accurate assessment of parathyroid function.

We developed a novel immunoradiometric assay (IRMA; whole parathyroid hormone [PTH] IRMA) for PTH, which specifically measures biologically active whole PTH(1-84). The assay is based on a solid phase coated with anti-PTH(39-84) antibody, a tracer of 125I-labeled antibody with a unique specificity to the first N-terminal amino acid of PTH(1-84), and calibrators of diluted synthetic PTH(1-84). In contrast to the Nichols intact PTH IRMA, this new assay does not detect PTH(7-84) fragments and only detects one immunoreactive peak in chromatographically fractionated patient samples. The assay was shown to have an analytical sensitivity of 1.0 pg/ml with a linear measurement range up to 2,300 pg/ml. With this assay, we further identified that the previously described non-(1-84)PTH fragments are aminoterminally truncated with similar hydrophobicity as PTH(7-84), and these PTH fragments are present not only in patients with secondary hyperparathyroidism (2 degrees -HPT) of uremia, but also in patients with primary hyperparathyroidism (1 degrees -HPT) and normal persons. The plasma normal range of the whole PTH(1-84) was 7-36 pg/ml (mean +/- SD: 22.7 +/- 7.2 pg/ml, n = 135), whereas over 93.9% (155/165) of patients with 1 degrees -HPT had whole PTH(1-84) values above the normal cut-off. The percentage of biologically active whole PTH(1-84) (pB%) in the pool of total immunoreactive "intact" PTH is higher in the normal population (median: 67.3%; SD: 15.8%; n = 56) than in uremic patients (median:53.8%; SD: 15.5%; n = 318; p < 0.001), although the whole PTH(1-84) values from uremic patients displayed a more significant heterogeneous distribution when compared with that of 1 degrees -HPT patients and normals. Moreover, the pB% displayed a nearly Gaussian distribution pattern from 20% to over 90% in patients with either 1 degrees-HPT or uremia. The specificity of this newly developed whole PTH(1-84) IRMA is the assurance, for the first time, of being able to measure only the biologically active whole PTH(1-84) without cross-reaction to the high concentrations of the aminoterminally truncated PTH fragments found in both normal subjects and patients. Because of the significant variations of pB% in patients, it is necessary to use the whole PTH assay to determine biologically active PTH levels clinically and, thus, to avoid overestimating the concentration of the true biologically active hormone. This new assay could provide a more meaningful standardization of future PTH measurements with improved accuracy in the clinical assessment of parathyroid function.

Effects of growth hormone-binding proteins on serum growth hormone measurements.

GH-binding proteins (GH-BPs) in human blood theoretically may interfere with measurements of immunoreactive GH by forming complexes with GH and competing with antibody reagents for ligand. Indeed, results of serum GH obtained by immunoassays are known to differ markedly depending on the assay employed. To assess the potential role of circulating GH-BPs in this phenomenon, we systematically examined their effect on GH measurement in four RIAs and two immunoradiometric assays. In all except one RIA, the effect of the BPs on tracer binding to antibody was mildly inhibitory. In both immunoradiometric assays, BPs increased the nonspecific association of the tracer with the solid phase reagent. However, these effects were minor at the BP concentrations realistically encountered in practice. Furthermore, the impact of BPs on GH standard curves and final results was negligible because BP effects fall into the category of nonspecific or zero dose counts, which are subtracted during data reduction. We conclude that GH-BPs are only a minor disturbance in GH immunoassays, which is completely compensated for by conventional assay design. Disparities among results yielded by different assays are probably not due to BP interference.

Evaluation of diagnostic accuracy of insulin-like growth factor (IGF)-I and IGF-binding protein-3 in growth hormone-deficient children and adults using ROC plot analysis.

