An immunoenzyme technique for the identification of granulocyte-macrophage colony-stimulating factor (GM-CSF) receptors using digoxigenated-GM-CSF.
A method for detecting granulocyte-macrophage colony-stimulating factor (GM-CSF) receptors has been devised using human macrophages and a GM-CSF/IL-3-dependent human megakaryoblastic leukemia cell line (M-07e). Recognition of the factor-binding site was accomplished by linking recombinant human (rh) unglycosylated GM-CSF previously labeled with digoxigenated compounds. Digoxigenates were able to link amino and sulphydryl groups of the soluble factor and an immunoperoxidase technique using monoclonal anti-digoxigenin antibody was employed to demonstrate the interaction. To support morphological data cross-linking analysis was performed with M-07e cells using digoxigenated-rh-GM-CSF. Macrophages and M-07e cells incubated with digoxigenated-rh-GM-CSF showed intense positivity by the immunoperoxidase technique. In cross-linking, M07e cells showed a 96 kDa band corresponding to receptor plus bound factor. This technique permits a high degree of specificity in the detection of GM-CSF receptors with good morphological preservation of cellular detail.
Roncaroli
F
F
Department of Pathology, Bellaria Hospital, University of Bologna, Italy.
Geuna
M
M
Reato
G
G
Ponti
R
R
Sanavio
F
F
Bellone
G
G
Palestro
G
G
D002478:Cells, Cultured; D004076:Digoxigenin; D004591:Electrophoresis, Polyacrylamide Gel; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D006801:Humans; D007124:Immunoenzyme Techniques; D008264:Macrophages / Q000737:chemistry; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor / Q000032:analysis*; D011994:Recombinant Proteins; D014407:Tumor Cells, Cultured
D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor; D011994:Recombinant Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D004076:Digoxigenin