Antigenic and allergenic changes during storage of a pollen extract

The changes that occur in the antigens and allergens of a timothy grass pollen extract during storage were measured by radioallergosorbent test (RAST) inhibition, isoelectric focusing, (IEF), crossed immunoelectrophoresis (CIE) and crossed radioimmunoelectrophoresis (CRIE) and a dye-binding protein assay. Storage intervals varied from 7 to 92 days at temperatures of 4 degrees, 22 degrees, and 35 degrees C and the freeze-dried extract was rehydrated and stored in either phosphate-buffered saline with 50% glycerol (PBSG) or Coca's solution (CS). For extracts stored in CS at 35 degrees C, allergenic activity as measured by RAST inhibition dropped by almost one half at 7 days and certain of the IEF, CIE, ad CRIE bands were no longer visible, showing that these properties correlated with allergenic activity. After 65 dys, no IEF bands were visible but three CIE bands could be visualized. At 22 degrees the changes were slower and at 4 degrees C no changes were observed for the 92-day storage time. Extracts stored in PBSG showed some of the same changes but were much more stable at all temperatures. During storage, the protein content as measured by dye binding was also reduced, showing tha proteins were fragmenting. Because some of the allergenic proteins disappeared more rapidly than others, there was not only a loss in total allergenic activity but also a change in specificity. This raises serious questions concerning the use of such extracts, especially for diagnosis.