Figure 2. Determining the size of active rat kidney nuclear and cytoplasmic ASPA. Pooled 150 mM NaCl anion-exchange chromatographed fractions were concentrated and injected into a size-exclusion column. Adjacent fractions corresponding to 500 µl/min elution were pooled and concentrated. A–B) Radiometric enzyme assay is described in Materials and Methods. The data are averages of two assays. Peak specific and total ASPA activity from both cytoplasmic (A) and nuclear (B) 150 mM NaCl anion-exchange fractions occur between fractions 36 and 40, corresponding to a MW of 38 kD as determined by standard curve (data not shown). Sizes (in kDa) of MW marker proteins are indicated by arrows. C) Western blot (1:2000 ASPA) of cytoplasmic protein (2.5 µg per lane) in size-exclusion chromatography fractions. D) Western blot (1:1000 ASPA) of nuclear protein (8 µg per lane) in size-exclusion chromatography fractions.