Reagentless amperometric immunosensors based on direct electrochemistry of horseradish peroxidase for determination of carcinoma antigen-125.
A novel strategy for immunoassay and the preparation of reagentless immunosensors was proposed. This strategy was based on the immobilization of antigen and the direct electrochemistry of horseradish peroxidase (HRP) that was labeled to an antibody. A reagentless immunosensor for carcinoma antigen-125 (CA 125) determination was developed. The immunosensor was prepared by immobilizing CA 125 with titania sol-gel on a glassy carbon electrode by the vapor deposition method. The incubation of the immunosensor in phosphate buffer solution (PBS) including HRP-labeled CA 125 antibody led to the formation of a HRP-modified surface. The immobilized HRP displayed its direct electrochemistry with a rate constant of 3.04 +/- 1.21 s(-1). With a competition mechanism, a differential pulse voltammetric determination method for CA 125 was established by the peak current decrease of the immobilized HRP. The current decrease resulted from the competitive binding of the CA 125 in sample solution and the immobilized CA 125 to the limited amount of HRP-labeled CA 125 antibody. Under optimal conditions, the current decrease was proportional to CA 125 concentration ranging from 2 to 14 units mL(-1) with a detection limit of 1.29 units mL(-1) at a current decrease by 10%. The CA 125 immunosensor showed good accuracy and acceptable precision and fabrication reproducibility with intraassay CVs of 8.7 and 5.5% at 8 and 14 units mL(-1) CA 125 concentrations, respectively, and interassay CV of 19.8% at 8 units mL(-1). The storage stability was acceptable in a pH 7.0 PBS at 4 degrees C for 15 days. The proposed method provided a new promising platform for clinical immunoassay.
Dai
Zong
Z
Department of Chemistry, Institute of Analytical Science, State Key Laboratory of Coordination Chemistry, Nanjing University, Nanjing 210093, P. R. China.
Yan
Feng
F
Chen
Jin
J
Ju
Huangxian
H
D000911:Antibodies, Monoclonal / Q000276:immunology; D001667:Binding, Competitive / Q000276:immunology; D015374:Biosensing Techniques / Q000379:methods*; D018394:CA-125 Antigen / Q000032:analysis* / Q000097:blood / Q000276:immunology; D004563:Electrochemistry / Q000379:methods; D006735:Horseradish Peroxidase / Q000737:chemistry*; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D009942:Organometallic Compounds; D010084:Oxidation-Reduction; D015203:Reproducibility of Results; D013696:Temperature; D014025:Titanium / Q000737:chemistry
D000911:Antibodies, Monoclonal; D018394:CA-125 Antigen; D009942:Organometallic Compounds; C102815:titanium isopropoxide; D014025:Titanium; D006735:Horseradish Peroxidase