Stimulation of mouse bone marrow cell membrane glycoconjugate synthesis by colony stimulating factor (CSF): basis for a rapid, simple assay for CSF.
A new method is presented for assaying mouse colony stimulating factor (CSF) based on the presumption that one of the earliest events after the induction of differentiation or maturation is an increase in synthesis and/or turnover of glycoconjugates in the target cell plasma membranes which may be detected as an increase in incorporation of [3H]galactose into acid precipitates of these cells. Using mouse bone marrow cells cultured in microtiter plates, we show that addition of CSF indeed results in dose-dependent stimulation of incorporation of galactose into their membrane glycoconjugates. Maximum stimulation of galactose incorporation occurs between 16 and 24 h of culture in the presence of CSF. A comparison of the standard colony test with the galactose incorporation assay showed similar dose-response patterns with 2 different CSFs. In a 3-step separation of bone marrow cells, there was a parallel enrichment of target cells for both assays. Furthermore, [3H]galactose-labeled marrow cells from our assay formed a discrete subpopulation which banded at a density of 1.065 g/cm3 on a linear gradient, and which contained all the identifiable CFU-c when further cultured in soft agar. The galactose incorporation assay does not require special medium or serum, and is simpler and quicker than the colony test. In comparative experiments it is demonstrated that this new 24 h assay is also reliable for screening CSF during purification procedures.
D000818:Animals; D001853:Bone Marrow / Q000378:metabolism; D001854:Bone Marrow Cells; D003114:Colony-Forming Units Assay; D003115:Colony-Stimulating Factors / Q000502:physiology*; D003470:Culture Media; D005690:Galactose / Q000378:metabolism; D006023:Glycoproteins / Q000096:biosynthesis*; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism; D046911:Macromolecular Substances; D008297:Male; D008565:Membrane Proteins / Q000096:biosynthesis*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA
D003115:Colony-Stimulating Factors; D003470:Culture Media; D006023:Glycoproteins; D046911:Macromolecular Substances; D008565:Membrane Proteins; D005690:Galactose