Monitoring internalization of Histoplasma capsulatum by mammalian cell lines using a fluorometric microplate assay.
We have developed a fluorometric microtiter plate assay to quantify the internalization of Histoplasma capsulatum yeasts by macrophages. The assay utilizes the fluorescent dye Calcofluor White to label the yeast cell wall and the vital dye trypan blue, which does not enter viable macrophages, to quench fluorescence of extracellular labeled yeasts. Murine RAW 264.7 cells showed more efficient internalization of strain G217B yeasts than human U937 cells. Both cell lines exhibited a dependence upon actin, and, to a lesser degree, microtubules, in G217B uptake.
Scott
A J
AJ
Department of Medical Microbiology and Immunology, University of Wisconsin Medical School, Madison 53706, USA.
Woods
J P
JP
D000818:Animals; D001557:Benzenesulfonates / Q000378:metabolism; D002460:Cell Line; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000378:metabolism; D005470:Fluorometry; D006658:Histoplasma / Q000276:immunology* / Q000502:physiology; D006801:Humans; D008264:Macrophages / Q000276:immunology* / Q000382:microbiology*; D051379:Mice; D010587:Phagocytosis; D013194:Staining and Labeling; D014343:Trypan Blue / Q000378:metabolism; D020298:U937 Cells
D001557:Benzenesulfonates; D005456:Fluorescent Dyes; C007061:C.I. Fluorescent Brightening Agent 28; D014343:Trypan Blue