Allergenic relevance of Cupressus arizonica pollen extract and biological characterization of the allergoid.
BACKGROUND
Cupressaceae (cypress) pollens can cause pollinosis in winter. However, the lack of specific commercial extracts combined with the early pollination period of cypress trees make a precise diagnosis difficult. The need for a reliable and effective cypress extract for diagnostic and therapeutic purposes is increasingly felt.


METHODS
Mixed or single Cupressus arizonica, lusitanica and sempervirens pollen extracts precipitated with ammonium sulfate (PPT) were compared by direct RAST, RAST inhibition and SDS-PAGE techniques. The major allergen of C. arizonica (Cup a 1), purified by anion exchange chromatography, was checked by immunoblotting experiments before chemical modification, in parallel with a C. arizonica extract, with potassium cyanate (KCNO) to obtain a monomeric allergoid. The allergoid extract was characterized for its biological, chemico-physical and immunological features by RAST inhibition, SDS-PAGE and ELISA assays.


RESULTS
Direct RAST, RAST inhibition, and SDS-PAGE data indicated that the PPT C. arizonica pollen extract showed the most allergenic potential, and it can be considered representative of the Cupressus spp. Immunoblotting data confirmed Cup a 1 as a major allergen. RAST inhibition and ELISA showed that modified PPT C. arizonica extract had less IgE reactivity than the native, non-modified extract, while preserving the immunogenic capacity typical for an allergoid. Finally, the SDS-PAGE profile of Cup a 1 allergoid was similar to native Cup a 1 allergen, suggesting the modified C. arizonica extract shows the characteristics of a monomeric allergoid.


CONCLUSIONS
The PPT C. arizonica pollen extract shows good in vitro diagnostic potential and its chemically modified form offers the features of a monomeric allergoid. It might therefore lend itself to the development of a product to be administered by the sublingual or oromucosal route for immunotherapy of individuals with cypress pollinosis.
Mistrello
G
G
Lofarma S.p.A., Milan, Italy. scient@lofarma.it
Roncarolo
D
D
Zanoni
D
D
Zanotta
S
S
Amato
S
S
Falagiani
P
P
Ariano
R
R
D000485:Allergens / Q000302:isolation & purification*; D000918:Antibody Specificity; D052179:Antigens, Plant; D029781:Cupressus / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007073:Immunoglobulin E / Q000097:blood; D066298:In Vitro Techniques; D010940:Plant Proteins / Q000302:isolation & purification*; D011058:Pollen / Q000276:immunology*; D011852:Radioallergosorbent Test; D006255:Rhinitis, Allergic, Seasonal / Q000175:diagnosis / Q000276:immunology
D000485:Allergens; D052179:Antigens, Plant; C419643:Cup a 1 protein, Cupressus arizonica; D010940:Plant Proteins; D007073:Immunoglobulin E