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OBJECTIVES: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods that allow accurate quantitation of thyroglobulin (Tg) in the presence of Tg antibodies (TgAbs) have recently become available. Due to cost differences between LC-MS/MS and immunoassay, some laboratories now offer a reflex test strategy that uses LC-MS/MS only for TgAb-positive samples. The goal of this study was to examine utilization of Tg testing strategies and cost savings. METHODS: Test ordering patterns were examined for over 150,000 orders for TgAb and Tg in our laboratory. The average list price was determined from three separate commercial laboratories offering this testing. RESULTS: Data showed that 89% of orders for Tg used the reflex test option, resulting in a savings of over $3 million compared with testing all samples by LC-MS/MS. Of the Tg by LC-MS/MS orders not using the reflex option, 1,663 also included a separate order for TgAb on the same patient sample, representing approximately $170,000 in potentially unnecessary costs from TgAb-negative samples. CONCLUSIONS: Identifying situations to use more expensive testing methods (eg, LC-MS/MS) only when necessary, such as for TgAb-positive patients, leads to considerable cost savings and a more economical use of valuable health care resources.

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The concentration of progesterone was measured in 60 plasma samples from bitches at various stages of the oestrous cycle, using commercially available quantitative and semi-quantitative ELISA test kits, as well as by two commercial laboratories undertaking radioimmunoassay (RIA). The RIA, which was assumed to be the 'gold standard' in terms of reliability and accuracy, was the most expensive method when analysing more than one sample per week, and had the longest delay in obtaining results, but had minimal requirements for practice staff time. When compared with the RIA, the quantitative ELISA had a strong positive correlation (r=0.97, P<0.05) and a sensitivity and specificity of 70.6 per cent and 100.0 per cent, respectively, and positive and negative predictive values of 100.0 per cent and 71.0 per cent, respectively, with an overall accuracy of 90.0 per cent. This method was the least expensive when analysing five or more samples per week, but had longer turnaround times than that of the semi-quantitative ELISA and required more staff time. When compared with the RIA, the semi-quantitative ELISA had a sensitivity and specificity of 100.0 per cent and 95.5 per cent, respectively, and positive and negative predictive values of 73.9 per cent and 77.8 per cent, respectively, with an overall accuracy of 89.2 per cent. This method was more expensive than the quantitative ELISA when analysing five or more samples per week, but had the shortest turnaround time and low requirements in terms of staff time.

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Bisphenol A (BPA) is widely used in the manufacturing of polycarbonate plastic food and drink packaging. Possessing a weak estrogenic activity, BPA is listed among a growing list of endocrine disrupting compounds. In this study, a polyclonal anti-BPA antibody was obtained by immunization with BPA-monocarboxymethylether covalently linked to BSA. The antibody demonstrates negligible cross-reactivity with most analogous BPA phenolic structures, and no cross-reactivity with endogenous steroids. An extraction step with ethyl acetate minimized matrix effects and allowed the BPA measurement in plasma and other biological samples. Recovery after loading test was 96 +/- 4% and dilution tests had a linear profile (r2 > 0.93). The limit of detection of the BPA RIA was 0.08 microg L(-1) with an IC50 of 1.25 microg L(-1). The intra- and inter-assay coefficients of variation were 5.6 and 8.6%, respectively at a BPA concentration of 0.7 microg L(-1) and 6.9 and 5.7% at a BPA concentration of 1.3 microg L(-1). A significant correlation was found between the values obtained by the RIA and HPLC-MS (r2 = 0.92) or HPLC coupled to a fluorescence detector (r2 = 0.80). In conclusion, we described a BPA-RIA that is a suitable tool for evaluating human exposure to BPA.

