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. 2010 Apr 12;285(24):18847–18857. doi: 10.1074/jbc.M109.098434

FIGURE 2.

FIGURE 2.

NDGGE analysis of apoA-I exchange. rHDL (apoA-I:A350) particles of 7.8-nm diameter were incubated at 37 °C in the presence (lanes 4a–6a) or absence (lane 3a) of a 5-fold molar excess of plasma-purified unlabeled lipid-free apoA-I. Equivalent amounts of rHDL-associated protein (1.5 μg) were separated from the lipid-free protein by electrophoresis on 4–20% nondenaturing Tris-glycine polyacrylamide gels. Fluorescently labeled protein was visualized using an UV trans-illuminator (excitation light, 365 nm; emission blue filter, 440–480 nm) (A). Total protein was detected with GelCode Blue staining (Pierce) (B). Lane 1, molecular weight markers (high molecular weight calibration kit from GE Healthcare). Lanes 2 and 3, rHDL (apoA-I:A350) stored at 4 °C or incubated at 37 °C for 5 h, respectively. Lanes 4–6, incubation mixtures of rHDL (apoA-I:A350) and a 5-fold molar excess of plasma-purified lipid-free apoA-I at 5 h, 24 h, and 7 days, respectively. Lanes 7, lipid-free apoA-I:A350 incubated with a 5-fold excess of lipid-free plasma-purified unlabeled lipid-free apoA-I and incubated at 37 °C for 5 h. The relative band intensities were estimated by densitometry (Imagequant 5.0 software; Amersham Biosciences).