Colorimetric Detection of Hemoglobin by Aptamer-Based Biosensor

Hemoglobin is an essential protein located in red blood cells. This protein's highly sensitive and selective detection plays a vital role in diagnosing several pathologies, including urinary, renal, and infectious diseases. Therefore, exploring simple and accurate methods for Hb detection is very important. In this work, we have developed an aptasensor based on a label-free colorimetric detection method for hemoglobin detection. The bovine serum albumin (BSA) adsorbed on the polystyrene microplate wells was used to immobilize an amino-modified aptamer specific for Hb. To avoid the labeling step, the peroxidase activity of Hb was explored for a direct generation of the colorimetric signal by catalyzing the peroxidase substrate TMB. The catalytic reaction of TMB occurs through the hemoglobin′s peroxidase activity, resulting in a blue-colored precipitate. Since the optimization steps are expensive, time-consuming, and require a large amount of reagents, response surface methodology based on Box-Behnken Design (RSM-BBD) and artificial neural network coupled with particle swarm optimization (ANN-PSO) were used as optimization methods to develop this colorimetric platform. The performance of RSM-BBD and ANN-PSO was compared, and the results were deeply analyzed. Colorimetric measurements have led to a linear range of 4-10 mg mL^-1and a detection limit of 0.044 mg mL^-1. In the proposed strategy, using the BSA molecule as an immobilization support and RSM-BBD and PSO-ANN as an optimization tool makes this label-free detection platform simpler and inexpensive.