Purification of murine endothelial cell cultures by flow cytometry using fluorescein-labeled griffonia simplicifolia agglutinin.
Griffonia simplicifolia agglutinin (GSA) is a valuable histochemical tool in the identification of endothelium. In this study GSA labeled with fluorescein isothiocyanate (GSA-FITC) was used to purify cultures of murine cerebral microvascular endothelium. Cultures were stained with GSA-FITC, then sorted using a fluorescence-activated cell sorter (FACS). GSA-positive endothelial cells were collected, re-cultured, and subsequently re-analyzed by FACS using GSA-FITC. Cultures that initially contained 80 +/- 3 to 89 +/- 3% (X +/- SE) GSA-positive cells were purified to 98 +/- 1% positivity. Immunohistochemistry with an anti-muscle-action antibody confirmed that FACS sorting of GSA-FITC-stained cells effectively removed contaminating smooth muscle cells from endothelial cell cultures. Viability, proliferation, and prostaglandin production of the cells was unaltered by lectin staining and FACS sorting. Thus, GSA-FITC can be used in conjunction with flow cytometry to enhance the purity of murine endothelial cell cultures without adversely affecting cell viability, growth, or metabolism.
Department of Pathology, University of Iowa, Iowa City 52242.
G
G
Sahagun
S A
SA
Moore
Z
Z
Fabry
R L
RL
Schelper
M N
MN
Hart
D000199:Actins / Q000378:metabolism; D000818:Animals; D002455:Cell Division; D002469:Cell Separation; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology* / Q000378:metabolism; D005434:Flow Cytometry / Q000379:methods*; D005452:Fluoresceins; D006651:Histocytochemistry; D037102:Lectins; D051379:Mice; D037121:Plant Lectins; D011453:Prostaglandins / Q000378:metabolism
D000199:Actins; D005452:Fluoresceins; C045475:Griffonia simplicifolia lectins; D037102:Lectins; D037121:Plant Lectins; D011453:Prostaglandins