Human placental conditioned medium contains monocyte-derived recruiting activity (MRA)

Monocyte-derived recruiting activity (MRA) stimulates the release of granulopoietic colony-stimulating factors (CSF) by endothelial cells. We carried out studies designed to test the hypotheses that human placental conditioned medium (HPCM), a widely utilized source of CSA for in vitro studies, contains both MRA and CSA and that these molecules could be separated on the basis of their isoelectric points. In five separate studies, concentrated samples of HPCM were chromatofocused and fractions tested in bioassays for MRA and CSA. We found that HPCM contains both MRA and CSA, that the majority of the MRA eluted at a pH of from 7.6 to 8.2 and was 2000-fold purified. CSA eluted at pH 5.6 or below. No CSA was detectable in the major MRA peak. We conclude that HPCM contains heterogeneous granulopoietic activities, that the isoelectric points of MRA and CSA are substantially different and that chromatofocusing provides a rapid single-step method for separating these two distinct granulopoietic factors from complex conditioned media.