Hahn S E, Parkes J G, Goldberg D M
Department of Clinical Biochemistry, University of Toronto, Ontario, Canada.
J Clin Lab Anal. 1992;6(4):182-9. doi: 10.1002/jcla.1860060404.
We describe an enzyme-linked immunosorbent assay (ELISA) to measure apolipoproteins AI and B secreted by Hep G2 cells and in cell homogenates. These assays utilize commercially available polyclonal antibodies, affinity-purified to improve their specificity, thereby achieving a dramatic increase in the sensitivity of the assay. These affinity-purified antibodies were also more sensitive than a series of monoclonal antibodies tested. We achieved a sensitivity of 0.4 ng in the apo AI assay, and a sensitivity of 5 ng in the apo B assay. By these methods, we measured secretion rates by Hep G2 cells of 358 +/- 41 ng/mg cell protein/hr for apo B and 137 +/- 8 ng/mg cell protein/hr for apo AI. These assays also allowed the measurement of intracellular apolipoproteins and thus can be used to facilitate investigations of human lipoprotein metabolism in cell culture systems.
我们描述了一种酶联免疫吸附测定法（ELISA），用于测量Hep G2细胞分泌的以及细胞匀浆中的载脂蛋白AI和B。这些测定法使用市售的多克隆抗体，经过亲和纯化以提高其特异性，从而使测定的灵敏度显著提高。这些亲和纯化的抗体也比所测试的一系列单克隆抗体更灵敏。我们在载脂蛋白AI测定中达到了0.4 ng的灵敏度，在载脂蛋白B测定中达到了5 ng的灵敏度。通过这些方法，我们测得Hep G2细胞的载脂蛋白B分泌率为358±41 ng/mg细胞蛋白/小时，载脂蛋白AI分泌率为137±8 ng/mg细胞蛋白/小时。这些测定法还能够测量细胞内载脂蛋白，因此可用于促进细胞培养系统中人类脂蛋白代谢的研究。
