Isolation of rat peritoneal mononuclear and polymorphonuclear leucocytes on discontinuous gradients of Nycodenz.
Centrifugation of rat leucocytes from thioglycollate-induced inflammatory peritoneal exudate on a discontinuous gradient of Nycodenz with a density of 1106 g/l and an osmolarity of 400 mosmol/l separated the polymorphonuclear from the mononuclear leucocytes. The cells on the interphase between the buffer and the gradient medium contained 96% mononuclear leucocytes with a recovery of greater than 60%, and the bottom fraction consisted of 98% polymorphonuclear leucocytes with a yield of 91%, when an exudate isolated 20 h after the injection of thioglycollate was fractionated. Leucocytes isolated from non-inflamed rat peritoneum could be enriched in the fraction of nonspecific esterase-positive cells from 86% to 96% with a recovery of 82% on a gradient with a density of 1091 g/l and an osmolarity of 325 mosmol/l. The viability of the isolated cells was greater than 95% (trypan blue exclusion test), and there was no measurable reduction in the fraction of phagocytosing cells (latex and opsonized zymosan) after exposure to the hypertonic gradient material.
Fisker
S
S
Institute of Physiology, University of Aarhus, Denmark.
Kudahl
K
K
Sonne
O
O
D000818:Animals; D002469:Cell Separation / Q000379:methods*; D002470:Cell Survival; D002499:Centrifugation, Density Gradient / Q000379:methods*; D004950:Esterases / Q000378:metabolism; D007472:Iohexol; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000201:enzymology; D008297:Male; D009504:Neutrophils / Q000187:drug effects; D010529:Peritoneal Cavity / Q000166:cytology*; D010587:Phagocytosis; D051381:Rats; D011919:Rats, Inbred Strains; D013864:Thioglycolates / Q000494:pharmacology; D015054:Zymosan / Q000494:pharmacology
D013864:Thioglycolates; D007472:Iohexol; D015054:Zymosan; D004950:Esterases