Production of a monoclonal antibody (IND.64) identifying a cell cycle‐associated antigen using spleen cells from nude mice bearing ichikawa tumour

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Abstract
Using spleen cells from athymic nude mice grafted with Ichikawa tumour, we have generated the monoclonal antibody IND.64, which detects a proliferation‐associated nuclear antigen. Immunoblotting analysis with IND.64 showed a double band with apparent molecular weights of 395 and 345 kD. In normal human tissues, the antigen detected by IND.64 was expressed only by the nuclei of proliferating cells, such as germinal centre cells of reactive lymph nodes, cortical thymocytes, the basal layer of the skin, and proiiferative compartments of the stomach, small intestine, and colon. IND.64 did not react with cells known to be non‐proliferative or to show only a low turnover, such as cells of the kidney, liver, smooth muscle, cardiac muscle, and brain. The expression of this antigen during the cell cycle was determined using two approaches: IND.64 immunostaining of synchronized adult bovine aortic endothelial cells and flow cytometric analysis of double‐labelled PHA‐stimulated peripheral mononuclear blood leucocytes with a DNA marker and IND.64. The antigen recognized by IND.64 was found to appear in the late G1 phase, and persisted in phases S, G2, and M, but was absent in the G0 and early Gl phases. IND.64 was further investigated in different tumour types to evaluate the correlation between the percentage of IND.64‐positive cells (IND.64 index) and the histological grade. In non‐Hodgkin's lymphomas, an excellent correlation was found between the percentage of IND.64‐positive cells and the cytomorphological grade. In nodular sclerosis and mixed cellularity Hodgkin's disease, a high number of Reed‐Sternberg cells were positive with IND.64. The non‐lymphoid neoplasms investigated showed a variable percentage of positive cells, IND.64 appears to be a promising tissue marker to complement the evaluation of prognosis in human cancer.

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