Fig. 6.
Fig. 6. PAI-1 in control and QPD platelets. / (A) PAI-1 in control (C) and QPD (Q; 3 patients [Pt] are shown) samples was analyzed by Western blotting with rabbit anti–human PAI-1 after 10% nonreduced SDS-PAGE. Lanes compare 5 μL lysate (L) and ionophore releasates (R), 20 μL K562 media, and 5 μL control ionophore releasate, incubated with 0 to 400 ng/mL tcu-PA (CR + tcuPA), as indicated. Lane * shows the PAI-1 affinity purified from QL using monoclonal anti–u-PA. QPD platelet lysates contained PAI-1 in high–molecular weight complexes (bands near the 98-kd marker) that comigrated with the complexes generated by adding tcu-PA to control releasates. Proteolyzed PAI-1 (arrow) was detected in QPD platelet lysates and in long exposures (not shown) of control releasates incubated with tcu-PA. (B) u-PA–PAI-1 complex ELISA indicated that, unlike control samples, QPD platelet lysates and releasates were unable to generate additional u-PA–PAI-1 complexes in vitro with added (+) tcu-PA (data representative of 2 separate experiments with pooled samples).

PAI-1 in control and QPD platelets.

(A) PAI-1 in control (C) and QPD (Q; 3 patients [Pt] are shown) samples was analyzed by Western blotting with rabbit anti–human PAI-1 after 10% nonreduced SDS-PAGE. Lanes compare 5 μL lysate (L) and ionophore releasates (R), 20 μL K562 media, and 5 μL control ionophore releasate, incubated with 0 to 400 ng/mL tcu-PA (CR + tcuPA), as indicated. Lane * shows the PAI-1 affinity purified from QL using monoclonal anti–u-PA. QPD platelet lysates contained PAI-1 in high–molecular weight complexes (bands near the 98-kd marker) that comigrated with the complexes generated by adding tcu-PA to control releasates. Proteolyzed PAI-1 (arrow) was detected in QPD platelet lysates and in long exposures (not shown) of control releasates incubated with tcu-PA. (B) u-PA–PAI-1 complex ELISA indicated that, unlike control samples, QPD platelet lysates and releasates were unable to generate additional u-PA–PAI-1 complexes in vitro with added (+) tcu-PA (data representative of 2 separate experiments with pooled samples).

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