Falahat Rana, Wiranowska Marzenna, Gallant Nathan D, Toomey Ryan, Hill Robert, Alcantar Norma
Department of Chemical & Biomedical Engineering, College of Engineering, University of South Florida, Tampa, FL 33620, USA.
Department of Pathology and Cell Biology, Morsani College of Medicine, University of South Florida, Tampa, FL 33612, USA.
Cell Immunol. 2015 Nov-Dec;298(1-2):96-103. doi: 10.1016/j.cellimm.2015.09.009. Epub 2015 Sep 28.
Quantitative analysis of MUC1, a cell membrane associated mucin, expressed by intact cells of epithelial origin previously has been limited to flow cytometry, which requires using large quantities of cells and antibodies. Here, for the first time, we report the development of a novel Cellular-based Enzyme Linked Immunosorbent Assay (Cell ELISA) to quantify the expression of MUC1 by cell lines of epithelial and neuroectodermal origin using an antibody recognizing a specific tandem repeat found in the extracellular domain of MUC1. In contrast to flow cytometry, this method requires a much lower number of cells. We report here the results obtained from two variants of this Cell ELISA in live and fixed cells. We found that the Cell ELISA in live cells was not sensitive enough to detect a difference in MUC1 levels between the normal cells and tumor cells. However, we found that Cell ELISA in fixed cells followed by whole cell staining was a dependable method of MUC1 level detection in the normal and tumor cells showing significantly higher levels of MUC1 receptor in the tumor cells when compared to the normal controls. Therefore, we conclude that the Cell ELISA in fixed cells is an efficient method for quantifying the expression of MUC1 by epithelial and neuroectodermal cancer cell lines.
对上皮来源的完整细胞所表达的细胞膜相关黏蛋白MUC1进行定量分析，此前仅限于流式细胞术，该方法需要使用大量细胞和抗体。在此，我们首次报告了一种新型的基于细胞的酶联免疫吸附测定法（细胞ELISA）的开发，该方法使用识别MUC1细胞外结构域中特定串联重复序列的抗体，来定量上皮和神经外胚层来源的细胞系中MUC1的表达。与流式细胞术不同，该方法所需的细胞数量要少得多。我们在此报告了从这种细胞ELISA的两种变体在活细胞和固定细胞中获得的结果。我们发现，活细胞中的细胞ELISA对检测正常细胞和肿瘤细胞之间MUC1水平的差异不够敏感。然而，我们发现，固定细胞后进行全细胞染色的细胞ELISA是检测正常细胞和肿瘤细胞中MUC1水平的可靠方法，与正常对照相比，肿瘤细胞中MUC1受体水平显著更高。因此，我们得出结论，固定细胞中的细胞ELISA是定量上皮和神经外胚层癌细胞系中MUC1表达的有效方法。
