Prakash B S, Metten M, Schams D, Wuttke W
Abteilung Klinische und Experimentelle Endokrinologie, Frauenklinik Goettingen, Georg-August Universitaet, Germany.
Anim Reprod Sci. 1998 May 15;51(3):185-94. doi: 10.1016/s0378-4320(98)00069-4.
A highly sensitive and specific second antibody enzymeimmunoassay (EIA) on microtiterplates for oxytocin determination in bovine plasma using the biotin-streptavidin amplification system was developed. Biotin was coupled to oxytocin and used to bridge between streptavidin-peroxidase and the immobilized oxytocin antiserum in the competitive assay. The assay was carried out directly in 200 microliters of bovine plasma. Oxytocin standards prepared in hormone-free plasma were used. The sensitivity of the assay was 0.25 pg/well which corresponded to 1.25 pg/ml plasma; the 50% relative binding was seen at 2.8 pg/well. Plasma volumes for the assay ranging from 50 to 200 microliters did not influence the shape of the oxytocin standard curve; however a distinct drop in the OD450 was observed with higher plasma volumes. The oxytocin antiserum used in the assay showed no significant cross-reaction with other octapeptides tested. The assay was compared with a radioimmunoassay (RIA) procedure employing prior solvent extraction of plasma samples. The oxytocin concentrations assayed by EIA and RIA in plasma samples obtained from four cows before, during and after milking were highly correlated and very similar (r = 0.97). Hence the assay developed offers an attractive alternative to the RIA since no prior laborious plasma extraction is needed. Further, the assay has the distinct advantage of being non-radioactive in nature.
开发了一种用于测定牛血浆中催产素的高灵敏度和特异性的微量滴定板二抗酶免疫测定法（EIA），该方法使用生物素-链霉亲和素放大系统。生物素与催产素偶联，并用于在竞争测定中介导链霉亲和素-过氧化物酶与固定化催产素抗血清之间的连接。该测定直接在200微升牛血浆中进行。使用在无激素血浆中制备的催产素标准品。该测定的灵敏度为0.25 pg/孔，相当于血浆中1.25 pg/ml；在2.8 pg/孔时观察到50%的相对结合率。测定中使用的血浆体积范围为50至200微升，这不会影响催产素标准曲线的形状；然而，当血浆体积较大时，OD450会明显下降。该测定中使用的催产素抗血清与所测试的其他八肽没有明显的交叉反应。将该测定与采用血浆样品预先溶剂萃取的放射免疫测定法（RIA）程序进行了比较。通过EIA和RIA测定的从四头奶牛挤奶前、挤奶期间和挤奶后获得的血浆样品中的催产素浓度高度相关且非常相似（r = 0.97）。因此，所开发的测定法为RIA提供了一种有吸引力的替代方法，因为无需预先进行繁琐的血浆萃取。此外，该测定法具有本质上非放射性的明显优势。
