Measurement of midregional proadrenomedullin in plasma with an immunoluminometric assay

BACKGROUND: Adrenomedullin (ADM) is a potent vasodilatory peptide, and circulating concentrations have been described for several disease states, including dysfunction of the cardiovascular system and sepsis. Reliable quantification has been hampered by the short half-life, the existence of a binding protein, and physical properties. Here we report the technical evaluation of an assay for midregional pro-ADM (MR-proADM) that does not have these problems. METHODS: MR-proADM was measured in a sandwich immunoluminometric assay using 2 polyclonal antibodies to amino acids 45-92 of proADM. The reference interval was defined in EDTA plasma of 264 healthy individuals (117 male, 147 female), and increased MR-proADM concentrations were found in 95 patients with sepsis and 54 patients with cardiovascular disease. RESULTS: The assay has an analytical detection limit of 0.08 nmol/L, and the interassay CV was <20% for values >0.12 nmol/L. The assay was linear on dilution with undisturbed recovery of the analyte. EDTA-, heparin-, and citrate-plasma samples were stable (<20% loss of analyte) for at least 3 days at room temperature, 14 days at 4 degrees C, and 1 year at -20 degrees C. MR-proADM values followed a gaussian distribution in healthy individuals with a mean (SD) of 0.33 (0.07) nmol/L (range, 0.10-0.64 nmol/L), without significant difference between males or females. The correlation coefficient for MR-proADM vs age was 0.50 (P < 0.001). MR-proADM was significantly (P < 0.001) increased in patients with cardiovascular disease [median (range), 0.56 (0.08-3.9) nmol/L] and patients with sepsis [3.7 (0.72-25.4) nmol/L]. CONCLUSIONS: MR-proADM is stable in plasma of healthy individuals and patients. MR-proADM measurements may be useful for evaluating patients with sepsis, systemic inflammation, or heart failure.     

Technical evaluation of a new immunoradiometric and a new immunoluminometric assay for thyroglobulin

BACKGROUND: After removal of differentiated thyroid carcinoma (DTC), serum thyroglobulin (Tg) can indicate persistent or recurrent disease. We describe two novel two-step assays designed to measure low Tg concentrations. METHODS: We evaluated prototypes of the new IRMA, DYNOtest Tg-pluS, and the new immunoluminometric assay (ILMA), LUMItest) Tg-pluS. In the first step, a high-salt incubation buffer leads to dissociation of Tg-Tg antibody complexes in serum and is intended to reduce nonspecific interference and interference of potential Tg autoantibodies in the system. We studied recovery of human Tg (from thyroid glands) added to horse serum. We also studied 58 patients with DTC in whom Tg values under thyroid-stimulating hormone (TSH) suppression and TSH stimulation (without thyroxine) were available. RESULTS: The detection limits were 0.04 microg/L Tg for the IRMA and 0.02 microg/L for the ILMA. Intraassay imprecision (CV) was <10% over the range of the calibration curve in both assays. The day-to-day CV was <20% at 0.2 microg/L for the IRMA and at 0.06 microg/L for the ILMA. No high-dose hook effect was seen with up to 200 000 microg/L added Tg or in dilutions of 12 patient sera with Tg values of 307-38 880 microg/L. Mean recovery of 50 microg Tg/L was 96% in those patients. Among 77 samples with Tg antibody values of 65.2-8150 kilounits/L, recovery by the IRMA was disturbed in 7 cases (9%) and by the ILMA in 9 cases (12%). Tg increased as measured in both assays in 50 of 58 patients after thyroxine withdrawal. CONCLUSIONS: The new assays have improved precision for Tg <1 microg/L, and even low measured Tg concentrations respond physiologically to thyroxine withdrawal. The assays are free of a high-dose hook effect up to a Tg concentration of at least 38 000 microg/L and may further reduce Tg antibody interference.     

Evaluation of the first automated thyroglobulin assay.

The aim of this study was to investigate technical and analytical performance of the first automated thyroglobulin (Tg) assay (DPC-Immulite; Diagnostic Products Corporation, Los Angeles, USA). In imprecision studies using several human serum pools ranging from 21 to 58 replicates, a coefficient of variation of 9.0% was obtained at a mean Tg concentration of 0.84 ng/ml and of 6.1% at a Tg concentration of 62.1 ng/ml. In a method comparison with a non-automated assay (BRAHMS LUMItest Tg, BRAHMS, Berlin, Germany) using 383 sera of 303 patients with thyroid carcinoma, regression analysis according to Passing and Bablock yielded in the following equation: Immulite Tg = 1.6 x BRAHMS Tg-0.1 ng/ml (Pearson's r = 0.979). Sera obtained from 59 patients with thyroid carcinoma enabled comparative follow-up studies; in all cases qualitative agreement was found with regard to increase or decrease of serum Tg; in eight cases, however, Tg was detected with the Immulite assay but not with the BRAHMS assay. Further follow-up proved the presence of thyroid tissue in these patients. From these and further methodological data (dilution linearity, interference studies, carry-over study, high-dose hook properties, and short report time) it is concluded that the DPC-Immulite Tg assay meets the requirements of routine diagnostic use.     

