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[en] The adaptation of human somatotropin and insulin assays to the automated Centria radioimmunoassay system [Clin. Chem. 21, 1305 (1975)] is reported. For the somatotropin assay, reaction conditions include a borate/bovine serum albumin buffer (pH 8.4) and a 20-h incubation at 40C. Assay results for clinical samples compared favorably (correlation coefficient = 0.930) with values obtained from a reference laboratory. The means determined for 92 patients' samples were 4.3 μg/liter (reference laboratory) and 5.1 μg/liter (Centria). Intra- and inter-run precision ranged from 3.2 to 15.9%. For the insulin assay, a phosphate/bovine serum albumin buffer (pH 7.4) is used, with a 20-h incubation at 40C. Previously analyzed insulin samples from a reference laboratory were determined by the Centria analyzer with excellent correlation (r = 0.965). Means for patients' samples were 43.0 milli (USP) units of insulin per liter (reference laboratory) and 47.5 milliunits of insulin per liter (Centria). In both assays an anionicexchange gel is used in the separation step. The criterion of parallelism, an indication of the validity of a radioimmunoassay, was satisfied in both assays. The Centria radioimmunoassay system offers the advantage of automating all the critical steps of these radioimmunoassays
[en] A methodological approach that permits, in a radioimmunoassay, the evaluation of rat insulin by using bovine insulin as reference is presented. As in general the technics for radioimmunoassay of different substances follow the same principles (competitive inhibition), we believe that the methodology presented here could be used for evaluation of other hormones when the adequated referential, of know biological activity, is not available. (Author)
[pt]Apresenta-se metodologia que torna possivel, em um radioimunoensaio, o uso de insulina bovina como referencial para avaliacao de insulina de rato. Como os radioimunoensaios utilizados para dosagem de diferentes substancias o mesmo principio (inibicao competitiva), acredita-se que a metodogologia apresentada possa ser utilizada tambem para avaliacao de outros hormonios, quando nao se dispuser do referencial adequado, de atividade biologica conhecida. (Autor)
[en] A soluble-phase proinsulin assay has been developed which does not require solid-phase antibody-binding. A human proinsulin standard curve is prepared in insulin-free and proinsulin-free plasma for comparison with unknown plasma samples. Proinsulin and insulin are bound with excess anti-insulin antiserum, and free C-peptide is removed by charcoal adsorption. The supernatant is then assayed using a routine C-peptide radioimmunoassay which utilises anti-C-peptide antiserum. The sensitivity of the assay (2 standard deviations above zero) is 9 pmol/L using 200 μL plasma sample. The assay is free from insulin cross-reactivity up to 100 mU/L and C-peptide up to 2000 pmol/L. Between-assay CV is 13% at 100 pmol/L. The assay has been used in subjects with hypoglycaemia of various aetiologies and has shown that a raised plasma proinsulin in the presence of hypoglycaemia can occur in sulphonylurea-induced and reactive hypoglycaemia as well as in insulinomas. After hyperglycaemic clamps at 7.5, 10 and 15 mmol/L glucose, type II diabetics both on and off sulphonylurea, were found to have lower proinsulin concentrations compared with normal subjects, commensurate with the diabetics' lower insulin responses. (author)
[en] (1) 125I labelling of insulin and the standardization of the labelled insulin for RIA use are well established. (2) A protocol of routine kit preparation is established. (3) Maximum two 100 tube kits can-be prepared from one batch. (4) The kits show steep dose-response curve. The expire date is usually 40 days after preparation.
