Lateral flow immunoassay for detection of active tuberculosis utilising CFP10-ESAT6 as biomarker

Tuberculosis (TB) is one of the greatest health care problems in the world. Traditional diagnostic techniques based on the isolation of the tuberculosis bacillus in culture media are time consuming, and it is necessary to wait for several weeks to obtain a result. Therefore, possible biosensor that easy to use and cheap is the lateral flow immunoassay (LFIA), which is affordable, sensitive, specific, and user-friendly. LFIA has been introduced as a handheld immunoassay-based point- of-care platform for an automated detection of TB. The CFP10-ESAT6 antigen of M. tuberculosis were used as the target in early detection of TB using LFIA strip- based point of care strategy Gold nanoparticles (AuNPs) were prepared in various shape, nanosphere, nanorod and nanostar. AuNPs in nanosphere shape were used as the colour probe for the detection of a target of interest. AuNPs were prepared through reduction of Aurum (III) Chloride with trisodium citrate. The prepared AuNPs were further conjugated with antibody (Rabbit anti Mycobacterium Tuberculosis). The high-resolution transmission electron microscopy (HRTEM) image and ultraviolet- visible spectrophotometer (UV-Vis) analysis confirmed that the synthesized AuNPs were appropriate for conjugation with the antibody for the immunoassay designed. As a proof of concept, conventional enzyme-linked immunosorbent assay (ELISA) method was carried out to validate our research finding. Sputum samples were spotted onto the LFIA strips and the result were obtained after 5-10 minutes. The positive and negative sample sputum were obtained from Hospital Universiti Sains Malaysia (HUSM) Kubang Kerian Malaysia which were confirmed by smear microscopy technique and culture technique. 12 μg/ml antibody was used for conjugation of antibody with gold nanoparticle in sphere shape. LFIA strips that were spot off with positive Tuberculosis sputum sample showed two red signals appeared on the membrane. One signal appeared on the test line and one in control line. For LFIA strips that were spot off with negative sputum sample, only one red signal appeared in membrane which is on the control line

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