Evaluation of an automated commercial immunoluminometric assay for alpha-foetoprotein.

We evaluated a two-site immunoluminometric assay (AFP LIA-mat Byk Sangtec) for the determination of alpha-foetoprotein (AFP). The assay, involving two monoclonal antibodies which recognize two different alpha-foetoprotein epitopes, is rapid (4 h) with a wide working range (0-600 x 10(3) IU/l), a good lower limit of detection and good reproducibility (CV less than 10%). The regression equation for the AFP LIA-mat (y) and the immunoradiometric assay AFP Bridge Serono (x) was y = 1.175x - 2.27 (n = 95, r = 0.996) for serum and y = 1.16x + 479.2 for amniotic fluid. It did not display a hook effect, and when we assessed the linearity by assaying a high concentration of alpha-foetoprotein, it gave a linear response down to 3.1 x 10(3) IU/l. We also evaluated the clinical response of AFP LIA-mat in 278 patients with different diseases. Eighteen of the 19 patients with hepatocellular carcinoma had alpha-foetoprotein levels greater than 100 x 10(3) IU/l. In contrast, only 5 of the 47 patients with cirrhosis showed values above 50 x 10(3) IU/l, demonstrating that this assay discriminates fairly well between hepatocellular carcinoma and cirrhosis. Data in agreement with the literature were obtained for testicular tumours; all of seven seminomatous tumours presented values below 5 x 10(3) IU/l, whereas 50% of the non-seminomatous tumours (n = 8) presented values above 20 x 10(3) IU/l.     

Serum lysophosphatidic acid concentrations measured by dot immunogold filtration assay in patients with acute myocardial infarction

In this study the relation between lysophosphatidic acid (LPA) and myocardial infarction was investigated, the typical and simplified methods for measuring serum LPA concentration by dot immunogold filtration assay (DIFA) based on a polyclonal antibody to LPA were developed, and serum LPA concentrations were measured in 31 patients with acute myocardial infarction (AMI) and 12 controls (blood donors) by DIFA. Serum LPA levels were raised more than twofold 8 h after the onset of AMI. Maximal elevation (10.43 mg/L) was found at 48-72 h following onset and remained higher than the control concentration (1.66 mg/L) 7 days after AMI. The rise in serum LPA concentration in AMI patients suggests that LPA might be involved in AMI-related pathophysiology in the cardiovascular system. The simplified DIFA developed in the present study for measuring serum LPA concentration is convenient and highly sensitive.     

Serum lysophosphatidic acid concentrations measured by dot immunogold filtration assay in patients with acute myocardial infarction

In this study the relation between lysophosphatidic acid (LPA) and myocardial infarction was investigated, the typical and simplified methods for measuring serum LPA concentration by dot immunogold filtration assay (DIFA) based on a polyclonal antibody to LPA were developed, and serum LPA concentrations were measured in 31 patients with acute myocardial infarction (AMI) and 12 controls (blood donors) by DIFA. Serum LPA levels were raised more than twofold 8?h after the onset of AMI. Maximal elevation (10.43?mg/L) was found at 48–72?h following onset and remained higher than the control concentration (1.66?mg/L) 7 days after AMI. The rise in serum LPA concentration in AMI patients suggests that LPA might be involved in AMI‐related pathophysiology in the cardiovascular system. The simplified DIFA developed in the present study for measuring serum LPA concentration is convenient and highly sensitive.     

Diagnostic value of thyrotropin concentrations in serum as measured by a sensitive immunoradiometric assay

A new, highly sensitive immunoradiometric thyrotropin (TSH) assay involving solid-phase-coupled monoclonal antibodies (Boots-Celltech Sucrosep IRMA-TSH) has been evaluated in a wide variety of patients with thyroidal and nonthyroidal illnesses and the results compared with those obtained by conventional diagnostic TSH RIAs. The sensitivity of the present assay ranged from 0.036 to 0.1 milli-int. unit/L (mean 0.056). TSH, measurable in serum of each of 128 euthyroid patients, ranged from 0.1 to 6.3 milli-int. units/L (mean 1.7, SD 1.1). Similar concentrations were found in 15 healthy pregnant women. TSH was undetectable in 27 hyperthyroid patients, of whom six were tested with thyroliberin stimulation and failed to respond. The mean TSH concentration measured in 62 seriously ill hospital patients of 2.7 (SD 2.5) milli-int. units/L was significantly higher (p less than 0.05) than in the euthyroid patients. Basal values and peak TSH responses to thyroliberin testing correlated well (r = 0.63, n = 48), irrespective of clinical diagnosis. We conclude that the present assay readily discriminates between euthyroid and hyperthyroid patients and should replace conventional TSH RIAs in diagnostic laboratories.     

