Impact of cell harvesting methods on detection of cell surface proteins and apoptotic markers

Assays based on the flow cytometry technique allow a convenient analysis of multiple cellular parameters; however, their results should be interpreted cautiously due to a strong impact of confounding factors. Different techniques in cell culturing such as either enzymatic or mechanic detachment of adherent cells can heavily influence the structure of the cell membrane or presence of the surface antigens leading to strong false positive signals, and finally, substantial experimental bias. The aim of our study was to assess and compare the impact of cell harvesting methods (both enzymatic and non-enzymatic) on the apoptosis process and on the surface antigen cytometric analyses. We found significant differences in the quality of analysis in terms of the amount of detected surface markers determined by the detachment method. Our results demonstrated clearly how important it is to carefully choose the appropriate detachment method and may help to avoid mistakes in experiment planning. In conclusion, we recommend to adjust the detachment method to the type of analyzed markers (surface antigens or translocated phosphatidylserine).

基于流式细胞术（flow cytometry）的检测技术可便捷分析多项细胞参数，但由于混杂因素会对检测结果产生显著影响，因此需谨慎解读其结果。在细胞培养过程中，针对贴壁细胞（adherent cells）所采用的酶解或机械解离等不同方式，会严重影响细胞膜结构或表面抗原（surface antigens）的表达情况，进而产生强烈的假阳性信号，最终导致显著的实验偏倚。本研究旨在评估并比较细胞收获方法（cell harvesting methods，包括酶解与非酶解两类）对细胞凋亡（apoptosis）过程及流式细胞术检测表面抗原结果的影响。研究发现，不同解离方式会影响所检测到的表面标志物数量，进而导致分析质量存在显著差异。本研究结果清晰表明，谨慎选择合适的细胞解离方法至关重要，该结论可助力避免实验规划中的失误。综上，我们建议根据所分析的标志物类型（表面抗原或易位磷脂酰丝氨酸（phosphatidylserine））调整细胞解离方法。