Highly enriched preparations of cultured myocardial cells for biochemical and physiological analyses.
Cultures of enzyme-dissociated myocardial cells contain myocytes as well as other cell types (e.g. fibroblasts); therefore, cell separation is necessary to interpret accurately biochemical measurements of substances or physiological measurements of processes common to more than one type of cell. Previous methods of cell separation have resulted in cultures enriched with, at most, about 80% myocytes. In the present study, a monoclonal antibody to cell-surface adhesion factors, CSAT, was used to obtain four highly enriched preparations: cultures of 7-day embryonic chick ventricular cells that contained either 99.1 +/- 0.2% (n = 10) myocytes or 96.5 +/- 0.8% (n = 6) fibroblasts; and cultures of 17-day chick cells that contained 99.1 +/- 0.2% (n = 11) myocytes or 92.5 +/- 0.8% (n = 8) fibroblasts. Following the removal of CSAT from myocyte-enriched cultures, 96.9 +/- 0.4% (n = 7) of the myocytes obtained from 7-day embryos and 93.8 +/- 1.4% (n = 6) of the myocytes obtained from 17-day embryos attached to culture dishes. Using these four preparations, we found the sialic acid content of myocytes to be significantly less than that of fibroblasts at both 7 and 17 days of development. In fact, the sialic acid content of myocytes did not change during this period, whereas that of fibroblasts increased significantly. These differences underscore the necessity of using highly enriched cell preparations. The antibody/differential adhesion method described provides an opportunity to evaluate and correlate the biochemical, physiological and pharmacological properties of myocytes without interference from other cell types.
Department of Physiology, Texas Tech University Health Sciences Center, Lubbock 79430.
J C
JC
McDonagh
E K
EK
Cebrat
R D
RD
Nathan
D000818:Animals; D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D015815:Cell Adhesion Molecules; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D002642:Chick Embryo; D006321:Heart / Q000196:embryology; D009206:Myocardium / Q000166:cytology*; D012794:Sialic Acids / Q000032:analysis
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D015815:Cell Adhesion Molecules; D012794:Sialic Acids