Schlueter A J, Segre M, Segre D
Department of Veterinary Pathobiology, University of Illinois, Urbana 61801.
J Immunol Methods. 1989 Nov 13;124(1):35-42. doi: 10.1016/0022-1759(89)90183-x.
A method is described in which sheep red blood cells (SRBC) are coated with anti-immunoglobulin (Ig) antibodies (Ab) for use in reverse hemolytic plaque assays (RHPA) as follows. The non-complement fixing F(ab')2 fragments of rabbit anti-mouse Ig Ab are derivatized with the N-hydroxysuccinimide ester of palmitate. The hydrophobic palmitate tails spontaneously insert into the SRBC membranes, thus coating the cells with anti-Ig F(ab')2 molecules. The SRBC are lysed by successive additions of mouse Ig, rabbit anti-mouse Ig and complement. When this procedure is carried out in agar gel, Ig-secreting mouse cells produce localized hemolytic areas (plaques). The procedure is more reproducible and more sensitive than RHPA performed with protein A-coated SRBC. In principle, this procedure should be adaptable to the detection of cells secreting any molecule for which specific antibodies are available.
本文描述了一种方法，即通过以下方式用抗免疫球蛋白（Ig）抗体（Ab）包被绵羊红细胞（SRBC），用于反向溶血空斑试验（RHPA）。兔抗小鼠Ig Ab的非补体结合F(ab')2片段用棕榈酸N-羟基琥珀酰亚胺酯进行衍生化。疏水性的棕榈酸尾部会自发插入SRBC膜中，从而使细胞被抗Ig F(ab')2分子包被。通过连续添加小鼠Ig、兔抗小鼠Ig和补体来裂解SRBC。当在琼脂凝胶中进行此操作时，分泌Ig的小鼠细胞会产生局部溶血区域（空斑）。该方法比用蛋白A包被的SRBC进行的RHPA更具可重复性且更灵敏。原则上，该方法应适用于检测能分泌任何有特异性抗体的分子的细胞。
