testosterone ria ct ref ks24ct - Radim S.p.A.

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RADIOIMMUNOASSAY FOR QUANTITATIVE DETERMINATION OF TESTOSTERONE IN HUMAN SERUM FOR IN VITRO DIAGNOSTIC USE ONLY CLINICAL APPLICATIONS Testosterone (M.W.: 288.4) is an androgen steroid of 19 carbon atoms. Mainly, it is synthesized in the testes by Leydig interstitial cells, from 5-pregnenolone and progesterone, by following both Δ4 and Δ5 metabolic pathways. The Δ5 pathway seems to be predominant in the testes. A small amount is produced by the reticular adrenal cortex, with Δ4-Androstenedione and DHEA as precursors. In the blood it is carried by a beta-globulin. About 97-99% of circulating testosterone is bound to this carrier protein; the remaining free fraction is considered to be biologically active. It is then excreted in the form of glucuronides or else converted into Δ4-androstenedione which is then further metabolized, the main catabolites being androsterone, etiocholanolone and epiandrosterone. It can also turn into epitestosterone, an important catabolite acting on the hypothalamus as well as dihydrotestosterone (DHT), by a 5-α-reductase. DHT is more potent than testosterone on target cells. Testosterone determines the changes which are typical in the various pubertal stages, by stimulating the development of the secondary sexual features (voice, height, body hair). It also acts on the penis, scrotum, seminal vesicles and prostate. It enhances protein synthesis, makes the nitrogen, phosphorus as well as potassium balance positive and stimulates the germinal epithelium of hair follicles. It is involved in the differentiation of sexual behavior and regulates normal spermatogenesis. Moreover there is a hypothalamic-pituitary negative feedback mechanism working on FSH and LH gonadotropins. In males, testosterone levels are evaluated in cases of: hypogonadism, testicular neoplasia and Klinefelter syndrome. In females, testosterone levels are significant in cases of: ovarian neoplasia, hypertrichosis and Stein- Leventhal syndrome. PRINCIPLE OF THE ASSAY The present method is based on a competition between labeled antigens (radioactive conjugate) and non-labeled antigens (calibrators, samples) for binding to a number of specific sites of the antiserum coated tubes. After the incubation, the liquid in the tubes is removed by aspiration and the radioactivity is measured in a gamma counter. REAGENTS PROVIDED WITH THE KIT: PREPARATION AND STABILITY − The reagents are sufficient for 100 tubes. − Store the kit at 2-8°C. − The expiry date of each reagent is shown on the vial label. CT Coated Tubes: 100 tubes (4x25) coated with rabbit polyclonal anti-Testosterone antibody. Unused tubes should be stored at 2-8°C in the appropriate bag and accurately sealed. CAL Calibrators: 6 vials of Testosterone in serum matrix: 1 vial of Zero Calibrator (2 mL) and 5 vials (1.5 mL) at the following concentrations: 0.25, 0.5, 1.5, 6.0 and 15.0 ng/mL. Ready for use. Conversion: 1 ng/mL = 3.467 nmol/L. Preservative: NaN3 (< 0.1%). NOTE: refer to the C.Q. sheet, for exact concentrations. CONJ I 125 Radioactive Conjugate: 1 vial (50 mL) of 125 I-Testosterone in phosphate and protein buffer. Radioactivity contents: 77.7 KBq. Ready for use and red colored. Preservative: NaN3 (< 0.1%). CTR Control Serum: 1 vial of Testosterone in animal serum. Preservative: Thimerosal (< 0.05%). Lyophilized. Reconstitute with 2 mL of distilled H2O. After reconstitution, store at 2-8°C for 2 weeks; for longer periods freeze at -20°C. NOTE: refer to the C.Q. sheet, for exact concentrations. MATERIAL REQUIRED BUT NOT SUPPLIED − Plastic test tubes (12 x 75 mm). − Test tube racks. − Adjustable, automatic micropipettes with disposable tips. KS24CT TESTOSTERONE RIA CT M104 - Rev.10 – 09/2007 - Pag. 8/32
