Quantitation of platelet derived growth factor receptors on cells from human arterial segments.
The study of receptor expression in tissue containing multiple cell types presents a two-fold problem. Firstly, in situ studies are difficult to perform quantitatively and secondly, enzymatic disruption of the primary tissue results in the loss of cell surface receptors which may or may not be resynthesised. In vitro culture of cells following isolation may also alter receptor expression. To circumvent these problems, we have devised a reproducible non-enzymatic method for releasing endothelial and smooth muscle cells from human arterial segments and separating the mixture into constituent cell populations, in order to facilitate semi-quantitative receptor investigation by ELISA. The method involves the mechanical disruption of small artery segments and passage through progressively smaller sieves, resulting in a mixed population of single cells. Magnetic beads covalently attached to identifying lectins or antibodies are then used in a multistep procedure to give highly enriched cell populations. The dissociation/enrichment steps can be modified to select other cell types which may be present in the primary tissue, such as macrophages. We have used these preparations for locating and semi-quantitating PDGF and EGF receptors on the constituent cells of human umbilical artery preparations by ELISA using monoclonal receptor antibodies.
Hicks
C
C
Department of Haematology, Prince of Wales Hospital, Randwick, NSW, Australia.
Sleigh
M J
MJ
Chesterman
C N
CN
D000911:Antibodies, Monoclonal; D002460:Cell Line; D002469:Cell Separation; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D004730:Endothelium, Vascular / Q000737:chemistry* / Q000166:cytology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D066246:ErbB Receptors / Q000032:analysis; D005347:Fibroblasts / Q000737:chemistry; D006801:Humans; D018189:Immunomagnetic Separation; D009131:Muscle, Smooth, Vascular / Q000737:chemistry* / Q000166:cytology; D017479:Receptors, Platelet-Derived Growth Factor / Q000032:analysis*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014469:Umbilical Arteries
D000911:Antibodies, Monoclonal; D066246:ErbB Receptors; D017479:Receptors, Platelet-Derived Growth Factor