Different rosette assays for detecting Fc receptor-bearing lymphocytes measure different subpopulations

Abstract

The capacity of purified lymphocytes from human peripheral blood to bind the Fc portion of IgG was investigated by the rosette technique using ox erythrocytes sensitized with rabbit anti-ox IgG (EA_ox) and human erythrocytes sensitized with anti-CD IgG (EA_CD). With unfractionated lymphocytes EA_ox always detected more rosette-forming cells (RFC) than did EA_CD; however, in lymphocyte populations specifically depleted of B lymphocytes by passage through copolymer styrene bead columns, the proportion of rosettes formed with EA_ox and with EA_CD was the same. Double labelling for Fc receptors and surface immunoglobulin (SIg) demonstrated that most of the lymphocytes which formed rosettes with either EA_ox or EA_CD also carried SIg. Pre-incubation of lymphocytes at 37° to remove heatlabile SIg did not affect their ability to form EA rosettes but reduced the proportion of SIg-bearing cells. Following pre-incubation a significant proportion of EA_ox RFC still remained SIg positive but the lymphocytes which formed rosettes with EA_CD no longer carried SIg.

These studies suggest that rosette formation with EA_CD detects only `K' lymphocytes (non-T, non-B cells bearing heat-labile SIg) while EA<sub>ox</sub> detects some B lymphocytes as well. By reducing the amount of IgG bound to ox erythrocytes sensitivity was reduced to the point where EA<sub>ox</sub> no longer formed rosettes with B lymphocytes but still detected `K' lymphocytes indicating either a qualitative or quantitative difference between the Fc receptors on B and `K' lymphocytes. Treatment of lymphocytes with trypsin decreased the percentage of rosettes formed with EA<sub>ox</sub> but not with EA<sub>CD</sub> supporting the conclusion that there is a structural difference between the Fc receptors on B and `K' lymphocytes although a difference in receptor density is not excluded.

When EA_CD and fluorescein-labelled ox erythrocytes sensitized with a low concentration of rabbit anti-ox IgG were mixed, most RFC bound both the indicator erythrocytes simultaneously suggesting that the Fc receptors on `K' lymphocytes do not exhibit species specificity.