Detection of Intracellular Factor VIII Protein in Peripheral Blood Mononuclear Cells by Flow Cytometry. (28th February 2013)

• Record Type: Journal Article
• Authors: Pandey, Gouri Shankar; Tseng, Sandra C.; Howard, Tom E.; Sauna, Zuben E.
• Other Names: Kesari Akanchha Academic Editor.
• Is Part Of: BioMed research international. Volume 2013(2013)
• Publication Date: 2013-02-28
• Subjects: Medicine -- Periodicals; Biology -- Periodicals; Biotechnology -- Periodicals; Life sciences -- Periodicals; 610.5
• DOI: 10.1155/2013/793502
• Languages: English
• ISSNs: 2314-6133

Abstract

Abstract : Flow cytometry is widely used in cancer research for diagnosis, detection of minimal residual disease, as well as immune monitoring and profiling following immunotherapy. Detection of specific host proteins for diagnosis predominantly uses quantitative PCR and western blotting assays. In this study, we optimized a flow cytometry-based detection assay for Factor VIII protein in peripheral blood mononuclear cells (PBMCs). An indirect intracellular staining (ICS) method was standardized using monoclonal antibodies to different domains of human Factor VIII protein. The FVIII protein expression level was estimated by calculating the mean and median fluorescence intensities (MFI) values for each monoclonal antibody. ICS staining of transiently transfected cell lines supported the method's specificity. Intracellular FVIII protein expression was also detected by the monoclonal antibodies used in the study in PBMCs of five blood donors. In summary, our data suggest that intracellular FVIII detection in PBMCs of hemophilia A patients can be a rapid and reliable method to detect intracellular FVIII levels.