Fig. 3

Characterization of PR3/PRTN3 activity in subclinical rejection urine. a The PR3/PRTN3 activity in urines from three patients displaying subclinical rejection at the time of sample collection. b Samples of the same urines as a) (lanes 1, 2, 13) or purified enzyme (lane PRTN3) were SDS PAGE separated, blotted and probed with anti- PR3/PRTN3. Molecular weight markers (lane M). c Purified proteinase 3 or d a proteinase 3 active subclinical urine were treated with DMSO alone (black square, black-up pointing triangle) or the PR3/PRTN3 specific inhibitor Bt-Pro-Tyr-Asp-(nor)Val^P(O-C₆H₄-4-Cl) 2 dissolved in DMSO (black circle). e Aliquots of the inhibitor treated samples in c and d or an untreated sample of the same urine were separated by SDS-PAGE, blotted and probed with Alexa fluor™ 488-streptavidin. Lanes M) Molecular weight markers; (1) Inhibitor-labelled purified PR3/PRTN3; (2) Untreated urine; lane 4) Inhibitor-treated subclinical rejection urine (same urine as in d)