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Cyclic AMP (cAMP) is an important signal transduction second messenger that is commonly used as a functional mirror on the actions of G protein-coupled receptors that can activate or inhibit adenylate cyclases. A radioimmunoassay for cAMP with femtomole sensitivity was first reported by Steiner more than 30 years ago, and there have been several subsequent modifications that have improved this assay in various ways. Here we describe additional improvement to existing methods that markedly improve speed and reduce cost without sacrificing sensitivity, and is also adaptable to analysis of cGMP. The primary antibody is coupled directly to magnetic beads that are then separated from unbound marker using filtration on microplates. This eliminates the need for a secondary antibody, and markedly increases throughput. In addition, we report a simple, reproducible, and inexpensive method to make the radiomarker used for this assay. Although still requiring the use of radioactivity, the resulting method retains a high degree of accuracy and precision, and is suitable for low-cost high throughput screening. Use of aspects of this method can also improve throughput in other radioimmunoassays.

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BACKGROUND: There are documented associations between elevated maternal corticotropin-releasing hormone (CRH) levels and adverse pregnancy outcomes. However, reports of these findings often lack sufficient detail and rationale regarding the bioassay methodology. This shortcoming can be problematic for researchers who do not possess in-depth laboratory sciences knowledge but who want to include bioassays in their investigations or to evaluate published reports. The quality and reliability of CRH measurement results can be significantly affected by variables encountered during sample collection, processing, storage, and bioassay. Thus, it is important to establish research laboratory protocols that are based on well-informed rationales and to carefully consider and control for relevant variables. APPROACH: A synthesis of laboratory sciences literature regarding variables affecting CRH measurement in pregnancy is presented. Additionally, consultation with experienced researchers provided an in-depth understanding of CRH measurement. From these sources, a laboratory protocol for clinical research was developed. RESULTS: Multiple variables that are specific to the reliability of CRH measurement in pregnancy have been identified. These include sample collection methods, sample processing, sample integrity, sample storage, and the actual assay selected. CONCLUSION: The reliability of CRH measurements can be significantly improved by identifying and controlling for variables encountered during sample collection, processing, storage, and bioassay. Adequate methodological details are difficult to glean solely from the published literature, thus consultation with well-informed researchers is necessary. A protocol for CRH bioassay in clinical research is proposed.

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Iron deficiency anaemia and the thalassemia syndromes, a group of disorders resulting from inherited abnormality of globin chain production, are common causes of anaemia in Thailand, and in the Far East in general. Monitoring erythropoiesis in these patient is very important in evaluating the disease and the response to treatment. Recently, our group has just reported the feasibility in using serum erythropoietin (EPO) for monitoring purposes in thalassemia and demonstrated that the determination of serum EPO can be an alternative choice in the follow up of these conditions. This study reports a cost effectiveness analysis comparing the recently reported radioimmunoassay (RIA) for serum EPO determination and the previously used tool, the reticulocyte count. The study reports a higher detection rate for ineffective erythropoiesis when using serum EPO determination, however, the increased sensitivity is at balanced by a higher unit cost. In this analysis, the cost effectiveness for serum EPO and reticulocyte are 208.33 and 50 baht/detection, respectively. () Therefore, the reticulocyte count is more cost effective and is recommended for routine usage in our current medico-economic setting.

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Studies of the role of leptin in patients with anorexia nervosa and bulimia nervosa have conflicted in their data and interpretation. Such differences may be a result of the assay methods used or the way results are compared with those from normal controls. To investigate these possibilities, we analyzed serum leptin levels in anorexic, bulimic, obese, and control individuals, thereby spanning the full range of human body weights, using three frequently employed commercial kits. Kits from Linco (St Louis, MO) and DSL (Webster, TX) employ a radioimmunoassay method, and the R&D Systems kit (Minneapolis, MN) uses an enzyme-linked immunosorbent assay. We found that the three kits provide results that are highly linearly correlated with each other and remarkably linearly related to percent ideal body weight (%IBW) over more than three orders of magnitude (Linco, r = 0.90; R&D, r = 0.87; DSL, r = 0.86). For very low leptin levels, the more sensitive kits from R&D and Linco appeared to give more reliable results. Measurement method does not appear to explain the literature conflicts. We found that patients with anorexia nervosa have serum leptin values that lie above the line extrapolated from the %IBW/leptin curve generated from analysis of all non-anorexic patients. Therefore, in anorexia nervosa, inappropriately high leptin levels for %IBW may contribute to a blunted physiologic response to underweight and consequent resistance to dietary treatment. By contrast, most bulimic patients have leptin levels significantly below those predicted from the same %IBW/leptin curve. The relative leptin deficiency in bulimic subjects may contribute to food-craving behavior. We propose that using the %IBW/ leptin curve can facilitate identification of true pathophysiologic abnormalities in eating-disordered individuals and provide a basis for the design of therapeutic interventions or monitoring of response to treatment.

