Isolation of Human scFv Antibodies Specific for House Dust Mite Antigens from an Asthma Patient by Using a Phage Display Library.
- Author:
Wang lim JUNG
1
;
Hee kyung LEE
;
Tae Soon YONG
;
Sang Hoon CHA
Author Information
- Publication Type:Original Article
- Keywords: Phage display library; scFv; asthma; type-1 hypersensitivity; house dust mite; allergen
- MeSH: Allergens; Antibodies; Antibody Formation; Asthma*; Bacteriophages*; Clone Cells; Dust*; Genes, Immunoglobulin; Hand; Humans*; Immunoglobulins; Lymphocytes; Mites; Polymerase Chain Reaction; Pyroglyphidae*; Sensitivity and Specificity; Sequence Analysis; Single-Chain Antibodies*
- From:Immune Network 2002;2(2):91-95
- CountryRepublic of Korea
- Language:English
- Abstract: BACKGROUND: In order to characterize human antibodies with specificity for mite allergens at the molecular level, a scFv phage display library was constructed using peripheral blood mononuclear lymphocytes from an asthma patient allergic to mite as Ig gene sources. METHODS: Immunoglobulin VH and V gene fragments were obtained by polymerase chain reaction, and randomly combined in pCANTAB-5E vector. The resulting human scFv phage display library had 3 X 10(4) independent clones, and biopanning was performed with house dust mite extracts. RESULTS: Four scFv clones specific for house dust mite extract were isolated. Immunoblot assay showed that our clones reacted to 25 kDa and 50~60 kDa proteins with unknown identity in mite extracts. Sequence analysis indicated that two clones (b7 and c15) are identical, and all clones belong to human VH3 subgroup. On the other hand, light chain usage was different in that two clones (a2 and b7/c15) belonging to V kappa 4 subgroup, but a4 used V kappa 1 light chain gene. CONCLUSION: Our approach should facilitate provision of useful information on the antibody responses against allergens at the molecular level in humans.
