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Serum samples obtained from non-immunocompromised patients treated therapeutically with recombinant cytokines (e.g. Il-1alpha; Il-1beta; Il-2 to Il-18; IFNalpha; GM-CSF; G-CSF; etc.) often reveal the presence of high affinity anti-cytokine antibodies. Antibody Fab binding in a saturable manner by ELISA and RIA or western immunoblotting prove their specificity. Antibody level often increases in these patients with repeated cytokine administration, suggesting boosts of antigen stimulation. However, the appearance in circulation of auto-antibodies to exogenous cytokine is not always associated with a decreased clinical response to therapy. The demonstration that non-neutralizing auto-antibodies to several natural cytokines can be found even in sera of normal healthy individuals never treated before with cytokines and particularly during the last trimester of pregnancy and in cord-blood, suggests that these naturally- occurring and therapeutically-induced auto-antibodies may exert different functions, not only as inhibitors or antagonists but also as beneficial physiological cytokine carriers or regulators of their activity.

Serum samples were obtained from patients with lymphoid and plasma cell malignancies who received after chemotherapy human recombinant GM-CSF or G-CSF. Sera from some patients revealed the presence of anti-cytokine antibodies, particularly after repetitive cytokine injections. Antibody Fab binding in a saturable manner by ELISA and Western immuno-blotting confirmed antibody specificity. Anti-cytokine antibodies were detected before the exogenous cytokine injections in some patients, but increasing antibody levels were found after one or subsequent treatments. Low levels of anti-GM-CSF and anti-G-CSF antibodies were also detected in a relatively large proportion (about 10-30%) of normal sera from different adult healthy volunteers who had never been treated before with exologous cytokines as well as from cord blood. EBV-immortalized cord blood derived B-cell cultures were also found to produce anti GM-CSF and/or anti-G-CSF antibodies with high frequency.

Sera were obtained from two groups of patients. Group A included 7 patients with low-grade non-Hodgkin's lymphoma treated with three or more cycles of standard-dose chemotherapy and recombinant human granulocyte-colony stimulating factor (rhG-CSF). The cytokine was administered to half the patients after the first chemotherapy cycle and to the other half after the second according to a randomized design and then to all patients from the third chemotherapy cycle on, until documented hemopoietic reconstitution. Group B included 3 patients with high-grade non-Hodgkin's lymphoma, 1 patient with resistant Hodgkin's disease, and 1 patient with multiple myeloma who received high-dose chemotherapy and rhG-CSF. Anti-G-CSF antibodies were detected in the sera of 4 patients. Both immunoglobulin IgM and IgG antibodies were detected at low levels in pretreatment sera from one group A patient. IgG antibody titers increased markedly during the first and second periods of G-CSF administration. IgG class antibodies developed in 3 groups B patients during the first course of rhG-CSF administration. Circulating anti-G-CSF antibodies did not seem to affect hematological recovery. Low levels of anti-G-CSF antibodies were also detected in sera (15/135) from different healthy adults and in sera (5/40) from umbilical cord blood. Saturable antibody binding and competition enzyme-linked immunosorbent assay (ELISA) and immunoblotting confirmed antibody specificity.

Spleen leukocytes from rats and mice immunized against mouse 2.5S nerve growth factor (NGF) and peripheral blood leukocytes from rabbits hyper-immunized against the same antigen were fused with the mouse plasmacytoma P3X63Ag8. Hybridomas were screened by immunological assays (micro-complement fixation test and solid phase radioimmunoassay) for production of antibodies that reacted with NGF. Significant variations were seen between culture fluids from different hybrid cells. In addition, most but not all hybridoma antibodies that reacted immunologically with NGF prevented neurite outgrowth from 8-day chick embryo sensory ganglia explants after binding to NGF. These results suggest that the hybridoma antibodies produced by the different clones react with different antigenic sites on the NGF molecule.