We critically evaluated the diagnostic value of IGF-I and IGF-binding protein-3 (IGFBP-3) in GH deficiency (GHD) in children and adults using receiver operating characteristic (ROC) plot analysis. Sixty-six children (chronological age, 1.3-15 yr) were studied: 34 GHD and 32 idiopathic short stature (ISS). Ninety-two adults (chronological age, 18-70 yr) were also evaluated: 72 GHD, 34 of childhood onset (AGHD-CO), and 38 of adult onset (AGHD-AO); and 20 healthy volunteers. The SD score (SDS) for IGF-I was calculated from 596 normal subjects (212 children and 384 adults), and the SDS for IGFBP-3 was calculated from 350 normal subjects (212 children and 138 adults). The ROC plot showed that the best IGF-I SDS cut-off line was -1.65 for children [sensitivity (S), 68%; specificity (Sp), 97%, diagnostic efficiency (DEf), 81%], the cut-off line for AGHD was -1.65 for AGHD-CO (S, 91%; Sp, 100%; DEf, 94%), and the cut-off line for AGHD-AO was -1.80 (S, 81%; Sp, 100%; DEf, 88%). For IGFBP-3 SDS, the best cut-off line was -1.80 for children (S, 90%; Sp, 60%; DEf, 78%); it was -1.45 for AGHD-CO (S, 90%; Sp, 75%; DEf, 82%) and -0.90 for AGHD-AO (S, 90%; Sp, 68%; DEf, 77%). An accurate diagnosis was obtained using IGF-I SDS alone in GHD children 65%; ISS, 97%; AGHD-CO, 92%; AGHD-AO, 86%, with IGFBP-3 SDS alone in GHD children 60%; ISS, 90%; AGHD-CO, 75%; AGHD-AO, 68%. Considering both, an accurate diagnosis was obtained in GHD children 60%; ISS, 87%; AGHD-CO, 71%; AGHD-AO, 64%. In conclusion, our findings support the need to use cut-off lines expressed in SDS obtained using an appropriate statistical methodology for better characterization of the various clinical presentations. IGF-I proved to be more useful because of its good diagnostic efficiency and accuracy in both children and adults, whereas IGFBP-3 did not significantly contribute to the diagnosis of GHD.

Standards for assay of von Willebrand factor concentrates: a collaborative study.

A multilaboratory collaborative study was undertaken to assess the feasibility of using a plasma standard for expressing the results of assays for the von Willebrand factor content of von Willebrand factor concentrates and of factor VIII concentrates. Thirteen laboratories tested six concentrates for von Willebrand factor antigen, ristocetin cofactor activity, and multimer content using the World Health Organization plasma standard for factor VIII/von Willebrand factor, 87/718, as a standard. Only a few assays were invalid because of nonparallelism or nonlinearity. Significant interlaboratory and interassay differences were found for both von Willebrand factor antigen and ristocetin cofactor activity. There was generally good agreement between the laboratories with respect to the multimer content in the preparations. With respect to assay validity, a plasma standard could be suitable for assaying concentrated preparations of von Willebrand factor.

Remission criteria for the follow-up of patients with acromegaly.

OBJECTIVE: The aim was to evaluate the validity of current remission criteria in acromegaly, a random GH level of <2.5 microg/l, a glucose-suppressed GH level of <1 microg/l and a normal IGF-I level. DESIGN: In forty-one patients treated for acromegaly (23 males and 18 females, 20-69 years) and 94 healthy subjects (50 males and 44 females, 20-78 years), basal GH and IGF-I levels and nadir GH levels after 75 g oral glucose were evaluated in decade blocks; these were assayed by sensitive immunoradiometric assays. RESULTS: Basal GH levels varied widely from 0.022 to 10.4 in healthy subjects and were >2.5 microg/l in 19%. The mean post-glucose GH nadir was 0.067+/-0.009 microg/l (range 0.003-0.4 microg/l) and the upper limit of the GH nadir was 0.26 microg/l (means+2 S.D.) in healthy subjects. Thirty-five patients with acromegaly had high-for-age IGF-I levels in relation to our healthy subjects. In this group, 15 (42.9%) patients had basal GH levels of <2.5 microg/l, 14 (40%) patients had nadir GH levels of <1 microg/l, and three (8.6%) patients had GH suppression to <0.26 microg/l which was defined as normal GH suppression in our healthy subjects. Only six patients with acromegaly had normal-for-age IGF-I levels and all of these patients had basal GH levels of <2.5 microg/l and all but one had nadir GH levels of <0.26 microg/l. CONCLUSIONS: A basal or random GH level of <2.5 microg/l is not a reliable criterion for remission in acromegaly and the currently accepted normal upper limit of 1 microg/l for post-glucose GH suppression is too high. Post-glucose nadir GH levels, measured with sensitive assays, can be <1.0 microg/l in 40% and basal GH levels can be <2.5 microg/l in 43% of the active acromegalic patients. IGF-I levels appeared to correlate better with a nadir GH cut-off of 0.26 microg/l rather than 1 microg/l in the determination of disease activity.

Urinary excretion rate of ceruloplasmin in non-insulin-dependent diabetic patients with different stages of nephropathy.

