International Immunology, Vol. 13, No. 12, 1445-1452,
December 2001
© 2001 Japanese Society for Immunology
Expression, purification, characterization and clinical relevance of rAed a 1—a 68-kDa recombinant mosquito Aedes aegypti salivary allergen
Department of Pediatrics and Child Health, and
1 Department of Immunology, University of Manitoba, Winnipeg, Manitoba R3E 3P5, Canada
2 Department of Molecular Biology and Biochemistry, University of California, Irvine, CA 92697, USA
Correspondence to: Z. Peng, Department of Pediatrics and Child Health, University of Manitoba, 532 John Buhler Research Centre, 715 McDermot Avenue, Winnipeg, Manitoba R3E 3P5, Canada; E-mail: zpeng{at}ms.umanitoba.ca
Accurate diagnosis of mosquito allergy has been precluded by the difficulty of obtaining salivary allergens. In this study, we expressed, purified, characterized and investigated the clinical relevance of a recombinant Aedes aegyptisalivary allergen, rAed a 1. Two cDNA segments were ligated together to form the full-length Aed a 1 gene. rAed a 1 was expressed using a baculovirus/insect cell system, and purified using a combination of anion-exchange and gel-filtration chromatography. The purified rAed a 1 bound to human IgE, as detected by ELISA, ELISA inhibition tests and immunoblot analyses. Epicutaneous tests with rAed a 1 and a commercial whole-body Ae. aegyptiextract, and Ae. aegyptibite tests were performed in 48 subjects. Nine of 31 (29%) of the subjects with positive immediate bite tests also had a positive rAed a 1 immediate skin reaction and 32% had an positive immediate test to the commercial extract. Six of 33 (18%) of the subjects with positive delayed bite tests also had a positive rAed a 1 delayed skin reaction and 6% had a positive delayed test to the commercial extract. Furthermore, rAed a 1-induced flare sizes significantly correlated with mosquito bite-induced flare sizes. None of the subjects with negative bite tests had a positive skin test to rAed a 1 or to commercial extract. We conclude that the rAed a 1 has identical antigenicity and biological activity to native Aed a 1, can be used in the in vitroand in vivodiagnosis of mosquito allergy, and is more sensitive than mosquito whole-body extract for detecting delayed skin reactions.
Keywords: Aed a 1, baculovirus/insect cell expression system, cDNA, insect allergy, skin test
Transmitting editor: S. H. E. Kaufmann
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