Quantification of mucosal mononuclear cells in tissues with a fluorescent bead-based polychromatic flow cytometry assay

Since the vast majority of infections occur at mucosal surfaces, accurate characterization of mucosal immune cells is critically important for understanding transmission and control of infectious diseases. Standard flow cytometric analysis of cells obtained from mucosal tissues can provide valuable...

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Bibliographic Details
Published in:Journal of immunological methods 2011-03, Vol.367 (1-2), p.95-98, Article 95
Main Authors: Reeves, R. Keith, Evans, Tristan I., Gillis, Jacqueline, Wong, Fay E., Connole, Michelle, Carville, Angela, Johnson, R. Paul
Format: Article
Language:English
Subjects:
Animals
Biological and medical sciences
biopsy
CD8-positive T-lymphocytes
dendritic cells
Flow cytometry
Flow Cytometry - methods
Fluorescence
Fluorescent beads
Fundamental and applied biological sciences. Psychology
Fundamental immunology
HLA-DR Antigens - analysis
infectious diseases
Leukocyte Common Antigens - analysis
Leukocyte Count
Leukocytes, Mononuclear - cytology
Lymphocyte enumeration
Macaca mulatta
Molecular immunology
monocytes
Mucosal lymphocytes
Mucous Membrane - cytology
natural killer cells
phenotype
Reproducibility of Results
Techniques
tissues
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Summary:Since the vast majority of infections occur at mucosal surfaces, accurate characterization of mucosal immune cells is critically important for understanding transmission and control of infectious diseases. Standard flow cytometric analysis of cells obtained from mucosal tissues can provide valuable information on the phenotype of mucosal leukocytes and their relative abundance, but does not provide absolute cell counts of mucosal cell populations. We developed a bead-based flow cytometry assay to determine the absolute numbers of multiple mononuclear cell types in colorectal biopsies of rhesus macaques. Using 10-color flow cytometry panels and pan-fluorescent beads, cells were enumerated in biopsy specimens by adding a constant ratio of beads per mg of tissue and then calculating cell numbers/mg of tissue based on cell-to-bead ratios determined at the time of sample acquisition. Testing in duplicate specimens showed the assay to be highly reproducible (Spearman R=0.9476, P
ISSN:0022-1759
1872-7905
DOI:10.1016/j.jim.2011.02.002