Due to their specificity, constant properties and virtually unlimited supply monoclonal antibodies have given an important stimulus to almost every field of biomedical research within the last 10 years. The generation of mouse monoclonal antibodies includes immunisation of mice followed by fusion of mice spleen cells with a murine myeloma cell line. With this procedure hybridomas secreting monoclonal antibodies of a predefined specificity can be obtained. For three major reasons we worked on the establishment of human hybridomas secreting specific antibodies: human antibodies are less immunogenic when used for diagnostic or therapeutic purposes, only human monoclonal antibodies allow the analyses of the human B cell repertoire, there is evidence that human monoclonal antibodies recognise epitopes different from those seen by murine monoclonal antibodies. Therefore, we set out to generate human B cell hybridomas by cell fusion using the human lymphoblastoid B cell line Wi-L2-729 HF2 and lymphocytes from melanoma patients. The lymphocytes were isolated from tumour-draining cervical lymph nodes, stimulated with pokeweed mitogens plus the autologous tumour cells in an enriched tissue culture medium and fused in the presence of polyethylene glycol. Supernatants of hybridomas were screened in a single cell immunosorbent assay with either autologous melanoma cells or established melanoma cell lines fixed to the bottom of Terasaki plates or on cytospin preparations of these cells using the immunoperoxidase staining procedure. We could demonstrate that the tumour draining lymph nodes of these melanoma patients contained B lymphocytes capable of producing antibodies reacting with the tumour cells.(ABSTRACT TRUNCATED AT 250 WORDS)