A simple and rapid technique for postelectrophoretic detection of proteases using azocasein.
Azocasein, diffused into gels after polyacrylamide gel electrophoresis, provides a useful substrate for simple and rapid protease detection. After electrophoresis, the gels are incubated in an azocasein solution, immersed in trichloroacetic acid and then treated with NaOH. Proteolytic zones appear as uncolored bands on an orange background. The gels can be stored in trichloroacetic acid for months. This technique gives good results with serine proteases, acid proteases and metalloproteases. The method can be used under nondenaturing conditions and in the presence of sodium dodecyl sulfate. The sensitivity of this method is at the submicrogram protein level for trypsin.
Vázquez Peyronel
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Cátedra de Bioquímica, Facultad de Química, Montevideo.
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D002364:Caseins / Q000378:metabolism*; D004396:Coloring Agents; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D010450:Endopeptidases / Q000032:analysis* / Q000378:metabolism; D008666:Metalloendopeptidases / Q000032:analysis; D012680:Sensitivity and Specificity; D012697:Serine Endopeptidases / Q000032:analysis; D012972:Sodium Hydroxide; D014238:Trichloroacetic Acid; D014357:Trypsin / Q000032:analysis
D002364:Caseins; D004396:Coloring Agents; C026018:azocasein; D012972:Sodium Hydroxide; D014238:Trichloroacetic Acid; D010450:Endopeptidases; D012697:Serine Endopeptidases; D014357:Trypsin; D008666:Metalloendopeptidases