[en] The assay system was set up in such a way as to increase the sensitivity of the reaction, reducing the tracer concentration from the usual 1.000 cpm/ml to 500 cpm/ml levels of radioactivity (specific activity aproximately 200 mCi/mg) and/or increasing the antiserum dilution by one thirty of the usual values in the assay of the final volume of incubation of 2,5 ml. (author)
[pt]O sistema de ensaio foi desenvolvido para aumentar a sensibilidade da reacao, reduzindo a concentracao do tracador dos 1.000 cpm/ml usuais para niveis de radioatividade de 500 cpm/ml (atividade especifica aproximadamente 200 mCi/mg) e/ou aumentando a diluicao do anticorpo 30 vezes o valor usual no ensaio de volume final de incubacao de 2,5 ml. (autor)
[en] The present study was undertaken to evaluate the significance of the insulin and the C-peptide response to oral glucose loads in normal and diabetic subjects and to establish the effects of the obesity. In this study, the authors have measured plasma insulin and C-peptide by means of radioimmunoassay in 10 nonobese normal, 5 obese normal, 13 nonobese moderate diabetic patients, 9 obese moderate diabetic patients and 9 severe diabetic patients. The results obtained were as follows; 1) In 10 nonobese normal subjects, the plasma insulin level at fasting state and at 30, 60, 90, and 120 min after oral glucose loads were 15.7±3.4, 48.3±9.8, 40.4±6.7, 37.4±6.5 and 26.0±4.2 uU/ml (Mean±S.E.) and C-peptide were 1.9±0.3, 3.9±0.6, 6.3±0.6, 5.7±0.5 and 4.0±0.5 ng/ml. The change of C-peptide was found to go almost parallel with that of insulin and the insulin value reaches to the highest level at 30 min whereas C-peptide reaches to its peak at 60 min.. 2) The plasma insulin level in 5 obese normal subjects were 38.9±12.3, 59.5±12.3, 59.2±17.1, 56.1±20.0 and 48.4±17.2 uU/ml and the C-peptide were 5.5±0.4, 6.8±0.5, 7.9±0.8, 7.9±0.8 and 7.8±2.0 ng/ml. The insulin response appeared to be greater than nonobese normal subjects. 3) In 13 nonobese moderate diabetic patients, the plasma insulin levels were 27.1±4.9, 44.1±6.0, 37.3±6.6, 35.5±8.1 and 34.7±10.7 uU/ml and the C-peptide levels were 2.7±0.4, 4.9±0.7, 6.5±0.5, 7.0±0.3 and 6.7±1.0 ng/ml. There was little significance compared to nonobese normal groups but delayed pattern is noted. 4) In 9 obese moderated diabetic patients, the plasma insulin levels were 22.1±7.9, 80.0±19.3, 108.0±27.0, 62.0±17.6 and 55.5±10.l uU/ml and the C-peptide levels were 5.2±0.4, 8.0±1.0, 10.4±1.6, 10.4±1.7 and 10.1±1.0 ng/ml and its response was also greater than that of nonobese moderate diabetic patients. 5) The plasma insulin concentrations in 9 severe diabetic subjects were 8.0±3.8, 12.1±3.5, 16.8±4.6, 19.6±5.2 and 15.0±5.0 uU/ml and the C-peptide levels were 1.6±0.3, 2.4±0.4, 4.1±0.6, 4.0±0.8 and 4.5±0.7 ng/ml and the insulin and C-peptide responses were markedly reduced in severe diabetic groups. 6) There were significant differences between each groups of patients on the magnitude of total insulin or C-peptide areas, the insulinogenic index and the C-peptide index.
[en] Plasma glucose levels before and after oral glucose administration have been compared in g group of 76 thyrotoxic subjects and a group of 8 normal control subjects in order to study the effect of glucose loading in thyrotoxicosis. Following were the results: 1) The mean fasting plasma glucose level was elevated in thyrotoxic group (95.5 mg%) compared to normal control group (88 mg%). 2) The peak of glucose tolerance curve is at 30 minutes after glucose administration in both groups, but its mean value was 44 mg% higher in thyrotoxic group than in control group. 3) The plasma glucose levels returned towards the fasting level in the later stage of the test more rapidly in thyrotoxic group than in control group. 4) 69.6% of oral glucose tolerance tests were impaired in the thyrotoxic group, and the occurrence of abnormal glucose tolerance could be related to the degree of thyrotoxicity, sex and age. 5) The mechanisms of the impaired glucose tolerance in thyrotoxicosis are thought to be related to an increased rate of glucose absorption from gastrointestinal tract, abnormal liver function with decreased hepatic glycogenesis, increased glucose oxidation, decreased pancreatic release of insulin, and genetic relationship between diabetes and thyrotoxicosis.