Fluoresceinated alpha 2-macroglobulin as a probe for studying macrophages.
The potential value of fluorescein-conjugated human alpha 2-macroglobulin as a probe for studying macrophages in murine peritoneal exudate and spleen cell suspensions has been investigated using a fluorescence-activated cell sorter. These studies revealed that the number of alpha 2-macroglobulin-positive cells in the mixtures examined correlated closely with their macrophage content as determined morphologically. Furthermore cells separated on the FACS on the basis of their strong alpha 2-macroglobulin binding exhibited macrophage morphology and expressed Fc receptors on their surface. Conversely the alpha 2-macroglobulin-negative population contained few macrophages or Fc rosette-forming cells. Extensive blocking and comparative binding studies with a number of purified human alpha 2-macroglobulin preparations and derivatives thereof, and a variety of purified proteins, confirmed that the binding of fluorescein-conjugated alpha 2-macroglobulin to peritoneal exudate cells was specific. Furthermore the binding occurs in a highly reproducible manner. These observations suggest that fluorescein-conjugated alpha 2-macroglobulin in conjugation with flow cytometry is a sensitive and reliable method for both identifying and isolating subpopulations of macrophages.
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K
James
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I
Milne
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K
Donaldson
D000818:Animals; D001202:Ascitic Fluid; D002469:Cell Separation; D005434:Flow Cytometry; D019793:Fluorescein; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006801:Humans; D007122:Immunoelectrophoresis; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D008264:Macrophages / Q000032:analysis*; D051379:Mice; D011971:Receptors, Immunologic / Q000378:metabolism; D013154:Spleen / Q000166:cytology; D000511:alpha-Macroglobulins / Q000378:metabolism
D005452:Fluoresceins; D005456:Fluorescent Dyes; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D011971:Receptors, Immunologic; D000511:alpha-Macroglobulins; D019793:Fluorescein