Aldehyde-fixed human red cells have previously been used to assess the roles of electrostatic and electrodynamic forces in adhesion. We have attempted to test the prediction that enzymic removal of cell surface negative charges should increase adhesion in dilute salt solutions by reducing electrostatic repulsion. While this is indeed the case for neuraminidase-treated cells and also for Pronase- and trypsin-treated cells over much of the low ionic strength range, the latter two treatments cause very strong adhesion over a remarkably narrow range of ionic strength centred on 1 mM-NaCl. At 0.5 and 1.5 mM adhesion is negligible. After Pronase treatment a further adhesive peak occurs at 2.5 mM. Electrophoresis of protease-treated cells shows small but clear reductions in mobility at precisely these peak adhesion values. These electrophoretic potential changes are almost certainly not large enough to cause increased adhesion directly, and it is thought that they are second-order changes, symptomatic of a structural rearrangement of the cell surface. How this causes such vastly augmented adhesion is an intriguing problem.
The incorporation of a low ionic strength solution (LISS) in the micro-elution technique used for the detection of blood group antigens in stains markedly improves the test's sensitivity. This is because LISS increases the amount of antibody taken up by the antigen in the stain which results in a greater yield of antibody recovered from the slain by elution. LISS also enhances the activity of the eluted antibody if it is introduced as a suspension medium for the red cells used to detect the antibody. The introduction of suitably diluted AB serum as diluent when testing the eluates is an additional advantage. The improvement in the sensitivity of the micro-elution technique is great enough in some instances to allow the detection of an antigen in a stain which is undetectable in the absence of LISS. Moreover some doubtful positive reactions are enhanced sufficiently for the presence of an antigen to be definitely established.
Abstract
1. Treatment of normal human red cells with various proteolytic enzymes, cholera vibrio filtrate, influenza virus, and periodate ion results in red cells susceptible to acid hemolysis in compatible normal human serum.
2. The kinetics of hemolysis of these artificially altered red cells resembles in many respects those observed in paroxysmal nocturnal hemoglobinuria (PNH).
3. While the observed differences in behavior between these artificially altered cells and PNH cells do not allow direct comparison, it is felt that these models may offer some clues in the understanding of PNH cell hemolysis, and some insight into the nature of the red cell lesion in paroxysmal nocturnal hemoglobinuria.
Abstract
C3b was bound to human red cells when serum complement was activated by addition of antibodies directed against different red cell antigens, and the rate of cleavage to C3dg was determined by assay for loss of bound C3c antigens using radiolabeled monoclonal anti-C3c. When C3b was bound by antibodies to antigens on branched-chain glycoproteins, cleavage to C3dg occurred more rapidly than when C3b was bound by antibodies to antigens closer to the red cell lipid bilayer. The rate of cleavage to C3dg also correlated directly with the number of complement receptors (CR1) per red cell, reflecting their role as cofactors in the cleavage of iC3b by factor I. Thus, the life span of C3b/iC3b on human red cells, which may be important for determining the rate and mechanism of clearance of C3-coated red cells, appears to depend on the CR1 status of the red cells and the characteristics of the antigen sites around which complement is bound.
Haemolysis and Reversible Agglutination of Trypsinized Normal Red Cells by a Normal Human Serum