FIGURE 1.

PFL-AE and PFL binding kinetics using surface plasmon resonance under anaerobic conditions. PFL-AE was covalently attached to a CM5 biochip using thiol-coupling procedures. The analyte PFL was injected at a rate of 30 μl/minute in 20 mm HEPES, 10 mm NaCl, pH 7.4 at 25 °C for 180 s to measure association rates. Experiments were run in triplicate using five experimental injections of 0.123, 0.370, 1.11, 3.33, and 10 μm PFL dimer (arrows indicate the start of each injection) in single cycle kinetics mode. At the end of each injection of PFL, buffer was injected for 60 s to dissociate the analyte; these regions show a decrease in response. The green trace corresponds to experimental data, and the black trace corresponds to a 1:1 Langmuir interaction model fit of the experimental data. At the end of each cycle, PFL-AE was regenerated using 20 mm HEPES, 500 mm KCl, 0.005% polysorbate 20, 200 mm imidazole, pH 7.4, for 180 s, which completely removed the bound PFL. RU, resonance units.