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PURPOSE: Carcinoembryonic antigen (CEA) is still a widely used test for monitoring breast cancer, although recent reports discourage its routine use because of low sensitivity. This is a prospective study evaluating the efficacy of CEA and CA 15.3 in monitoring breast cancer. EXPERIMENTAL DESIGN: Serum CEA and CA 15.3 were measured in 2191 patients with either benign (n = 738) or malignant (n = 1453) breast diseases. Five hundred and forty-nine patients were monitored during postsurgical follow-up for either a minimum of 5 years or until time of recurrence. Fifty-three patients with metastases were also monitored during chemotherapy. RESULTS: Elevated CEA and CA 15.3 levels were found in 16.7% and 33.0% of patients, respectively. CEA sensitivity rose to 41.3% and CA 15.3 sensitivity rose to 80.8% in metastatic patients. The adjunct of CEA increased the CA 15.3 sensitivity by 6% in the overall population and by only 2.1% for patients with metastases. During postsurgical follow-up, CEA was elevated in 38.0% and CA 15.3 in 70.2% of patients with recurrence. The combination of CEA and CA 15.3 increased the overall sensitivity by only 1.4%. Longitudinal monitoring of 53 metastatic patients undergoing chemotherapy demonstrated that, when positive, both CEA and CA 15.3 paralleled response to treatment, although CA 15.3 was a significantly more powerful marker for determining response to treatment. The cost effectiveness ratio of CEA was clearly less favorable than that of CA 15.3. CONCLUSIONS: CEA monitoring should be considered an expensive and inefficient method of follow-up evaluation for breast cancer patients, and it provides no additional value when used in combination with CA 15.3.

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Serial monitoring by plasma progesterone measurement is advised in the literature for fertility work-up, to detect ovulation disturbances in women presenting with regular menstrual cycles. Three strategies to diagnose such 'subtle ovulation disorders' (SOD, defined as anovulation, inadequately timed ovulation or ovulation of a follicle of reduced size in regularly cycling women) were evaluated, in order to investigate costs of such a diagnosis. On the basis of a 'maximal', an 'ultrasound-only', and a 'preselection' strategy, total medical costs and costs including non-medical costs were calculated for each SOD diagnosis. A 'maximal' diagnostic strategy resulted in a total medical cost of ECU 9057 per diagnosis (including non-medical costs ECU 12,787); an 'ultrasound-only' strategy in ECU 4520 (ECU 6791) per diagnosis. By use of a 'preselection' strategy, 4.25% of the women were found to have an SOD, at a cost of ECU 3036 (ECU 6868) for each diagnosis. As the real significance of SOD diagnosis for the prognosis of the patient to become pregnant without treatment remains unclear, and as no randomized trials on treatment effectiveness have as yet been undertaken, it is questionable whether this approach is worthwhile.

