Isolation and characterization of Poa p I allergens of Kentucky bluegrass pollen with a murine monoclonal anti-Lol p I antibody.
The Poa p I allergens were isolated from the retentate fraction of a dialyzed preparation of an aqueous extract of Kentucky bluegrass pollen by means of a reverse immunosorbent prepared with a murine anti-Lol p I monoclonal antibody, Mab 290-A-167. By sodium dodecyl sulphate-polyacrylamide gel electrophoresis and preparative isoelectrofocusing, Poa p I was found to consist of a 35.8-kD component with an isoelectric point of 6.4 and a 33-kD component with one of 9.1 and designated as Poa p Ia (acidic) and Poa p Ib (basic), respectively. The relative protein content of these components was estimated from the intensity of the stained bands following sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Thus, Poa p Ia appeared to be the major protein constituent, and on Western immunoblot it also bound the monoclonal antibody to a greater extent than Poa p Ib. On the other hand, Poa p Ib was shown by Western immunoblot and autoradiographic analysis, to bind to a greater extent the IgE antibodies present in a pool of sera from grass-allergic individuals. Therefore, Poa p Ib was considered as the major allergenic component of Poa p I. By competitive inhibition of the radioallergosorbent test, it was demonstrated that the Mab inhibited the binding of Poa p I allergens to human IgE antibodies to the extent of 70%. Hence, it is suggested that Mab and human IgE antibodies recognize identical or closely related determinants of Poa p I allergens.
Lin
Z W
ZW
Department of Immunology, University of Manitoba, Winnipeg, Canada.
Ekramoddoullah
A K
AK
Kisil
F T
FT
Hebert
J
J
Mourad
W
W
D000485:Allergens / Q000276:immunology / Q000302:isolation & purification*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D000918:Antibody Specificity; D001345:Autoradiography; D001667:Binding, Competitive; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D007526:Isoelectric Point; D051379:Mice; D008807:Mice, Inbred BALB C; D010940:Plant Proteins; D006109:Poaceae / Q000276:immunology; D011058:Pollen / Q000032:analysis* / Q000276:immunology; D011852:Radioallergosorbent Test
D000485:Allergens; D000911:Antibodies, Monoclonal; D010940:Plant Proteins; C058089:Poa p I proteins, Poa pratensis