Inhibition of Hemagglutination

Pepper (1968) investigated the role of sialic acid in horse serum on the inhibition of hemagglutination by Asian influenza virus. Horse serum has N-acetyl, N-glycoyl, and the 4-0-acetylated derivatives of neuraminic acid (cf. Chapter 1). The biological activity of horse serum specifically toward the Az strain of influenza virus appears to be determined by the N,0-diacetylneuraminic acid component in c -ma-croglobulin. 

As early as 1964, Perona et a/., observed that sialidase treatment of erythrocytes resulted in a decrease in half-life, as determined by loss of Cr-labeled cells from the circulation. However, the mechanism of this reduction was not determined. See Chapter 7. 

Morell et al. (1966) observed that a radioactive preparation of asialoceruloplasmin upon injection into rabbits was cleared from the circulation in a matter of minutes, vastly more rapidly than native ceruloplasmin, whose half-life is about 56 hr. In subsequent work (Morell et al., 1968) they observed that the rapid disappearance of the asialoceruloplasmin from the serum was accompanied by an equally rapid accumulation of radioactivity within the liver, specifically in the lysosomal fraction of the parenchymal cells (Gregoriadis et al., 1970). 

Woodruff and Gesner (1968) studied the effect of sialidase treatment on Cr-labeled rat lymphocytes. They found that sialidase treatment caused a change in the cells electrophoretic mobility and in agglutinabil-ity. Upon injection of the sialidase-treated Cr-labeled lymphocytes into the rat, it was observed that the asialolymphocytes were rapidly taken up by the liver, with very few accumulating in lymphoid tissues. These results led them to conclude that the sialic acid components of the cell 

Regoeczi et al. (1974) were able to find a 15% and a 29% increase in the rate of removal of homologous asialotransferrin preparations from the plasma of rabbits and humans, respectively. However, they observed that when human asialotransferrin was injected into rabbits, it was removed 3.5 times more rapidly than the homologous asialotransferrin. Removal of normal human transferrin from rabbit plasma was not significantly different from the normal rabbit compound. In subsequent work (Regoeczi and Hatton, 1974) was found that progressively desialylated human transferrin molecules had plasma half-lives inversely proportional to the loss of sialic acid. 

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SCHEME 3. Tetrameric galabioside having an IC50 of 2 nM in hie inhibition of hemagglutination of human erythrocytes by S. suis 2... 

Studies on the carbohydrate-binding specificity of the fava-bean lectin, as determined by hapten inhibition of hemagglutination,140,213,466-469 showed that the lectin is inhibited by Makela s group III sugars (see Table VI). Hemagglutination by the lectin was inhibited by... 

Characterization of the carbohydrate-binding specificity of Ulex europeus II is still very rudimentary. Studies by inhibition of hemagglutination have been completed, using only a few of the appropriate sugars necessary to define specificity of the binding. 

It is also conceivable that steric stabilization plays a role in the inhibition of hemagglutination by the longer polymers although it should be noted that the polymers generated via ROMP are considerably smaller than the acrylamide- based polymers that benefit from the steric mechanism. Moreover, the assays for the ROMP-derived ligands do not involve direct binding to a surface-immobilized receptor. It is also possible that the ability of ligands to cluster multiple copies of ConA is a key factor in their potency. 

B. DETERMINATION OF CARBOHYDRATE SPECIFICITY BY HAPTEN INHIBITION OF HEMAGGLUTINATION... 

Discussion of the effect of ligand structure on protein-carbohydrate affinity requires an evaluation of complex stability constants. A munber of biophysical techniques are appropriate for the study of protein-carbohydrate interaction many of the more enlightening strategies are the topics of separate chapters elsewhere in this volume. We describe below three techniques used extensively in glycobiology— inhibition of hemagglutination, enzyme-linked lectin assay (ELLA), and isothermal titration microcalorimetry—and we consider the types of information provided by each technique in order to facilitate appropriate interpretation of the data. 

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