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Binding of macrophage colony-stimulating factor to serum proteins
1Division of Hematology, Department of Internal Medicine, Jichi Medical School, Tochigi, Japan.
Abstract:
Although three molecular forms of macrophage colony-stimulating factor (M-CSF) have been reported, Western blot analysis of immunoaffinity-purified M-CSF from human blood has shown that the major species of M-CSF in the serum has a molecular weight (MW) of 85 kD. Superose-12 gel filtration chromatography of immunoaffinity-purified serum M-CSF showed the presence of M-CSF-positive fraction in a higher MW area compared with the elution profile of recombinant human (rh) 85-kD M-CSF. Western blot analysis of the higher MW fraction showed that the M-CSF was the same as rh 85-kD M-CSF (not proteoglycan form of M-CSF), indicating that, in the serum, M-CSF exists bound to some serum proteins. To detect the serum proteins, we performed M-CSF-bound column chromatography. The eluate contained at least three serum proteins including albumin and IgG. This result was supported by chromatography using biotinylated M-CSF and avidin-agarose. The binding between rhM-CSF and albumin or IgG was also demonstrated by high-performance liquid chromatography (HPLC) fractionation of the mixture and enzyme-linked immunosorbent assay (ELISA) of the fractions. In the serum, a fraction of M-CSF seems to be complexed with serum proteins such as albumin and IgG.
Insights
Macrophage colony-stimulating factor (M-CSF) in human serum primarily exists as an 85-kD form bound to serum proteins like albumin and IgG, not as a proteoglycan. This binding influences its molecular weight and behavior in biological systems.
Area of Science:
- Immunology
- Molecular Biology
- Biochemistry
Background:
- Macrophage colony-stimulating factor (M-CSF) is crucial for hematopoiesis.
- Three molecular forms of M-CSF have been reported, but their in-vivo state is unclear.
Purpose of the Study:
- To investigate the molecular form and serum binding of M-CSF in human blood.
- To identify serum proteins interacting with M-CSF.
Main Methods:
- Immunoaffinity purification and Western blot analysis of human serum M-CSF.
- Superose-12 gel filtration chromatography.
- M-CSF-bound column chromatography and chromatography with biotinylated M-CSF.
- High-performance liquid chromatography (HPLC) and enzyme-linked immunosorbent assay (ELISA).
Main Results:
- The major M-CSF species in human serum is an 85-kD form.
- Serum M-CSF exhibits a higher molecular weight than recombinant 85-kD M-CSF, indicating complex formation.
- M-CSF binds to serum proteins, including albumin and immunoglobulin G (IgG).
Conclusions:
- In human serum, M-CSF exists primarily as an 85-kD protein complexed with other serum proteins.
- Albumin and IgG are identified as binding partners for M-CSF in serum.