Development of a noncompetitive phage anti-immunocomplex assay for brominated diphenyl ether 47.
We present a new application of the noncompetitive phage anti-immunocomplex assay (PHAIA) by converting an existing competitive assay to a versatile noncompetitive sandwich-type format using immunocomplex binding phage-borne peptides to detect the brominated flame retardant, brominated diphenyl ether 47 (BDE 47). Three phage-displayed 9-mer disulfide-constrained peptides that recognize the BDE 47-polyclonal antibody immunocomplex were isolated. The resulting PHAIAs showed variable sensitivities, and the most sensitive peptide had a dose-response curve with an SC(50) (concentration of analyte producing 50% saturation of the signal) of 0.7ng/ml BDE 47 and a linear range of 0.3-2ng/ml, which was nearly identical to the best heterologous competitive format (IC(50) of 1.8ng/ml, linear range of 0.4-8.5/ml). However, the PHAIA was 1400-fold better than homologous competitive assay. The validation of the PHAIA with extracts of house furniture foam as well as human and calf sera spiked with BDE 47 showed overall recovery of 80-113%. The PHAIA was adapted to a dipstick format (limit of detection of 3.0ng/ml), and a blind test with six random extracts of local house furniture foams showed that the results of the PHAIA and dipstick assay were consistent, giving the same positive and negative detection.
Department of Entomology and UCD Cancer Center, University of California, Davis, CA 95616, USA.
Hee-Joo
HJ
Kim
Martin A
MA
Rossotti
Ki Chang
KC
Ahn
Gualberto G
GG
González-Sapienza
Shirley J
SJ
Gee
Ruthie
R
Musker
Bruce D
BD
Hammock
D000818:Animals; D000906:Antibodies / Q000276:immunology; D002417:Cattle; D003429:Cross Reactions; D055768:Halogenated Diphenyl Ethers / Q000032:analysis / Q000097:blood / Q000737:chemistry*; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D019151:Peptide Library; D012997:Solvents / Q000737:chemistry
D000906:Antibodies; D055768:Halogenated Diphenyl Ethers; D019151:Peptide Library; D012997:Solvents