Lectin histochemistry to detect altered glycosylation in cells and tissues.
Lectins are naturally occurring carbohydrate-binding molecules. A very wide range of purified lectins are commercially available which exhibit a diversity of carbohydrate-binding preferences. They can be used in the laboratory to detect carbohydrate structures on, or in, cells and tissues in much the same way that purified antibodies can be employed to detect cell- or tissue-bound antigens using immunocytochemistry. As lectins can distinguish subtle alterations in cellular glycosylation, they are helpful in exploring the glycosylation changes that attend both transformation to malignancy and tumour progression. In this chapter, methodologies are given for appropriate preparation of many types of cell and tissue preparations, including cells cultured on coverslips (which can be used for live-cell imaging), cell smears, and frozen (cryostat) and fixed, paraffin wax-embedded tissue sections. Heat- and enzyme-based carbohydrate retrieval methods are covered. Basic detection methods, which can be readily adapted to the researcher's needs, are given for direct (labelled lectin), simple indirect (labelled secondary antibody directed against the lectin), and avidin-biotin (biotinylated lectin) and avidin-biotin complex. The use of both the enzyme label, horseradish peroxidase, and fluorescent labels is considered.
Department of Biological & Medical Sciences, Oxford Brookes University, Oxford, Headington, UK. sbrooks@brookes.ac.uk
Susan A
SA
Brooks
Debbie M S
DM
Hall
D000818:Animals; D005629:Frozen Sections; D006031:Glycosylation; D006651:Histocytochemistry / Q000379:methods*; D006358:Hot Temperature; D006801:Humans; D037102:Lectins / Q000378:metabolism*; D016612:Paraffin Embedding; D009195:Peroxidase / Q000378:metabolism; D013194:Staining and Labeling; D016707:Tissue Fixation; D014357:Trypsin / Q000378:metabolism
D037102:Lectins; D009195:Peroxidase; D014357:Trypsin