FIGURE 1.

Commercial human A2M has no detectable contaminants. A, human A2M (100 μg; Sigma) was separated on a Superose 6 size exclusion column. There is only one protein peak present, representing the A2M. Fractions of 0.5 ml were collected and then subjected to SDS-PAGE followed by silver staining (fractions from below the horizontal line are represented in B). B, silver-stained SDS-polyacrylamide gel showing proteins from fractions collected at 8.16 ml through 12.16 ml. Each fraction was 0.5 ml. Only A2M from under the peak is visible on the gel. C, SP-D binds only to fractions containing A2M. Fractions from the separation (samples from every 0.5 ml of eluate, boxed lanes in B) were coated on ELISA plates. SP-D was incubated with these samples in the presence of calcium (5 mm), and the amounts of SP-D that bound to these fractions were determined by standard ELISA procedure. SP-D binding profile corresponds to the fractions containing A2M. mAU, milliabsorbance unit.