Grenett H E, Garver F A
J Lab Clin Med. 1980 Oct;96(4):597-605.
The experimental details of ELISA for the identification and quantitation of Hb S are presented; the assay is based upon the passive adsorption of Hb S top a solid phase (polystyrene tubes) and the addition of monospecific rabbit antibodies capable of recognizing the (beta 6 Glu leads to Val) substitution in Hb S. After the addition of alkaline phosphatase-conjugated goat antibody to rabbit IgG and substrate, the yellow color produced by hydrolysis of substrate is measured spectrophotometrically. For the identification and quantitation of Hb S in unknown samples, the hemolysate is added to the Hb S-coated tubes before the addition of antibody to Hb S, thus causing an inhibition of the antigen-antibody reaction as evidenced by an absence or reduction of color formation. With this procedure, there is no cross-reactivity with normal hemoglobins, and the immunoassay has a sensitivity in detecting 50 ng quantities of the abnormal hemoglobin in a 5 microgram hemolysate. The assay can be performed on multiple samples in 1 day and offers many advantages over other techniques currently used for the identification and quantitation of Hb S and other abnormal hemoglobins in the clinical laboratory.
本文介绍了用于鉴定和定量血红蛋白S（Hb S）的酶联免疫吸附测定（ELISA）的实验细节；该测定基于Hb S被动吸附到固相（聚苯乙烯管）上，并添加能够识别Hb S中（β6谷氨酸突变为缬氨酸）取代的单特异性兔抗体。在添加碱性磷酸酶偶联的山羊抗兔免疫球蛋白G和底物后，通过分光光度法测量底物水解产生的黄色。为了鉴定和定量未知样品中的Hb S，在添加抗Hb S抗体之前，将溶血产物添加到包被有Hb S的管中，从而抑制抗原-抗体反应，表现为颜色形成缺失或减少。采用该方法，与正常血红蛋白无交叉反应，免疫测定在检测5微克溶血产物中50纳克量的异常血红蛋白时具有灵敏度。该测定可在1天内对多个样品进行检测，与临床实验室目前用于鉴定和定量Hb S及其他异常血红蛋白的其他技术相比具有许多优势。