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The immunoassay is one of the most sensitive and reliable analytical techniques available in the clinical laboratory. The original label for immunoassays was radioisotopes, and these methods, radioimmunoassay (RIA) and immunoradiometric assay (IRMA) are still the reference methods, because of invulnerability of the radioactive emission with respect to environmental interference. Labels other than radioisotopes have been tested for use in immunoassay to improve the sensitivity and reliability and to avoid some of the disadvantages of radioisotopic techniques. New labels have continued to be developed (Horseradish peroxidase-HPR-, pyrophosphatase, luciferases, pyrodopirazines, europium cryptates, porphirins, phosphors) and new label detection methods have been set up (e.g. chemiluminescence assay, thermometric assay, NADP+ and FADP- based coupled assay). New immunoassay strategies such as simultaneous multianalyte automated test have been developed and the reliability of the assays has in some cases caused division among researchers about the choice between the radioisotopic immunoassay or the non-radioisotopic immunoassay, as considerable effort and investment had been devoted to the search for more sensitive and practicable tests than the classic RIA-IRMA methods. The evolution of immunoassays (Monoclonal Antibodies, non-radioactive tracers, automation) has produced systems which allow a large number of laboratories to determine a great number of analytes with very good practicability. The availability of fully automated systems has generated the opinion that analytical performance of immunoassays can be considered similar to that of many traditional parameters of clinical chemistry. This conclusion seems however too optimistic, in fact data collected from interlaboratory studies demonstrate that problems concerning the analytical reliability of the measurements still remain not completely solved. In the authors' opinion, this opposition between immunological assay based on isotopic or non-isotopic labels is misleading, because each assay (whether it uses isotopic, enzymatic, fluorimetric or luminescent labels) has its own analytical characteristics and performance. For this reason the term "alternative", used to indicate all non-isotopic assays as a unique class of tests, should be abandoned. From a theoretical point of view the choice should not be between isotopic and non isotopic techniques. For each analyte to be tested, it is advisable to use the immunological assay that suits the requirements of the laboratory, irrespective of type of label. From a practical point of view, the choice should be based on the analytical performance and on the characteristics of each assay, on its cost and the type of instrumentation available in the laboratory, and on the experience and the knowledge of the laboratory personnel.

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OBJECTIVES: The study was designed to assess the reliability of measurement of 24-hour urinary 17 alpha-hydroxyprogesterone (17-OHP) by radio-immunoassay (RIA) as an alternative biochemical assessment for monitoring the treatment of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD) and to assess the need for sample purification by column chromatography to improve assay specificity. METHODOLOGY: Morning serum 17-OHP was measured using RIA and 24-hour urinary pregnanetriol using gas chromatography. Twenty-four-hour urinary 17-OHP was measured in samples from 17 prepubertal patients with CAH due to 21-OHD, and 20 normal prepubertal children as controls. In 24 urine samples, RIA of 17-OHP was performed with and without column chromatography. RESULTS: There was a good correlation between 24-hour urinary 17-OHP and 24-hour urinary pregnanetriol (r = 0.962, P < 0.01) and between 24-hour urinary 17-OHP and morning serum 17-OHP (r = 0.955, P < 0.01). There was no significant difference in the RIA of the urine samples with and without purification by column chromatography. CONCLUSIONS: The measurement of 24-hour urinary 17-OHP is a reliable alternative for the biochemical monitoring of 21-OHD, and RIA specificity is unaffected by omission of column chromatography.

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To determine whether the new immunometric (sandwich) assays for gonadotropins achieve the alleged improvements over RIAs, we compared 13 commercial kits for LH and FSH with our established and validated polyclonal RIAs. Although the kits claimed detection limits as low as 0.05 IU/L, when sensitivity was tested with a uniform hormone standard (the Second International Reference Preparation of human menopausal gonadotropin urinary standard) and the requirement that the limit must be determined from a detectable standard point rather than the variance around zero, only 4 kits surpassed the detectability achieved by the existing LH and FSH RIAs. Seven of the 10 LH kits tested displayed greater than 10% cross-reactivity with either the alpha- or LH beta-subunit. The relationship between the kit standard and the Second International Reference Preparation of human menopausal gonadotropin uniform standard in each kit was nonlinear, so that attempts to compare results between assays with simple multiplication factors are inaccurate. Only 2 assays were designed to eliminate a hook effect. The following conclusions were reached. 1) Most available gonadotropin immunometric assays do not yet offer features not already available in existing validated polyclonal research assays. 2) Conversion of data from one assay to another is not straightforward. 3) Many of the manufacturers' claims do not appear justified. 4) Determination of the detection limit and other immunometric assay characteristics requires standardization of criteria. We propose the following minimum criteria for validating gonadotropin immunometric assays: 1) a uniform definition of the detection limit based on the lowest distinguishable standard concentration, 2) determination of the standard concentration at which a hook occurs, 3) determination of cross-reactivity to subunits as well as intact gonadotropins, and 4) establishment of an adequate normative data base.

