Localization of the two major allergens in rye-grass pollen using specific monoclonal antibodies and quantitative analysis of immunogold labelling |
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Authors: | P E Taylor I A Staff M B Singh and R B Knox |
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Institution: | (1) School of Botany, University of Melbourne, 3052 Parkville, Victoria, Australia;(2) Botany Department, LaTrobe University, 3083 Bundoora, Victoria, Australia |
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Abstract: | Summary The intracellular localization of the two major allergens, Lol p I and Lol p IX, in rye-grass anthers was examined using monoclonal antibodies FMCA1 (specific for Lol p I) and FMCA7 (specific for Lol p IX) with immunocytochemical techniques and quantitative analysis. A newly developed anhydrous fixation technique in a mixture of glutaraldehyde, paraformaldehyde and 2, 2-dimethoxypropane followed by embedding in LR Gold resin resulted in both improved infiltration of pollen grains compared with existing techniques and the localization of these water-soluble antigens in their original sites compared with diffusion artefacts following aqueous methods. After anhydrous fixation, Lol p I was predominantly located in the electron-opaque regions of the cytosol of the vegetative cell of the tricellular pollen grains (24 counts m-2), whereas Lol p IX was detected mainly within starch granules (16 counts m-2). For both Lol p I and Lol p IX, similar labelling was detected in the cells of the endothecium and middle layer (18 counts m-2), but none was found in the tapetal cells or orbicules. |
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