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 iso-electric point of 6.4 and a 33-kD component with one of 9.1 and designated as Poa p la (acidic) and Poa p lb (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 la appeared to be the major protein constituent, and on Western immunoblot it also bound the monoclonal antibody to a greater extent than Poa p lb. On the other hand, Poa p lb 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 lb 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.

Allergen Poa p I isolated from the dialysed aqueous extract of Kentucky blue grass pollen by affinity chromatography with an anti-Lol p I murine monoclonal antibody (MAb) 290A-167 was previously shown to consist of a 35.8-kilodalton (kD) component with a pI of 6.4, designated as Poa p Ia, and a 33-kD component with a pI of 9.1, designated as Poa p Ib. The present study reports on the comparative antigenic analyses of these two components, using MAbs produced separately against Poa p I and Lol p I. Thus, anti-Poa p I MAbs 60 and 61 and anti-Lol p I MAb 290A-167 recognized Poa p Ia and Poa p Ib whereas anti-Poa p I MAbs 62, 63 and 64 and anti-Lol p I MAb 348A-6 recognized only Poa p Ia. The specificities of the MAbs were further resolved by comparing their respective abilities to inhibit the binding of ¹²⁵I-Poa p I or ¹²⁵I-Lol p I to the different MAbs prepared in the form of solid phase. These studies revealed that at least 4 distinct epitopes (designated as E₁, E₂, E₃ and E₄) were shared by both Poa p I and Lol p I. All 4 epitopes were present on Poa p Ia whereas only E₁ and E₃ were detected on Poa p Ib. E₁ was recognized by MAbs 60 and 61, E₂ by MAbs 62, 63 and 64, E₃ by MAb 290A-167 and E₄ by MAb 348A-6. The observations that MAb 290A-167 inhibited significantly the binding of human IgE antibodies to Poa p I and Lol p I were interpreted to indicate that epitope E₃ may also constitute an allergenic site on both Poa p I and Lol p I. These studies revealed that Poa p I and Lol p I share extensive antigenic and allergenic cross-reactivities with one another.