A monoclonal antibody (MAb NKI/C-3) produced against a purified membrane preparation of human melanoma cells reacts preferentially with sections of formaldehyde-fixed and paraffin-embedded tissues of melanoma, nevocellular nevi, carcinoids and medullary carcinomas of the thyroid. NKI/C-3 did not react with basal-cell carcinoma, brain tissue or brain tumors, and in only 14/196 other tumors was a clear cross-reactivity observed, e.g. with prostate carcinomas and a minority of primary breast, ovarian, lung and clear-cell carcinomas. This antibody was used in an immuno-electron microscopic study for the cellular localization of the antigen. The antigen was dispersed in the cytoplasm of melanoma cells, and more concentrated inside vacuoles and sometimes also on the melanosomes. Occasionally, the antigen was seen on the cell surface. The nature of the antigen was determined in an enzyme immunoassay (EIA). It was found that the antigen is a glycoprotein with a disulfide-dependent configuration that is essential for recognition by the MAb. The antigen was distributed heterogeneously during gel filtration as well as during SDS-polyacrylamide gel electrophoresis in the region of 25-110 kd proteins. A purified antigen preparation that was obtained after affinity chromatography on a column of MAb NKI/C-3 linked to Sepharose 4B contained a carbohydrate:protein ratio of 1:3.5.

By means of a modified immune adherence (IA) technique, sera from melanoma patients were tested for the presence of antimelanoma antibodies. In total 13/73 sera tested showed a positive IA reaction of which 4/6 sera showed a positive reaction in the autologous situation. Sera from 33 patients with other tumors, 7 patients with non-neoplastic diseases and 50 healthy individuals did not show any IA reactivity towards melanoma cells. The reaction seemed to be selectively directed against tumor-associated antigens (TAA) on melanoma cells. No correlation with the stage of the disease could be found. Longitudinal studies indicated that conversions in antibody activity did not correlate with the clinical state of the patients. There was also no correlation with the corresponding in vitro data obtained in cell-mediated immunity tests. Cell lines and short-term cultures originating from tumors from different melanoma patients shared a common antigenicity. The expression of TAA on cells from a melanoma cell line fluctuated significantly during prolonged culture. The expression of TAA was influenced by the culture conditions and the growth state of the cells. A relation between TAA-expression and cell cycle phase was demonstrated.

No abstract

A post-microsomal fraction (Fraction X) was isolated from cell sap from rat liver by centrifugation a t 1050OOxg for 15 h. The activity of a number of aminoacyl-tRNA synthetases was considerably higher in the resulting pellet than in the supernatant. These enzymes (glutaminyl-, isoleucyl-, leucyl-, lysyl-, and methionyl-tRNA synthetase) were purified by gel filtration on Sephadex G-200, chromatography on DEAE-Sephadex A-50 and on hydroxyapatite. As compared with crude cell sap a 120-to 170-fold purification was achieved.I n all purification steps, including additional centrifugation on sucrose gradients or isoelectric focusing, the major peak of activity of the five enzymes coincided.I n the electron microscope circular and rectangular particles, 11-14nm in size, could be visualized. From these findings we suggest that in rat liver cells, if not in all animal cells, part of the aminoacyl-tRNA synthetases occur in a particulate state.

In order to investigate their possible role as prognostic markers, staining with the antibodies NKI/C-3 and anti-S100, that are applicable on paraffin sections, was examined using a group of primary cutaneous melanomas and autologous metastases using the immunoperoxidase procedure. All melanoma lesions stained with anti-S100, and the large majority with NKI/C-3. In primary melanomas showing a moderate or dense associated lymphocytic infiltrate, significantly more tumour cells stained with anti-S100 than in primary melanomas with a slight or absent infiltrate. In markedly pigmented metastases, significantly more tumour cells stained with NKI/C-3 than in less pigmented lesions; in primary melanomas this phenomenon just failed to be significant. In metastases with a high mitotic index a significantly lower proportion of tumour cells stained with NKI/C-3 than in lesions with a low mitotic index. No significant differences in staining were found between a group of primary melanomas with metastases and a group without metastases within a follow-up period of 5 years. Therefore, although staining with NKI/C-3 and anti-S100 appears to be associated with certain histopathological characteristics, it has no direct contribution to the assessment of prognosis in primary melanoma.