Surface flow cytometry analysis

H., Nakahata, T. (2005). Prospective characterization of neural stem cells by flow cytometry analysis using a combination of surface markers. J Neurosci Res. 80, 456-66. 

Data are analyzed by flow cytometry analysis software (BD FACS Diva, BD Biosciences Flowjo, TreeStar). Data presentation using the frequency of positive events is appropriate only in presence of a bimodal distribution of the emitted fluorescence. In this case, there is no particular preference for the use of the histogram or the dot plot to present data. In case of a non-bimodal distribution of the emitted fluorescence, data should be reported as relative MFI (i.e., the ratio between the MFI values of the sample stained with aU the experimental markers and the MFI values of the negative control sample) and the histogram layout should be preferred. Differently from frequency data, this analytical approach provides relative quantitative information of the chemokine receptor expression levels on the surface of each... 

Figu re 4.8 Flow cytometry analysis of silica nanoparticles surface-modified with proteins. MPMS nanoparticles (a, c) and TEOS nanoparticles (b, d) modified with GFP (a, b) or phycoerythrin-conjugated streptavidin... 

N5. Noronha, A., and Richman, D. R, Simultaneous cell surface phenotype and cell cycle analysis of lymphocytes by flow cytometry. J. Histochem. Cytochem. 32, 821-829 (1984). 

Roe, R., Robins, R. A., Laxton, R R., and Baldwin, R W (1985) Kinetics of divalent monoclonal antibody binding to tumor cell surface antigens using flow cytometry—standardization and mathematical analysis Mol Immunol. 22, 11—21... 

Successful cell surface display of the protein can be verified by inducing gene expression via addition of anhydrotetracycline to the culture medium and immunofluorescence staining of the cells using an antibody directed against the protein to be displayed (Protocol 1). Analysis can be performed by fluorescence microscopy or flow cytometry. 

Krutzik PO, Clutter MR, Nolan GP. Coordinate analysis of murine immune cell surface markers and intracellularphosphoproteins by flow cytometry. J Immunol 2005 175(4) 2357-65. 

Pollice, A A, McCoy, J. P, Jr., Shackney, S E, Smith, C A., Agarwal, J., Burholt, D R., Janocko, L. E, Hornicek, F J, Singh, S. G, and Hartsock, R. J. (1992) Sequential paraformaldehyde and methanol fixation for simultaneous flow cytometric analysis of DNA, cell-surface proteins, and intracellular proteins. Cytometry 13,432-444... 

Cell-enzyme-linked immunosorbent assay (cell-ELISA) is an useful technique for the quantitative analysis of cell surface antigen expression that was developed on the basis of enzyme immunohistochemistry (EIH) and ELISA. Since its development, which was made possible by the establishment of monoclonal antibody technology, a wide range of cell types and surface molecules were analyzed by cell-ELISA. Here we show four variants of this method and provide a brief comparison of cell-ELISA with flow cytometry (FACS) and radioimmunobinding assay (BIA), which are other methods for the quantitative detection of cell-surface molecules. We describe step-by-step procedures for both direct and indirect cell-ELISA using either adherent or nonadherent live cells. 

---