Development of a high capacity microassay for measurement of neutrophil adhesion.
A high capacity semiautomated assay for neutrophil adhesion was developed utilizing the 96 well microtiter plate format. Optimal adhesion occurred with about 150 microliters/well of neutrophils at 5 X 10(6) cells/ml in tissue culture plates that had been precoated with 5% serum. Optimal incubation times were 10 min for f-Met-Leu-Phe-treated cells and 20 min for A23187 or phorbol myristate acetate stimulation. Optimal washing occurred after three washes with a Cetus Pro/pette pump. Adhesion could be effectively blocked by the inhibitors of cellular protein kinase C, an enzyme known to be necessary for the occurrence of neutrophil adhesion.
D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002448:Cell Adhesion / Q000187:drug effects; D002860:Chromium Radioisotopes; D006801:Humans; D007958:Leukocyte Count / Q000295:instrumentation / Q000379:methods*; D008297:Male; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology / Q000502:physiology*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D051381:Rats; D011919:Rats, Inbred Strains; D012710:Serum Albumin, Bovine; D019311:Staurosporine; D013997:Time Factors
D000470:Alkaloids; D002860:Chromium Radioisotopes; D012710:Serum Albumin, Bovine; D000001:Calcimycin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D011493:Protein Kinase C; D019311:Staurosporine