Detection of alpha-fetoprotein through biological signal amplification by biosensor based on imaging ellipsometry

Abstract

High sensitive and quantitative detection of alpha-fetoprotein (AFP) by biosensor based on imaging ellipsometry (BIE) through biological amplification was investigated. AFP firstly reacted with the rat monoclonal antibody (rat-mAb) initially immobilized on glutaraldehyde modified silicon surface, then rabbit anti-human AFP polyclonal antibodies (Rabbit-pAb) and goat anti-rabbit IgG (goat-IgG) were sequentially applied to amplify signal. Results revealed that signal was enhanced approximately six fold. The linear range of AFP detection was 20.0–200.0ng/ml with a low limit of 5.0ng/ml (S/N=3). The cross-reaction rate was less than 5.2% evaluated by biomarker (carcinoembryonic antigen, carbohydrate 19-9 and carbohydrate antigen 242) and two common proteins (human serum albumin, fibrinogen) and their mixture. Coefficient variation (CV) for intra-slide and inter-slide reproducibility were 10.3%, 6.6%, 6.3% and 10.7%, 7.9%, 6.4% for 41.6ng/ml, 83.2ng/ml and 128.4ng/ml AFP. In clinic application, cut-off value at 28.6ng/ml was also determined with sensitivity 0.72 and specificity 0.94. Results of 47 clinic patient samples detected by BIE were in good agreement with those of electrochemiluminescence immunoassay (ECLIA) (R2=0.9949).The area under receiver operating characteristic curve (ROC) curve is 0.89. Statistical analysis showed that BIE is high agreement with ECLIA (Kappa=0.733, U=3.21>U0.01). It shows a potential for hepatocellular carcinoma (HCC) diagnosis.