Serum thyrotropin as measured with a one-step monoclonal-antibody radioimmunometric assay compared with two commercial radioimmunoassay kits

We compared results obtained with a commercial immunoradiometric assay kit for human thyrotropin in which monoclonal antibodies are used (Tandem -R TSH (ONE STEP) ImmunoRadioMetricAssay; Hybritech Inc.) with those obtained with two commercial radioimmunoassay methods: GAMMA-DAB [125I]HS-hTSH RIA (Clinical Assays) and Thyro-SHure TSH Diagnostic Kit (Nuclear Medical Laboratories). The correlation of all results obtained in the Hybritech assay with those of the two commercial RIAs exceeded 95%. Mean values for 100 euthyroid samples measured in the Hybritech assay were lower than for the other two methods. The separation between hyperthyroid and euthyroid patients was much clearer with the Hybritech assay than with the RIA methods. The Hybritech assay was far more sensitive than the Clinical Assays or the Nuclear Medical Laboratories assay.     

The experiences of new fathers during the first 3 weeks of life

Research has consistently demonstrated that the transition to parenthood is a stressful event. As well, the literature recognizes that the role of the father in North American society is in the process of change. The purpose of this qualitative study was to clarify our understanding of the experience of new fathers during the first 3 weeks postpartum. Twenty-two fathers were interviewed in their homes using a semi-structured interview format. Findings suggest that new fathers go through a predictable three-stage process during the transition to fatherhood. In addition, factors were identified which affect the transition. Nursing interventions were suggested to facilitate this process and implications for future study included.     

Clinical utility of an automated immunochemiluminometric thyroglobulin assay in differentiated thyroid carcinoma

BACKGROUND: Thyroglobulin (Tg) measurements are important in the follow-up of patients with differentiated thyroid carcinoma (DTC). We evaluated the analytical and clinical performance of a new automated immunochemiluminometric assay for Tg (Tg-ICMA; Nichols Advantage Tg; Nichols Institute Diagnostics). METHODS: We used the Tg-ICMA to measure Tg concentrations in serum samples from 110 Tg antibody-negative DTC patients undergoing thyroid-hormone suppression therapy. Disease state at the time of measurement was assessed on the basis of routine follow-up data. We compared the clinical performance of this assay with the routinely used IRMA (ELSA-hTG; CIS Bio International). RESULTS: The detection limit and functional sensitivity of the Tg-ICMA, based on direct calibration to CRM-457, were 0.05 and 0.6 microg/L, respectively. No Tg-IRMA-positive cases were missed by the Tg-ICMA. Tg was measurable by Tg-ICMA (0.6-8.6 microg/L) but undetectable by Tg-IRMA (<1.5 microg/L) in 12 patients (11%). Clinical data showed evidence of disease in 4 of 12 patients (33%). CONCLUSIONS: The Tg-ICMA is a sensitive and reproducible assay for identifying patients in follow-up for DTC with evidence of disease, but uncertainty remains with regard to interpreting findings of measurable serum Tg in patients with no evidence of disease. Follow-up data are required to determine the predictive value of these isolated Tg results. New concepts, i.e., serial Tg measurements and risk stratification of patients, need to be tested to confirm the applicability of this assay for clinical practice.     

Serum thyroglobulin in the monitoring of differentiated thyroid cancer

Abstract

Patients with differentiated thyroid cancer (DTC) usually have an excellent prognosis. Following surgical and radioiodine treatment to remove the cancer cells and suppressive doses of levothyroxine, long-term follow-up, including measurement of serum thyroglobulin (Tg) using a sensitive assay is required to detect recurrence. To interpret Tg results clinicians need to know the corresponding serum TSH concentration, have an appreciation of the clearance of Tg from patient serum following various interventions and the limitations of its measurement. The limitations of Tg immunoassay are well described and include potential interference from TgAb. For the majority of patients with DTC who are TgAb-negative, Tg measurement remains the most useful method of follow-up. For the TgAb-positive minority, interference and the possibility of producing erroneous results is a concern. Some assays are less badly affected than others and laboratories are advised to choose their assays carefully. Laboratories have sought to identify interferences using measurement of TgAb, lack of concordance between RIAs and immunometric assays and recovery of added Tg. More recently LC-MSMS assays to quantify Tg have been developed. They are not currently as sensitive as Tg immunoassays and it is likely these assays will, like immunoassays, be limited by Tg heterogeneity and standardization issues, although initial evaluations indicate that they may have value in the clinical setting as a second line test in antibody-positive DTC patients in whom Tg is unmeasurable by immunoassay.