Epstein-Barr virus in an American patient with Burkitt's lymphoma: detection of viral genome in tumor tissue and establishment of a tumor-derived cell line (NAB).

Epstein-Barr virus (EBV) DNA (17.7 genome equivalents/cell) was found in tumor tissue from an American patient with Burkitt's lymphoma who had never traveled outside the United States. A lymphoid cell line (NAB) containing the EBV genome was established from tumor tissue from this patient; characteristics of this cell line were described. Previous Burkitt's tumors found in Americans and examined by molecular hybridization were negative for EBV DNA. Our results suggested that EBV is associated with at least some American Burkitt's tumors.

Monoclonal antibodies to human melanoma-associated antigens: an amplified enzyme-linked immunosorbent assay for the detection of antigen, antibody, and immune complexes.

An amplified, indirect biotin-avidin micro-enzyme-linked immunosorbent assay was developed for the measurement of human melanoma-associated antigens, either free or circulating with associated immunoglobulin in patient sera. Parameters and specificity of detection were assessed using monoclonal antibody to human melanoma-associated antigens. The main advantages of the assay are its flexibility, through the use of indirect detection and a variety of formats, and its sensitivity, with a lower limit of antibody detection at 100 pg/well and a lower limit of soluble antigen detection at 10 pg/well. The assay was applied to cell surface antigen detection with monoclonal antibody 9.2.27 to a melanoma-associated antigen against a panel of glutaraldehyde-fixed cells, and gave similar binding specificity as assessed by a previous 125I-Protein A assay. Utilizing a unique "sandwich" format, aMr 100,000 melanoma-carcinoma-associated antigen was quantitated in melanoma patient sera and found highly elevated in Stage IV disease. The same sandwich format was also used to detect and determine the class of human immunoglobulin associated with circulating Mr 100,000 human melanoma-associated antigens in normal donor sera. Thus, the sensitivity and flexibility of this enzyme-linked immunosorbent assay system make it particularly suitable for numerous applications in the study of monoclonal antibody-defined tumor-associated antigens.

Monoclonal antibody defined-human melanoma-associated antigens: molecular and phylogenetic studies in normal serum.

Normal human sera were analyzed for the presence and molecular form of two human melanoma-associated antigens (MAAs), the 250 "melanoma-specific" glycoprotein and the 100 K "common tumor antigen". The 250 K MAA was not synthesized by any cultures other than human melanoma and was not detectable in normal human serum. In contrast, the 100 K MAA, which is present in spent medium of cultured human melanoma, carcinoma and fetal melanocytes but not of adult normal cells, was found in normal human serum in nanogram quantities. This serum form of the 100 K MAA was also found in pooled sera of higher apes but not of lower species. The cell-derived form of the 100 K MAA, present in spent culture medium, had a similar phylogenetic distribution. The molecule was produced by cultured brain glia from gorilla, but not by melanoma cells from miniature swine or dog. The 100 K MAA from gorilla glia had a mol. wt identical to the molecule produced by human melanoma cells. Molecular characterization of this MAA in normal human serum showed that it was heterogeneous in size and was present in fractions greater than 100 kd after analytical HPLC gel sieving under non-denaturing conditions. In contrast, MAA from spent culture medium of melanoma cells was 100 kd or less in chemically defined medium (CDM) with no protein supplement, but had a higher mol. wt in CDM with BSA or fetal calf serum supplement, similar to the serum form of the molecule. An association of the 100 K MAA with albumin was demonstrated by analytical HPLC gel sieving and SDS-PAGE analysis of monoclonal antibody immunoprecipitates. The 100 K MAA was dissociated from albumin in normal human serum by treatment with SDS and fractionation by gel sieving. Under these conditions 100 K MAA from serum co-migrated with similarly treated 100 K from melanoma cells. These results indicate that the 100 K MAA is a normal serum constituent which forms a strong, non-covalent association with albumin and is evolutionarily restricted to higher apes or humans.

Occult (non-surface expression) of T, B and monocyte markers in human large granular lymphocytes.

Human large granular lymphocytes were examined for non-surface expression with a panel of monoclonal antibodies to T cell, B-cell and monocyte markers. T101, antibody to the T65 antigen, showed binding to crude fractions containing intracellular membranes but not to immobilized whole cells. Non-surface expression of T65 was also demonstrated by flow cytometry using lysolecithin to transiently permeabilize cells. With the latter technique nonsurface expression was also demonstrated with monoclonal antibodies B2 and MO-2. T65 was shown to be synthesized by large granular lymphocytes by metabolic labeling, indirect immunoprecipitation and SDS-PAGE. T65 from large granular lymphocytes was the same mol. wt as antigen for T cells derived from the same donor. These results indicate that human large granular lymphocytes synthesize, but do not express on the surface, certain monoclonal antibody-derived markers heretofore considered specific for other cell lineages.