A sensitive enzyme immunoassay for urine 6β-hydroxycortisol has been developed. Enzyme labeling of 6β-hydroxycortisol was accomplished by the N-succinimidyl ester method. The use of N-succinimidyl ester of 6β-hydroxycortisol 21-hemisuccinate and β-galactosidase in an appropriate molar ratio provided a conjugate suitable for enzyme immunoassay. Antiserum was prepared by immunization with the 6β-hydroxycortisol 21-hemisuccinate-bovine serum albumin conjugate in the rabbit. Sufficient sensitivity and improved specificity of the assay system could be obtained by the selective blocking of less specific antibodies. The quantitation limit of 6β-hydroxycortisol was approximately 10 pg, which is comparable to that of radioimmunoassay. The intra- and inter-assay coefficients of variation for 6β-hydroxycortisol in human urine were 5.9-8.1% and 2.8-12.3%, respectively.
尿液 6β-hydroxycortisol 的灵敏酶免疫测定方法已经研制成功。6β-hydroxycortisol 的酶标记是通过 N-琥珀
酰亚胺酯法完成的。使用 6β-hydroxycortisol 21-hemisuccinate 的 N-琥珀
酰亚胺酯和
β-半乳糖苷酶以适当的摩尔比提供了适合酶免疫测定的共轭物。用 6β-hydroxycortisol 21-hemisuccinate-bovine serum albumin conjugate 免疫兔制备抗血清。通过选择性阻断特异性较低的
抗体,可以获得足够的灵敏度并提高检测系统的特异性。6β-hydroxycortisol 的定量限约为 10 pg,与放射免疫法相当。人体尿液中 6β-hydroxycortisol 的测定内变异系数为 5.9-8.1%,测定间变异系数为 2.8-12.3%。