Monji N, Gomez N O, Kawashima H, Ali H, Castro A
J Clin Endocrinol Metab. 1980 Feb;50(2):355-9. doi: 10.1210/jcem-50-2-355.
Enzyme immunoassay for cortisol was developed using beta-galactosidase as an enzyme label and m-maleimidobenzoyl derivatives of cortisol, i.e. cortisol-21-m-maleimidobenzoate (CT-MB) and cortisol-21-hemisuccinate conjugated with m-maleimidobenzoic acid through p-phenylenediamine linkage (CHS-MB), as the haptens coupled to sulfhydryl groups of the enzyme. This enzyme-coupling procedure was highly efficient; over 90% of the enzyme was labeled while full enzyme activity was retained. CHS-MB-beta-galactosidase conjugate showed high immunoreactivity to antibody produced against cortisol-21-hemisuccinate-bovine serum albumin but showed poor displacement with the added cortisol. CT-MB-beta-galactosidase conjugate, however, showed not only a high immunoreactivity to the antibody but also displaced well with cortisol, showing maximum sensitivity of 1 microgram/dl with a 20-microliter sample size. Modification around the linkage of cortisol derivative resulted in high sensitivity to cortisol. Cross-reactions to cortisol-21-acetate, cortisone, and corticosterone were 150%, 10%, and 7%, respectively. The accuracy, precision and correlation with RIA of this method were satisfactory for clinical application.
采用β-半乳糖苷酶作为酶标记物，以皮质醇的间马来酰亚胺苯甲酰衍生物，即皮质醇-21-间马来酰亚胺苯甲酸酯（CT-MB）和通过对苯二胺连接与间马来酰亚胺苯甲酸共轭的皮质醇-21-半琥珀酸酯（CHS-MB）作为与酶的巯基偶联的半抗原，开发了用于皮质醇的酶免疫测定法。这种酶偶联方法效率很高；超过90%的酶被标记，同时保留了全部酶活性。CHS-MB-β-半乳糖苷酶偶联物对针对皮质醇-21-半琥珀酸酯-牛血清白蛋白产生的抗体显示出高免疫反应性，但与添加的皮质醇的置换效果较差。然而，CT-MB-β-半乳糖苷酶偶联物不仅对抗体显示出高免疫反应性，而且与皮质醇的置换效果良好，对于20微升的样本量，最大灵敏度为1微克/分升。皮质醇衍生物连接部位周围的修饰导致对皮质醇具有高灵敏度。与皮质醇-21-乙酸酯、可的松和皮质酮的交叉反应分别为150%、10%和7%。该方法的准确性、精密度以及与放射免疫分析的相关性对于临床应用而言是令人满意的。
