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Some Ultrastructural Features of the Cell Surface after SV40 Transformation and Somatic Hybridization with Normal Untransformed Cells¹

The Wistar Institute of Anatomy and Biology, Philadelphia, Pennsylvania 19104

We have studied the surface properties of four cell lines: normal human fibroblasts (GM-177), SV40-transformed Lesch-Nyhan fibroblasts (LN-SV), mouse macrophages, and somatic cell hybrids between mouse macrophages and human SV40-transformed fibroblasts (LN-SV). The concanavalin A (Con A)-peroxidase method was used to visualize the localization of the Con A binding sites by means of electron microscopy. The mean thickness of the reaction product was calculated and the distribution pattern was visualized by drawing "glycograms," from which the ratio of covered to uncovered areas was measured.

Ruthenium red technique was used to study the ability of the cell surface to retain ruthenium red-positive materials after rinsing in phosphate-buffered saline. Before rinsing, there were no differences between thicknesses of the ruthenium red-positive surface layers of normal and virus-transformed Lesch-Nyhan fibroblasts. After rinsing, however, significant differences in the ability to retain ruthenium red-positive material were seen in favor of the SV40-transformed Lesch-Nyhan fibroblasts (54% over 35%).

Before rinsing, significant differences in the thickness of the ruthenium red-positive material were found between mouse macrophages and the hybrids between mouse macrophages and SV40-transformed human fibroblasts, with the hybrids expressing a thicker ruthenium red-positive layer than did the parental mouse macrophages. After washing, however, the ability to retain this material was greater in mouse macrophages than in hybrids. The hybrid cell surface resembles that of SV40-transformed human Lesch-Nyhan fibroblasts.

The Con A-peroxidase method shows a different distribution of the reaction product. In general, in all four cases the central portions of the cells had less continuous labeling patterns than did their peripheries, while the marginal portions were very often devoid of the reaction product.

Normal human fibroblasts had the most continuous surface reaction product, the thickest reaction product, and the greatest Con A:gap ratio. In all four of these measurements, the hybrid cell line expressed great similarity to the parental cell lines, especially to the mouse macrophages.

¹ Supported in part by USPHS Research Grants CA-10815 from the National Cancer Institute and RR05540 from the Division of Research Resources.

² WHO fellow of the Wistar Institute from the Department of Comparative Anatomy, Jagellonian University, Krupnicza 50 Krakow, Poland.

Received 1/ 6/75. Accepted 6/16/75.