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Slight albuminuria predicts clinically significant nephropathy in patients with insulin-dependent diabetes mellitus (IDDM) and early death in patients with non-insulin-dependent diabetes mellitus (NIDDM). This study compares four commercially available methods for measuring low concentrations of urinary albumin. We tested random spot urine specimens from 50 nondiabetic volunteers and 100 diabetic patients. This specimen was chosen to simplify collection in an outpatient setting. Two screening methods were evaluated for their ability to detect urinary albumin in the range of 15 to 200 mg/L. Sensitivity and specificity were 100% and 54.7%, respectively, for the Ames Micro-Bumintest; and 95.5% and 91.5%, respectively, for the Sclavo Albumin Screen. The high number of false-positive results made the Micro-Bumintest unacceptable. The Albumin Screen yielded fewer false-positive results, but also produced some false-negatives. Two quantitative methods, a radioimmunoassay (RIA) and a turbidimetric assay (the SPQ Microalbumin), yielded results that agreed well with each other.

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We adapted a commercial serum cortisol radioimmunoassay (RIA) kit for use with saliva specimen. Using 50 microliters sample volume, the lower sensitivity was found to be 0.02 microgram/dl with intraassay variation coefficients between 5.4 and 8.9% at different concentrations. The 50% intercept was either 0.5 or 0.26 microgram/dl (50 or 100 microliters standard/sample volume). Fifty-four early morning samples from healthy adults showed absolute concentrations which are closely comparable to respective data from other laboratories. A comparison of 35 saliva samples which were each assayed with the adapted RIA as well as with three other commercial kits revealed high correlations between these assays (r = .94 to r = .97). Data on salivary cortisol responses to CRH stimulation and dexamethasone suppression in healthy subjects further the validity of the assay results. The most important contribution of this assay modification, however, is thought to be its impact on analysis costs: The protocol presented in this paper allows for reliable salivary cortisol measures with a reduction of costs for analytical material to 25% compared to serum determinations.

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CA 19-9 is a promising radioimmunoassay for the detection of pancreatic cancer, but its clinical role and cost-effectiveness are not yet known. To investigate these factors, we used clinical decision analysis to study diagnostic strategies for patients with suspected pancreatic cancer presenting as pain or weight loss. Comprehensive diagnostic strategies were developed to reflect current and future patterns of practice utilizing CA19-9 radioimmunoassay (RIA) to yield biopsy-proved cancer or confidently exclude its presence. The performance of the strategies beginning with CA19-9 RIA and ultrasonography were equivalent in positive and negative predictive values over a range of prevalence of pancreatic cancer from 0.02 to 0.15. At higher prevalence, the negative predictive value of the ultrasonography strategy became significantly better. The CA19-9 RIA strategy used fewer noninvasive tests, endoscopic retrograde cholangiopancreatographic procedures, and invasive radiologic studies than did the ultrasonography strategy at each prevalence. The health care costs ranged between $848 and $1413 per patient for the CA19-9 RIA strategy and $1186 and $1848 per patient for the ultrasonography strategy. We conclude that the CA19-9 RIA is a useful, cost-effective initial test for the examination of patients with suspected pancreatic cancer.

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Quality assurance testing represents a substantial proportion of the clinical laboratory budget, but current guidelines are based on criteria that pertain to analytic error rather than to optimization of the cost-effectiveness of patient care. A general Bayesian mathematical model for the cost-effectiveness of assay quality control has been developed, and is demonstrated using previously published data. The cost-effectiveness of quality assurance as defined here depends upon the prevalence of disease, the shapes of the distributions of test results observed in the non-diseased and diseased populations, the decision limit selected for labeling results positive or negative, the costs and benefits associated with each of the possible therapeutic outcomes, the magnitude of random and systematic analytical errors, the statistical power of the quality control test in use, the costs associated with delays due to re-assay, and the proportion of total test cost attributable to quality control procedures. Given current clinical laboratory practice, much of this information will not be routinely available. The model combines these factors into a simple equation with three terms: one for the cost of the original and any required repeat laboratory analyses, one for the cost of delay entailed by the rejection of an assay batch, and one for the change in total costs consequent to rejection of erroneous assay results.

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