Amplification of free thiol content within TRX-1–treated nβ2GPI by HUVECs. (A-B) nβ2GPI (1μM) was pretreated with DTT (35μM) activated TRX-1 (1.75μM) for 1 hour and then incubated with HUVECs or empty wells. (C-D) rβ2GPI (1μM) was pretreated with TRX-R (10nM)/NADPH (200μM) activated TRX-1 (1.75μM) for 1 hour and then incubated with HUVECs or empty wells. The supernatant from each well was then labeled with MPB, transferred to a PVDF membrane, and probed with streptavidin–horseradish peroxidase (HRP). This confirmed that HUVECs are also capable of amplifying the free thiol content of (A-B) nβ2GPI pretreated with DTT-activated TRX-1 (mean enhancement ± SD, 58.1% ± 32.5%; *P ≤ .04; n = 3) and (C-D) rβ2GPI pretreated with TRX-R/NADPH–activated TRX-1 (207.6% ± 146.4%; *P ≤ .03; n = 4). Efficiency of MPB labeling of DTT (35μM) activated TRX-1 (1.75μM) was shown to be unaffected whether performed under argon or air; n = 2 (E-F).