Rapid assay and identification of human haemopoietic growth factors.
In order to facilitate the identification of human haemopoietic growth factors (HGFs), in complex conditioned media from limited cell sources, we have developed a method for the rapid assay of HGFs using bone marrow stem cells (BMSCs) obtained by Percoll density gradient enrichment of non-adherent cells from long-term human bone marrow culture. These BMSCs provide the basis for a 72 h assay where proliferation is triggered by the presence of GM-CSF, G-CSF and IL-3. The assay is up to 1000 times more sensitive than the conventional colony assay in detecting the presence of HGFs. We also describe methods for the rapid separation and identification of HGFs using fast protein liquid chromatography (FPLC) with phenyl-Superose and Mono Q columns. This enables individual HGFs present in complex conditioned media to be rapidly identified from the elution profile of HGF activity determined in the BMSC assay.
Pullman
W E
WE
Developmental Haematology Group, John Curtin School of Medical Research, Australian National University, Canberra.
Elsbury
S
S
Kobayashi
M
M
Hargreaves
J
J
Preston
L
L
van Leeuwen
B
B
Hapel
A J
AJ
D001681:Biological Assay; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D002853:Chromatography, Liquid; D003115:Colony-Stimulating Factors / Q000032:analysis* / Q000302:isolation & purification; D003470:Culture Media; D006412:Hematopoietic Stem Cells / Q000032:analysis; D006801:Humans; D007376:Interleukin-2 / Q000032:analysis; D007377:Interleukin-3 / Q000032:analysis
D003115:Colony-Stimulating Factors; D003470:Culture Media; D007376:Interleukin-2; D007377:Interleukin-3