Rapid radioimmunoassay for anabolic steroids in urine.
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Biomedical subjects
Publications and source records attributed to V A Rogozkin.
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The purpose of this investigation was to examine the effects of training either once or three times/day on the principal stages of skeletal muscle. The experiments were carried out on male albino rats fed either 3 or 5 times/day a diet containing 20% protein. Experimental animals swam either once or 3 times/day, 6 days/week for 10 weeks with weights attached. The total duration of daily activity was equal for both groups and at 10 weeks each animal was swimming for 60 min/day with 3% of his body weight attached. All the animals were examined at rest after the 10-week training programs. The adequacy of the weight-loading and training schedules was estimated by body weight dynamics and such energy metabolites as creatine phosphate and glycogen. Skeletal muscle RNA and protein synthesis were studied by means of 14C-orotic acid and 14C-leucine incorporation, respectively. Quadriceps and gastrocnemius muscles were used for analysis in all experiments. It was found that the increase in the number of daily training sessions resulted in an increased content and intensity of synthesis of skeletal muscle proteins as evidenced by an increase in the content of amino acids in the muscle, an increased synthesis of both microsomal and ribosomal RNA, an increased stability of (poly-A)-containing messenger RNA, and an increased synthesis of all skeletal muscle protein fractions: myofibrillar, sarcoplasmic, and myostromal.
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4hr incubation of the growing culture of chick embrio myoblasts in the presence of 5 mM creatine resulted, regardless of a well-defined lowering of cell membrane permeability to labelled precursors, in: (1) the 1.5-fold induction of 14C-orotic acid incorporation into total cellular RNA; (2) the 1.9-fold stimulation of DNA-dependent RNA-polymerase activity and (3) the preferable, in comparison with total proteins, 14C-leucine incorporation in the myosin heavy chain. The stimulating effect of creatine on RNA-poly-merase deals with a definite enzyme fraction and it is caused by the augmentation of the transcriptional complex enzyme activity. Polyacrilamide gel electrophoresis of total 14C-RNA revealed a relative increase of radioactivity after creatine treatment in both high-molecular pike and the fraction corresponding to 26S RNA, the relative proportion of mRNA containing Poly-A in the composition of total cellular RNA being invariable. The data obtained show that, though creatine as a positive regulator manifestates its effect at different stages of the sucessive chain of reactions leading to muscular protein synthesis, the direct realization of creatine regulatory function is connected with nucleus and is fullfilled at the transcription level.
It has been established that the activity of calpains and the amount of neutrophil myeloperoxidase in skeletal muscles of rats increases after the 24-48 h-rest, that followed an intensive single physical exercise. Adaptation which appears as a result of systematic physical exercises causes less pronounced changes in the activity of proteolytic enzymes. Free ubiquitin content in skeletal muscles remains unchanged during training and in posttraining periods.
The immunoradiometric assay of rat myeloperoxidase (MPO) was developed based on the usage of the CNBr-activated sepharose as a solid phase for antiMPO and of [125I] IgG (the affinity preparation of antiMPO) as a label. The analysis lasted for 8 h and standard curve limits were 0.5-500 ng per tube. Intra and inter assay coefficients of variation did not exceed 3% and 6.5%, respectively. It was shown by the above method that the intensive swimming of male rats led to an increase in the MPO concentration in blood plasma and to its decrease in neutrophils.
Radioimmunoassay (RIA) of human lysozyme was developed using 125I-lysozyme and specific antiserum to the enzyme. Titre of antibodies produced in the antiserum was as high as 1:81,000, affinity constant constituted 1.3.10(9) M/L. Optimum patterns of the RIA were achieved under the following conditions: B0 = 50-55%, T = 25-30.10(3) cpm, BSA = 0.5%, NaCl = 0.85%, charcoal 2.5%. The antibody-antigen reaction was carried out within 30 min, 30 and 15 min at 37 degrees, 20 degrees and 4 degrees, respectively. The RIA enabled to estimate lysozyme with sensitivity of 0.25 ng/ml, while the standard curve was optimal within the range of 0.1-1.0 ng/tube. Intra- and inter-assay coefficients did not exceed 4% and 8%, respectively. Recovery of the standard lysozyme concentration was equal to 96.5 +/- +/- 0.8%. As shown by the RIA intensive physical exercises resulted in an increase of lysozyme content in blood, which appears to occur due to the enzyme secretion from phagocytes. Calculation of the secretion index may be used for evaluation of blood phagocytes functional state.