Mengeling W L, Paul P S
National Animal Disease Center, USDA, Ames, IA 50010.
Am J Vet Res. 1988 Aug;49(8):1235-9.
During initial studies, we found that many fluorescein isothiocyanate-labeled anti-immunoglobulin conjugates were unstable and tended to aggregate and precipitate when used for indirect immunofluorescence microscopy. In some instances, the precipitate was extensive enough to interfere with interpretation of the test results. Attempts to resolve this problem resulted in a procedure by which such conjugates were digested with papain to Fab and Fc fragments before use. Aggregation and precipitation were prevented, while desired antibody activity was retained. Digestion with papain also reduced the diffuse background fluorescence (commonly referred to as nonspecific fluorescence or staining) that is often associated with conjugates before they are sorbed with tissue powders or chromatographed to remove highly labeled immunoglobulin molecules.
在初步研究中，我们发现许多异硫氰酸荧光素标记的抗免疫球蛋白缀合物不稳定，在用于间接免疫荧光显微镜检查时容易聚集和沉淀。在某些情况下，沉淀物足够多，足以干扰测试结果的解读。为解决这个问题所做的尝试产生了一种方法，即此类缀合物在使用前用木瓜蛋白酶消化成Fab和Fc片段。这样可以防止聚集和沉淀，同时保留所需的抗体活性。用木瓜蛋白酶消化还减少了弥漫性背景荧光（通常称为非特异性荧光或染色），这种荧光在缀合物被组织粉末吸附或进行色谱分离以去除高度标记的免疫球蛋白分子之前常常与之相关。
