OLE9 | Major pollen allergen Ole e 9

Varied amounts of recombinant OLE9 protein were loaded/well and separated on 10 % SDS-PAGE  and blotted 1h to PVDF (0.45 µm) using semi-dry transfer. Blot was blocked with 5 % milk for 2h/RT with agitation. Blot was incubated in the primary antibody at 2.8 µg/ml for 1h/RT with agitation in PBS-T. The antibody solution was decanted and the blot was rinsed briefly twice, then washed 4x in PBS-T at RT with agitation. Blot was incubated in matching secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated) diluted to 1:50 000 in  for 1h/RT with agitation. The blot was washed as above and developed with chemiluminescent detection reagent according to manufacture's instructions.