Article

Enhanced conductometric immunoassay for hepatitis B surface antigen using double-codified nanogold particles as labels

Department of Hepatobiliary Surgery, Research Institute of Surgery, Daping Hospital, Third Military Medical University, Chongqing 400042, PR China; Department of Dermatology, Research Institute of Surgery, Daping Hospital, Third Military Medical University, Chongqing 400042, PR China; Department of Cancer Centre, Research Institute of Surgery, Daping Hospital, Third Military Medical University, Chongqing 400042, PR China
Biochemical Engineering Journal DOI:10.1016/j.bej.2009.03.002

ABSTRACT A new conductometric immunoassay for hepatitis B surface antigen (HBsAg) was developed based bioelectrocatalytic reaction on a microcomb-type electrode by using double-codified nanogold particles as labels. This microcomb-type electrode was fabricated on an interdigitated transducer covered with a well-ordered anti-HBs/protein A/nanogold architecture. The double-codified nanogold particles were prepared by using nanogold-labeled anti-HBs antibodies conjugated with horseradish peroxidase (HRP). Sandwich-type immunoassay protocol was successfully introduced for the detection of HBsAg. The formation of the immunocomplex changed the direct electrical communication between the carried HRP and the electrode, and thus local conductivity variations could be assayed based on the bioelectrocatalytic reaction of the carried HRP in 0.01 M PBS (pH 7.0) containing 60 μM H2O2, 0.08 M KI and 0.1 M NaCl. Under optimized conditions, the linear range obtained by using HRP-conjugated anti-HBs as secondary antibodies was 1.5–450 ng/mL HBsAg, while the assay sensitivity by using double-codified nanogold particles could be further increased to 0.01 ng/mL with the linear range from 0.1 to 600 ng/mL HBsAg. The developed immunoassay method showed good precision, high sensitivity, acceptable stability and reproducibility, and could be used for the detection of real sample with consistent results in comparison with those obtained by the ELISA method.

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Keywords

acceptable stability
 
assay sensitivity
 
bioelectrocatalytic reaction
 
carried HRP
 
developed immunoassay method
 
double-codified nanogold particles
 
ELISA method
 
HRP-conjugated anti-HBs
 
interdigitated transducer
 
local conductivity variations
 
M KI
 
microcomb-type electrode
 
nanogold-labeled anti-HBs antibodies conjugated
 
new conductometric immunoassay
 
optimized conditions
 
real sample
 
reproducibility
 
Sandwich-type immunoassay protocol
 
secondary antibodies
 
well-ordered anti-HBs/protein A/nanogold architecture
 

Hongming Liu