The level of ceruloplasmin, which is a more negatively charged protein than albumin, was measured by an immunoradiometric assay in timed overnight urine and serum samples from patients with non-insulin-dependent diabetes mellitus and healthy controls. None of the plasma proteins examined showed any cross-reactivity in this assay. A linear correlation was seen between the ceruloplasmin level and the serial dilution of the sample. Western blot analysis using concentrated urine samples showed that the molecular weight of ceruloplasmin in the urine sample was the same as that of ceruloplasmin in the serum and standard samples. These findings indicated that the substance detected by this assay was truly ceruloplasmin. The urinary ceruloplasmin excretion rate (CER) and clearance of ceruloplasmin increased in parallel with the progression of albuminuria. The highest CER was found in macroalbuminuric patients, followed by micro- and normoalbuminuric patients and the healthy control subjects, the differences between the groups being significant. In view of the fact that the isoelectric point of ceruloplasmin (4.4) is more acidic than that of albumin, the present findings suggested that an enhanced CER was due either to the alteration of charge selectivity in the glomerular basement membrane with unaltered tubular function or to a defect of the non-discriminatory pores (shunt pathway) with unaltered tubular function.

Influence of apolipoprotein(a) phenotype on lipoprotein(a) quantification: evaluation of three methods.

Three commercially available assays (an enzyme-linked immunosorbent assay ELISA, an immunoradiometric assay, IRMA, and a nephelometric assay) for the determination of lipoprotein(a) [Lp(a)] were compared with respect to the dependency of these assays on the various apolipoprotein(a) [apo(a)] isoforms. Although there was a strong correlation between the three methods, a significant difference between the absolute values (mg/L) was observed (p < 0.001). Using purified Lp(a) preparations, we showed that the ELISA assay quantifies the Lp(a) concentration on a molar basis, independently of the apo(a) isoform size. The IRMA and the nephelometric assay however are apo(a) isoform size dependent and overestimate the Lp(a) concentration of large apo(a) isoforms whereas the amount of small apo(a) isoforms is underestimated. In general, the isoform dependency of the Lp(a) quantification is of limited clinical relevance. In this study, inconsistent risk assignments are made in approximately 3% of the cases, when the Lp(a) concentrations obtained with the apo(a) isoform dependent assays are compared with the isoform independent ELISA.

Analytical performance of CA 19.9, CA 125 and CA 15.3 assays as observed through an external quality assessment program.

Immunoassays of the tumor markers CA 19.9, CA125 and CA 15.3 are generally acknowledged to be a useful tool in the management of cancer patients. As a consequence, many methods developed by different companies are now commercially available. However, discrepancies have been described in the results of marker determinations even when the same monoclonal antibody was used. An external quality assessment (EQA) was carried out; starting from 1989 about 110 laboratories participated; since December 1991 the program was linked with the interlaboratory program Oncocheck organized by the Service de Radiopharmacie et Radioanalyse, University of Lyon. At present more than 200 laboratories of many European countries are involved: cumulatively 47 quality control samples have been prepared and sent to the participants. This manuscript is a report on data collected for CA 19.9, CA 125, and CA 15.3.

Parathyroid hormone: what are we measuring and does it matter?

Immunometric assays claiming to determine intact parathyroid hormone (PTH) generally cross-react with N-truncated forms such as PTH(7-84). Laboratories need to examine the relevance of new assays with probable PTH(1-84) specificity. It is logical that assays should measure what they state they do. However, it seems unlikely that use of older 'intact' PTH assays will affect the clinical interpretation of results in primary hyperparathyroidism or vitamin D deficiency. It is plausible that appropriate application of new PTH assays could improve outcome in chronic renal failure. However, it has never been suggested that straightforward replacement of existing assays with new PTH(1-84) assays will lead to this improved outcome. A better understanding of PTH fragments and their interaction with PTH receptors may shed light on the relevance of different PTH assays. In the meantime, older technologies will continue to work well for the vast majority of patients.

Sensitive calcitonin measurement by two-site immunometric assays: implications for calcitonin screening in nodular thyroid disease.

The aim of this study was to investigate the impact of analytical aspects on the clinical usefulness of calcitonin (CT) measurement. In a retrospective analysis, CT levels measured by a polyclonal immunometric assay (Scantibodies Laboratory, CA, USA) were evaluated in various clinical situations. CT in newly diagnosed medullary thyroid cancer (MTC) (n = 20) ranged from 15.5-87130 pg/ml (median 661 pg/ml). Levels >10 pg/ml were seen in 7.3% of 314 patients with benign nodules, 48.9% of 45 patients with impaired kidney function, 97.7% of 87 patients on hemodialysis, 30.2% of 43 patients after renal transplantation, and in 71.0% of 31 patients with critical illnesses. Subgroups of patients were reevaluated by two monoclonal immunometric assays specific for mature CT. CT levels measured by the monoclonal immunometric assays were highly correlated to the polyclonal assay results in MTC patients, but were significantly different with a lower incidence of elevated levels in patients with renal disease and critical illnesses. In conclusion, highly sensitive assays with cut-off values of 10 pg/ml or below are mandatory for CT screening in nodular thyroid disease. The specificity of CT measurement in patients with renal disease and critical illnesses is higher with monoclonal assays specific for monomeric CT. These methodological aspects have to be regarded if CT measurement is used for decision making in nodular thyroid disease.

