title: Heterologous antigen selection of camelid heavy chain single domain antibodies against tetrabromobisphenol A.
abstract: Tetrabromobisphenol A (TBBPA) is a ubiquitous flame retardant. A high-throughput immunoassay would allow for monitoring of human and environmental exposures as a part of risk assessment. Naturally occurring antibodies in camelids that are devoid of light chain, show great promise as an efficient tool in monitoring environmental contaminants, but they have been rarely used for small molecules. An alpaca was immunized with a TBBPA hapten coupled to thyroglobulin and a variable domain of heavy chain antibody (VHH) T3-15 highly selective for TBBPA was isolated from a phage displayed VHH library using heterologous coating antigens. Compared to the VHHs isolated using homologous antigens, VHH T3-15 had about a 10-fold improvement in sensitivity in an immunoassay. This assay, under the optimized conditions of 10% methanol in the assay buffer (pH 7.4), had an IC50 for TBBPA of 0.40 ng mL(-1) and negligible cross reactivity (<0.1%) with other tested analogues. After heating the VHH at 90 °C for 90 min about 20% of the affinity for coating antigen T3-BSA remained. The recoveries of TBBPA from spiked soil and fetal bovine serum samples ranged from 90.3% to 110.7% by ELISA and agreed well with a liquid chromatography-tandem mass spectrometry method. We conclude the many advantages of VHH make them attractive for the development of immunoassays to small molecules.
authors: College of Resources and Environmental Sciences, Beijing Key Laboratory of Biodiversity and Organic Farming, China Agricultural University , Beijing 100193, P.R. China.
authors: Jia
authors: J
authors: Wang
authors: Candace R S
authors: CR
authors: Bever
authors: Zuzana
authors: Z
authors: Majkova
authors: Julie E
authors: JE
authors: Dechant
authors: Jun
authors: J
authors: Yang
authors: Shirley J
authors: SJ
authors: Gee
authors: Ting
authors: T
authors: Xu
authors: Bruce D
authors: BD
authors: Hammock
mesh_terms: D000818:Animals; D015478:Antigens, Heterophile / Q000276:immunology; D002161:Camelids, New World / Q000276:immunology; D002417:Cattle; D004784:Environmental Monitoring / Q000379:methods; D005411:Flame Retardants / Q000032:analysis*; D007118:Immunoassay / Q000379:methods*; D007143:Immunoglobulin Heavy Chains / Q000276:immunology*; D057230:Limit of Detection; D011075:Polybrominated Biphenyls / Q000032:analysis* / Q000097:blood / Q000276:immunology; D061905:Single-Domain Antibodies / Q000276:immunology*; D012987:Soil / Q000737:chemistry; D012989:Soil Pollutants / Q000032:analysis*
chemical_list: D015478:Antigens, Heterophile; D005411:Flame Retardants; D007143:Immunoglobulin Heavy Chains; D011075:Polybrominated Biphenyls; D061905:Single-Domain Antibodies; D012987:Soil; D012989:Soil Pollutants; C020806:tetrabromobisphenol A