[A new counting method for airborne Japanese red cedar and grass pollen allergens by the immunoblotting technique]

We devised a new counting method of pollen allergen particles which improved the fluorescence immunoblotting technique by Schumacher et al (1988). And by which airborne pollen allergens became visible under 10X magnifier or naked eyes. Airborne pollen allergens collected on the Burkard's sampling tape were transferred onto nitrocellulose membrane and were reacted with anti Cry j I rabbit serum or anti Lol p I rabbit serum, and then treated with alkaline phosphatase conjugated F(ab')2 anti rabbit IgG. Finally, bluish purple spots were obtained by staining with BCIP/NBT phosphatase substrate system. This technique does not require any skillful morphological observation, and is more suitable to measure the amounts of airborne pollen allergen for given pollinosis patients because total pollen allergen particles with common antigenicity are measured. In Japanese red cedar pollen counts, we could not count the spots more than 400 grains per 0.16 cm2 of the sample trapping area due to many overlapping spots. In this case, we tried to calculate the value from the ratio of bluish purple coloured area to one pollen area. However, a more suitable method for estimating the content of pollinosis caused airborne allergens may be colorimetric quantitation using densitometry and displaying the value as allergen content.