Evidence for membrane differentiation in polarised leucocytes: the distribution of surface antigens analysed with lg–gold labelling

Abstract

The distribution of a number of leucocyte surface antigens was studied on both round and polarised neutrophil or mononuclear leucocytes using Ig-gold conjugates with transmission electron microscopy. Thin sections of cells, which had been lightly fixed before antibody labelling, were analysed using a statistical method to determine: (1) whether the antigens had a non-random distribution or 'clustering' over the cell surface; and (2) whether there was any overall bias in labelling to give a polarised distribution. Comparison between the results of this analysis and cell morphology were made. The results indicated that with the antigens investigated here, CD45, CD15, HLA-DR and CR3, the majority of polarised cells had a calculated direction of overall asymmetry of gold particles that was aligned with the long axis of morphological polarity. Maximal asymmetry was seen in polarised cells labelled for CD45 and HLA-DR, with labelling ratios of up to 6:1 between the front and back of the cell. A number of round mononuclear cells demonstrated significant polarisation of gold particles but this had no apparent morphological correlation and, in general, round cells showed a low degree of asymmetry. However, there was evidence that both round and polarised cells had a non-random distribution or 'clustering' of gold particles, which was more marked in morphologically polarised cells and particularly significant in polarised neutrophil leucocytes labelled for CR3. The significance of these results for models of cell locomotion involving membrane flow is discussed.