A STUDY OF THE FORCES INVOLVED IN THE SECOND STAGE OF HEMAGGLUTINATION.

Abstract

Agglutination of antibody coated human erythrocytes has been found to depend on the zeta‐poten‐tial at the surface of shear and the dimensions of the antibody molecules. The critical zeta‐potential above which agglutination cannot occur has been shown to be —23 millivolts and —13 millivolts for saline and albumin antibodies respectively. A zeta‐potential below —18 millivolts for 19S molecules and below —8 millivolts for 7S molecules has been found to be necessary to give optimum agglutination and titer.The value of the enzymes used in blood group serology has been shown to be related to their ability to reduce the net surface charge density with a consequent reduction in zeta. It is postulated that this effect is due to esterase rather than protease activity.Bovine albumin and three synthetic polymers were found to bring about agglutination by reducing the zeta‐potential as a result of raising the dielectric constant. No evidence was found to support the hypothesis that they reduce the surface charge of the erythrocytes.A satisfactory equation for calculating zeta, that can be used to predict the course of a serological reaction, has been computed from a determined value of —3190 esu/cm2 for the net surface charge density of human erythrocytes in NaCl‐buffer at 25 C. This has been extended to include electrolytes of other valences by taking into account the mean activity coefficient of the electrolyte. Evidence is presented to show that the minimum zeta associated with erythrocyte stability is about —7 millivolts. This was considered to be the minimum potential required to overcome the cohesive forces of interfacial tension.In reaction mixtures imparting a zeta‐potential of —7 millivolts, tests with ten anti‐K and ten anti‐Fya sera show them to react similarly to anti‐D. There was no evidence to support the claim that these antibodies are monovalent.