New sensitive detection method for lectin hemagglutination using microscopy.

Abstract

The blood group system AB0 is determined by the composition of terminal oligosaccharides on red blood cells. Thanks to this structural feature, these groups can be recognized by saccharide-recognizing compounds. Lectins are proteins that are able to reversibly bind saccharide structures. They generally occur as multimers and are known as hemagglutination agents. Hemagglutination is a process in which blood cells are cross-linked via multivalent molecules. Apart from lectins, hemagglutination can also be caused by antibodies or viruses. A hemagglutination assay is commonly used for the detection of multivalent molecules that recognize blood cells, in order to search for their sugar specificity. It is traditionally performed on a microtiter plate, where the lectin solution is serially diluted and the lowest concentration of lectin causing agglutination is detected. This experimental set-up is utilized further for testing lectin specificity via a hemagglutination inhibition assay. We have developed a new way of detecting hemagglutination using microscopy, which was tested on purified lectins as well as cell lysates. Hemagglutination was performed on a microscope slide directly and detected using a microscope. Comparison with the standard hemagglutination assay using microtiter plates revealed that microscopic approach is faster and more robust and allows fast determination of lectin activities immediately in bacterial cytosols.