Electrochemical aptasensor using optimized surface chemistry for the detection of Mycobacterium tuberculosis secreted protein MPT64 in human serum

Tuberculosis (TB) remains one of the leading causes of mortality worldwide. There is a great need for the development of diagnostic tests, which are reliable, sensitive, stable, and low cost to enable early diagnosis of TB in communities with scarce resources. This study reports the optimization and evaluation of a synthetic receptor, an aptamer, for the detection of the secreted protein MPT64, which is a highly immunogenic polypeptide of Mycobacterium tuberculosis, a causative agent of TB. The study investigates combinatorial effects of an aptamer linker and a co-adsorbent onto a gold electrode for optimal binding efficiency and reduced non-specific interactions for label-free detection of MPT64 using electrochemical impedance spectroscopy. Two types of co-adsorbents and two types of aptamer linkers were studied and high specificity and sensitivity to MPT64 was observed for a surface prepared with a thiol PEGylated aptamer HS-(CH₂)₆-OP(O)₂O-(CH₂CH₂O)₆-TTTTT-aptamer and 6-mercaptohexanol in a ratio of 1:100. The developed aptamer-based sensor was successfully used with spiked human serum sample with a limit of detection of 81 pM This work demonstrates the use of the MPT64 aptamer as a lower cost, more sustainable and stable alternative of antibodies for the development of point-of-care TB biosensors decreasing the detection time from several days or hours to thirty minutes.