Endocrine Research Unit, Division of Endocrinology, Mayo Clinic, Rochester, MN, USA.
J Lipid Res. 2011 Feb;52(2):408-15. doi: 10.1194/jlr.M008995. Epub 2010 Nov 28.
CD36 is a transmembrane protein present in many tissues that is believed to facilitate inward fatty acid transport. Western blotting is the most widely used method to measure tissue CD36 protein content, but it is time consuming, technically demanding, and semiquantitative. To more precisely measure adipose tissue CD36 content we developed an enzyme linked immunosorbent assay (ELISA) after establishing that: 1) the anti-CD36 antibodies gave a single distinct band on traditional Western blots, and 2) the vast majority of adipocyte CD36 resides in the plasma membrane. By using serial dilutions of each sample and including a calibrator sample and quality control sample on each plate, we could achieve inter- and intra-assay variability of ∼ 10%. We found that CD36 content in omental and abdominal subcutaneous adipose tissue varied over a 2-5-fold range depending upon the means of data expression (per units of tissue protein, weight, or lipid). Omental CD36 content in women decreased markedly (P = 0.01) as a function of fat cell size. For the most part, tissue CD36 content was not correlated with CD36 mRNA. This ELISA method for tissue CD36 content should enhance research into the role of this protein on tissue fatty acid uptake.
CD36 是一种存在于许多组织中的跨膜蛋白，被认为有助于脂肪酸向内转运。蛋白质印迹法是测量组织 CD36 蛋白含量最广泛使用的方法，但它既耗时，技术要求高，且半定量。为了更精确地测量脂肪组织 CD36 含量，我们在建立以下两点后开发了一种酶联免疫吸附测定法（ELISA）：1）抗 CD36 抗体在传统蛋白质印迹中产生单一明显条带，2）绝大多数脂肪细胞 CD36 位于质膜中。通过对每个样本进行系列稀释，并在每个板上包含校准样本和质控样本，我们可以实现约 10%的组内和组间变异性。我们发现，网膜和腹部皮下脂肪组织中的 CD36 含量根据数据表达的方式（每单位组织蛋白、重量或脂质）在 2-5 倍范围内变化。女性网膜 CD36 含量随脂肪细胞大小的变化明显降低（P = 0.01）。在大多数情况下，组织 CD36 含量与 CD36 mRNA 不相关。这种用于组织 CD36 含量的 ELISA 方法应该会增强对该蛋白在组织脂肪酸摄取中的作用的研究。
