Development of enzymo-immunoassays (EIA) for macrophage colony-stimulating factor (M-CSF) and leukaemia inhibitory factor (LIF) by using the same capture and signal generating polyclonal antibody.

Specific high titre polyclonal antibodies rapidly obtained by intralymphnode immunization of rabbits with recombinant M-CSF and LIF (< 60 micrograms/animal) have been used to develop specific, accurate and sensitive EIAs. The same batch of purified anti-M-CSF or anti-LIF Igs has been used for the coating of 96-well plates (capture antibody) and for the quantitative detection of the bound cytokine molecules (soluble biotinylated Igs). The sensitivity (M-CSF: 10 IU/ml, LIF: 20 pg/ml), accuracy (intra-assay-CV: 8.2 to 12.8% for M-CSF; 0 to 19.9% for LIF) and reproducibility (inter-assay-CV: 7.9 to 13.6% for M-CSF; 4.9 to 17.5% for LIF) are equivalent to those for previously published RIAs or EIAs. These assays are highly specific since 11 other cytokines (Epo: 3 IU/ml; G-CSF: 100 IU/ml; CNTF, OSM, SCF, IL-1 beta, IL-2, IL-3, IL-6, IL-11, IL-13: 5 ng/ml) tested in both EIAs were not detectable. Finally, the M-CSF and LIF concentrations measured in various biological fluids were found to be similar to those measured by us and others with different assays. In conclusion, the methodology used for M-CSF and LIF EIAs presented in this work represents a valuable approach for most cytokines, particularly when they are still available in reduced amounts.