Thyroglobulin antibodies in differentiated thyroid cancer.

A retrospective review of patients with differentiated thyroid cancer (DTC) who were seen between 1984 and 1996 at the Royal Marsden Hospital identified 40 patients with serum thyroglobulin antibodies (TgAb). These antibodies can interfere with the immunoradiometric assay for serum thyroglobulin (Tg) used at this hospital, with resulting underestimation of the Tg level. A review of the case notes was carried out to ascertain the clinical significance of TgAb. The median follow-up from diagnosis of DTC was 26 months (range 3-401). The median age at diagnosis of DTC was 50 years (range 13-83). Patients were grouped according to the TgAb titre (high titre: TgAb >1/100, n = 28; low titre TgAb <1/100, n = 12). Thirteen patients relapsed, 11 in the high titre group and two in the low titre group. Sites of recurrence were: neck (n = 9), lung (n = 5), bone (n = 4) and brain (n = 2). No patient in the high titre group showed an elevated Tg with recurrence. One patient in the low titre group showed a Tg response to recurrence. Overall, the Tg assay failed to detect 92% of recurrences. Eight patients in the high titre group developed TgAb, apparently in response to tumour progression. In a third patient in the low titre group, the TgAb also acted as a 'tumour marker'. Thus, overall TgAb acted as a tumour marker in nine of the 40 (22.5%) patients in whom it was detected, and in nine of the 470 (1.9%) patients on follow-up during this time period. The overall survival of the whole group was 69% at 10 years. For patients with papillary carcinoma (n = 34) overall survival was 78% at 10 years. Laboratories should report routinely the presence of TgAb, with a caution indicating the direction of possible error (which depends on the assay used). Clinicians should appreciate that Tg measurements are unreliable in the presence of TgAb and that the development of TgAb can indicate active tumour.

Evaluation of second generation CA 125 assays.

OBJECTIVE: This study was performed in order to evaluate and compare the serum CA 125 values obtained using an immunoradiometric (IRMA-II) and an immunoenzymatic (ETI-II) second generation assay, and to establish whether or not the two methods may be used interchangeably. STUDY DESIGN: Serum CA 125 levels were measured in parallel using IRMA-II and ETI-II CA 125 assays (Sorin Biomedica), in 82 women with benign or malignant gynecological diseases. Statistical analysis was performed by linear regression analysis and Wilcoxon's test. RESULTS: Serum CA 125 levels measured using the immunoenzymatic method were lower than those obtained by the immunoradiometric assay. The largest discrepancies between the two methods were found at concentrations of 35-100 U/ml, within which fall cutoff values for the immunoradiometric assay. The cutoff values of 35 or 65 U/ml, frequently used in the original immunoradiometric assay and retained for the immunoradiometric second generation assay, corresponded to 18 and 47 U/ml in the immunoenzymatic second generation assay. CONCLUSION: The discrepancies in CA 125 results obtained by the two detection methods imply that the cutoff values used in the immunoenzymatic procedure should have a lower reference value in order to eliminate high rates of false negative results. Furthermore, their interchangeable use should be avoided in the monitoring of ovarian cancer and other gynecological diseases.

The validity of capillary blood sampling in the determination of human growth hormone concentration during exercise in men.

BACKGROUND: Studies measuring human growth hormone (hGH) in blood during exercise have mainly used venous sampling. The invasive nature of this procedure makes evaluation of hGH impossible in various exercise environments. OBJECTIVE: To determine whether capillary sampling could offer an alternative sampling method. METHODS: Capillary and venous blood samples were collected for determination of hGH at the end of each exercise stage during an incremental exercise test in 16 male club level competitive cyclists (mean (SD) age 30.8 (8.0) years, body mass 72.2 (7.1) kg, body fat 12.9 (3.5)%, peak oxygen consumption 4.18 (0.46) l x min(-1)). Linear regression, from a plot of venous v capillary blood hGH concentration, showed a correlation coefficient of r = 0.986 (p<0.001). When geometric means and log transformations were used, a coefficient of variation of 14.2% was demonstrated between venous and capillary flow for hGH concentration. The mean ratio limits of agreement were 0.62 (1.72)-that is, 95% of the ratios were contained between 0.36 and 1.07, with a mean of 0.62. CONCLUSIONS: Capillary blood sampling is an acceptable alternative to venous sampling for determining hGH concentration during rest and exercise. Sample sites should not be used interchangeably: one site should be chosen and its use standardised.