Lectins agglutination assay

Hynes, S.O., Hirmo, S., Wadstrom, T., Moran, A.P. Differentiation of Helicobacter pylori isolates based on lectin binding of cell extracts in an agglutination assay. J Clin Microbiol 37 (1999) 1994-1998. 

Detecting and assaying lectins are made possible by the ability of lectins to agglutinate red blood cells. The specificity of lectins is in terms of the simple sugars that inhibit the agglutination assay. Some lectins, of course, recognize somewhat more complex carbohydrate structures. Table 2 gives alist of some lectins to illustrate the features of this class of proteins. 

Figure 2. Agglutinating effect of increasing dilutions of the crude extracts from the pedicellariae of T pileolus and plant lectins. The assay of agglutinating activity with mouse erythrocytes was carried out using a 2.5 % suspension in PBS. G-type, the crude extract (5 pg/ml) from the giant type pedicellariae N-type, the crude extract (5 pg/ml) from the normal type pedicellariae Control, PBS WGA, wheat germ agglutinin (50 pg/ml) Con A, concanavalin A (100 pg/ml).

The agglutinating activity was assayed by using horse or mouse erythrocytes in microtiter plates. In some experiments, human erythrocytes was also used. 25 pi of 2.5 % (v/v) suspension of erythrocytes in 6.4 mM phosphate buffer saline, pH 7.2 (PBS) was added to 50 pi of serial 2-fold dilutions of the lectin fractions in PBS. The plates were incubated at room temperature for 1 hr. The results were expressed as the minimum concentration of lectin fractions (pg/ml). The agglutination inhibition was expressed as the minimum concentration of each sugar required for inhibition of the lectin fractions. 

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