Microfluidic electrochemical immunosensor for the determination of cystatin C in human serum

The fabrication of a nanointerfaced electrochemical immunosensor is described for the rapid determination of cystatin C, a biomarker that is elevated in diabetic retinopathy. A dispersion of graphene oxide-chitosan (GO-Chit) nanocomposite was used to modify the carbon working electrode, allowing for a high conjugation of anti-cystatin C antibody. This modified sensor was characterized both morphologically and electrochemically, and the sensor performance was evaluated towards selective quantification of cystatin C in simulated as well as serum samples using cyclic voltammetry and differential pulse voltammetry. The sensor was able to detect cystatin C in the concentration range1 - 10 mg/L with a detection limit of 0.0078 mg/L. The preparation time of the sensor was 420 s, which was faster than that of conventional ELISA and other electrochemical sensors reported in literature. The clinical applicability of the proposed electrochemical biosensor was demonstrated through quantification of cystatin C in human serum samples and identification of diabetic retinopathy. A cutoff value of 1.2 mg/L of cystatin C was used beyond which the samples were classified as positive for diabetic retinopathy. Two different working electrodes, namely a glassy carbon electrode (GCE) and paper electrodes, were used in the study. The working potential was set to 0.25 V vs. Ag/AgCl for experiments with the GCE and 0.15 V for the paper electrodes. The prediction was validated by clinical diagnosis wherein the prediction accuracy of the sensor exceeded 85%. The sensor platform was translated onto a paper substrate and characterized for achieving an optimum sensing performance. This work is the first attempt to employ an electrochemical cystatin C sensor for the diagnosis of diabetic retinopathy from serum samples. Graphical abstract.