Development of a screen-printed electrochemical immunosensor for the detection of prostate cancer biomarkers
Printed immunosensor; Conductive ink; Prostate cancer; Prostate-specific antigen; Human kallikrein 2
Prostate cancer (PCa) is the second most common type of cancer in the male population globally. The effectiveness of clinical treatment of PCa depends on the stage at which the disease is detected. Therefore, diagnosing the disease in early stages is critical to increase the patient's survival rate. Electrochemical sensors are devices used in analytical tests that aim to investigate the presence of species in a given sample and emerge as an alternative method to quantify PCa biomarkers. Printed electrodes are an excellent alternative in the area of sensor development, as they are easy to prepare, versatile, and, applicable in several areas, in addition to having high selectivity and sensitivity. Therefore, in this work, two screen-printed electrochemical immunosensors capable of detecting and quantifying the biomarkers PSA and KLK2 was developed. First, inks based on graphite, carbon black, nail polish, and acetone were produced for printing the sensor, and ink based on silver, nail polish, and acetone for the construction of the quasi-reference electrode. The sensors containing the working, quasi-reference, and auxiliary electrodes were printed using the screen printing technique on polyethylene terephthalate (PET) support. The printed electrode was characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), and the results showed the potential application of the electrochemical sensor. The immunosensor was constructed with the addition of gold nanoparticles (AuNPs), antibodies (anti-PSA or anti-KLK2), and bovine serum albumin (BSA) to the working electrode, for later application in the detection of the antigen of interest. Two immunosensors were developed, one for PSA detection and the other for KLK2. The parameters optimized for the best performance of the immunosensors were: antibody incubation time, BSA and antigen, BSA concentration, pH, and support electrolyte concentration. By monitoring the electrochemical behavior of the potassium ferrocyanide probe in front of the immunosensor, it was possible to observe a reduction in the signal of the anodic current, which is due to the immobilization of the proteins that act as an insulator. The immunosensor proposed for PSA detection had an LOD equal to 0.55 ng mL-1 and a LOQ of 1.82 ng mL-1. The interferents did not significantly affect the current signal and the reproducibility showed a relative standard deviation of 5.9%. The immunosensor was applied to synthetic human serum samples by the addition and retrieval method, and the results obtained demonstrated accuracy and potential application in clinical analyses. The immunosensor proposed for the detection of KLK2 showed LOD 0.17 ng mL-1 and LOQ 0.55 ng mL-1. These results suggest a potential application for analysis in biological samples. In addition, the methodology presented enables the development of an immunosensor that is easy to prepare and inexpensive, with an approximate cost of R$ 1.71 per unit.