Department of Biomedical Sciences, University of South Alabama, Mobile, Alabama 36688, USA.
Cytometry A. 2010 Apr;77(4):305-9. doi: 10.1002/cyto.a.20865.
Phagocytosis is a highly conserved mechanism exhibited by both free-living amebas and mammalian blood cells. Similarities demonstrated by either cell type during engulfment of the same bacterial species may imply analogous surface proteins involved in receptor-mediated endocytosis. The increased availability of anti-human leukocyte antibodies or clusters of differentiation (CD) markers used in conjunction with flow cytometric (FCM) and/or immunohistochemical (IHC) analysis provides investigators with a relatively easy method to screen different cell populations for comparable plasma membrane proteins. In this study, we incubated Naegleria and Acanthamoeba amebas with several directly conjugated anti-human leukocyte monoclonal antibodies (mAb) for similarly recognized amebic epitopes. CD marker selection was based upon a recognized role of each mAb in phagocyte activation and/or uptake of bacteria. These included CD14, CD45, and CD206. In FCM, only one CD45 antibody demonstrated strong reactivity with both Naegleria fowleri and Naegleria gruberi that was not expressed in similarly tested Acanthamoeba species. Additional testing of N. gruberi by IHC demonstrated reactivity to a different CD45 antibody. Our results suggest a possible utility of using anti-human leukocyte antibodies to screen amebic cells for similarly expressed protein epitopes. In doing so, several important items must be considered when selecting potential mAbs for testing to increase the probability of a positive result.
吞噬作用是自由生活的变形虫和哺乳动物血细胞都具有的一种高度保守的机制。在吞噬相同细菌物种时，两种细胞类型表现出的相似性可能意味着参与受体介导的胞吞作用的类似表面蛋白。越来越多的抗人白细胞抗体或分化群 (CD) 标记物与流式细胞术 (FCM) 和/或免疫组织化学 (IHC) 分析一起使用，为研究人员提供了一种相对简单的方法，用于筛选不同的细胞群体中具有可比性的质膜蛋白。在这项研究中，我们用几种直接偶联的抗人白细胞单克隆抗体 (mAb) 孵育 Naegleria 和 Acanthamoeba 变形虫，以识别类似的变形虫表位。CD 标记物的选择基于每种 mAb 在吞噬细胞活化和/或细菌摄取中的作用。这些包括 CD14、CD45 和 CD206。在 FCM 中，只有一种 CD45 抗体对 Naegleria fowleri 和 Naegleria gruberi 表现出强烈的反应性，而在类似测试的 Acanthamoeba 物种中则没有表达。对 N. gruberi 的进一步 IHC 测试表明对不同的 CD45 抗体有反应性。我们的研究结果表明，使用抗人白细胞抗体筛选具有类似表达蛋白表位的变形虫细胞可能具有一定的应用价值。在这样做时，在选择用于测试的潜在 mAb 时，必须考虑几个重要因素，以增加阳性结果的可能性。