− Vortex mixer. − Water-bath, adjustable at 37°C. − Aspiration pump or automated tube washing device. − Scintillation gamma counter. − Logit-log or semi-log graph paper. − Distilled H2O. RADIOACTIVE MATERIAL Radionuclide: 125 I ITALIAN DECREES (D.Lgs. 230/95 – D.Lgs. 241/00 art. 19) AND RULES OF PREVENTION AND SECURITY In order to obtain correct and reproducible results, the following rules must be observed: − Do not mix reagents of different lots. − Do not use reagents beyond their expiry date. − Do not store or leave reagents and samples at high temperatures or areas of possible contamination. − Use thoroughly clean glassware, free from metal ion contamination or oxidizing substances. − Use distilled or deionized water, stored in perfectly clean containers. − Carefully avoid any contamination among samples; for this purpose, disposable tips should be used for each sample and reagent. − Do not modify the "Assay Procedure", in any way. Altering: • volume of reagent added • exact temperature and incubation times may cause incorrect clinical results. − Reconstitute lyophilized reagents, if present, as described on the relative labels. Any deviation in reagent use or wrong volumes, may affect the reliability of results obtained. − In case of manual procedure, it is important to use calibrated pipettes and have appropriate technical manuals. Primary importance is a good precision preparing and dispensing the reagents. Ensure that all the equipment used is in perfect working order, has been correctly calibrated and is regularly maintained. − Ensure that all the equipment used (glassware, incubators, shakers, tube washers, gamma counter and fridge/freezers used for reagent and sample storage) is in perfect working order, has been correctly calibrated and is regularly maintained. Any deviation from the correct use of the equipment listed can produce errors in the methodology, this may affect the reproducibility and reliability of results obtained. − Utilise a suitable method for the correct identification of patient samples. Incorrect identification may cause a specificity losses of the system and wrong clinical results. In order to reduce physical, biological and chemical risks, the following precautions must be observed: − Use protective individual articles (ex.: disposable gloves, lab coats, etc.) while handling radioactive materials and/or potentially infectious material as well as during the assay. − Do not pipette reagents by mouth. − Do not smoke, eat, drink or apply cosmetics during the assay. − All radiological work should be done in a designated area. − Radioactive materials should be stored in their original container in a designated area. − A record book for logging receipt and disposal of all radioactive materials should be kept. − Any radioactive spills should be taken care of immediately in accordance with established procedures. − All material of human origin used for the preparation of this kit was tested negative for HBsAg, anti- HIV and anti-HCV. Since no test at present can guarantee complete absence of these viruses, all samples and reagents containing biological material used for the assay must be considered potentially infectious. KS24CT TESTOSTERONE RIA CT M104 - Rev.10 – 09/2007 - Pag. 9/32
Page 1 and 2: TESTOSTERONE REF KS24CT Italiano p.
Page 3 and 4: DOSAGGIO RADIOIMMUNOLOGICO PER LA D
Page 5 and 6: − I materiali di origine umana ut
Page 7: Siero Media ± D.S. C.V. Replicati
Page 11 and 12: The values shown below must be cons
Page 13 and 14: RADIO-IMMUNASSAY ZUR QUANTITATIVEN
Page 15 and 16: − Entsprechend der italienischen
Page 17 and 18: (ng/mL) % Nr. a 0.53 ± 0.0327 6.1
Page 19 and 20: N.B.: vérifier toujours l’exact
Page 21 and 22: échantillon et par interpolation s
Page 23 and 24: RADIOINMUNOENSAYO PARA LA DETERMINA
Page 25 and 26: − Es necesario eliminar inmediata
Page 27 and 28: Repetibilidad (intra-ensayo) Suero
Page 29 and 30: RCNS H2O Ricostituire con / reconst
Page 31 and 32: Schema del Dosaggio, Assay Scheme,

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