Calibrating gelatin zymograms with human gelatinase standards.
We describe the use of gelatinase standards from human capillary whole blood for calibrating gelatin zymograms. Capillary blood was obtained by fingerstick puncture and prepared in nonreducing Laemmli SDS-PAGE sample buffer without heating. Gelatin zymography revealed that whole blood (0.25- 1 microliter) contained substantial fibroblast-derived (72-kDa) and neutrophil-derived (92-, 130-, and 225-kDa) gelatinases which could be used for calibration purposes. Calibration was linear under different electrophoretic conditions using 6.5-10% polyacrylamide gels containing 1-2 mg/ml gelatin (correlation range, r = 0.990-0.998; perfect correlation, r = 1.000). The use of the standards for characterizing gelatinases from arthritic joint fluid is demonstrated. The standards are easily obtained and well characterized, and should facilitate interlaboratory comparison and standardization.
Makowski
G S
GS
Department of Laboratory Medicine, University of Connecticut, School of Medicine, Farmington 06030, USA.
Ramsby
M L
ML
D001168:Arthritis / Q000201:enzymology; D002138:Calibration; D002196:Capillaries; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000201:enzymology; D005780:Gelatin; D018093:Gelatinases; D006801:Humans; D007962:Leukocytes / Q000201:enzymology; D008666:Metalloendopeptidases / Q000097:blood*; D012015:Reference Standards; D013582:Synovial Fluid / Q000201:enzymology
D005780:Gelatin; D018093:Gelatinases; D008666:Metalloendopeptidases