Establish a Flow Cytometric Method for Quantitative Detection of Beclin-1 Expression

Flow cytometry is an advanced technology for efficient, rapid, specific and multi-parameter analysis of single cells in various basic research fields including cytobiology, immunology, genetic, hematology and other basic research. Beclin-1 protein is an important indicator in monitoring autophagic activity. However, quantitative flow cytometry had been rarely reported till now to be applied in the detection of Beclin-1 expression. The present study was aimed to establish a flow cytometric method for quantitative detection of Beclin-1 expression by employing the autophagy inhibitor 3-methyladenine as the control. A multi-parameter optimal method for Beclin-1 protein staining is as follows. 2?% bovine serum albumin in phosphate buffered saline was used for sample block. Concentration of primary antibody was 0.004?μg/μL. Samples were incubated at room temperature (25?°C) for 30?min. The prepared samples had better to be detected immediately or to be stored at 4?°C and detected within 6?h, otherwise the samples should be fixed in 1?% paraformaldehyde storing at 4?°C and detected within 3 d. Furthermore, we employed the immunohistochemistry to validate the method in vivo, the results confirmed flow cytometric method. The established flow cytometric analysis for Beclin-1 protein has the advantage of simpleness, speediness, sensitivity and reproducibility.