A Novel Amperometric Aptamer–Antibody Sandwich Assay for the Detection of Tuberculosis With Diazonium Electrografted Enhanced Modified Electrode

In this study, a sensitive aptamer-antibody-based sandwich biosensor utilizing chronoamperometry for early detection of
Mycobacterium tuberculosis
CFP10 antigen is reported. An anti-CFP10 aminated aptamer was covalently immobilized onto the disposable screen-printed carbon electrode via carbodiimide chemistry with the electrografted 4-carboxyphenyl diazonium salt. The CFP10 aptamer recognized and captured the target protein through affinity binding. The quantification of CFP10 antigen was based on amperometric responses from the electrocatalytic reaction between horseradish peroxidase (HRP) and H
₂
O
₂
, with hydroquinone as an electron transfer mediator. The developed aptasensor displayed an excellent analytical performance with a linear range of 5 to 500 ng mL
⁻¹
and a detection limit of 1.22 ng mL
⁻¹
in buffer solution. The diazonium-based sensor was also successfully applied on spiked human serum sample and exhibited a detection limit of 1.05 ng mL
⁻¹
. Furthermore, the fabricated aptasensor demonstrated high sensitivity to CFP10 antigen over other proteins. The proposed aptamer-based sensor is rapid, sensitive, and disposable, thus suitable for point-of-care tuberculosis detection. This work also highlights the use of CFP10 aptamer as a less expensive and stable alternative reagent to antibodies for the development of a better diagnosis of tuberculosis.