Chitosan gold nanoparticle-based dot-blot assay for sensitive visual detection of histidine-tagged recombinant proteins

There are various techniques for detecting recombinant proteins such as western blot, SDS-PAGE, ELISA and fluorescence microscopy. However, these methods are difficult to handle, time-consuming and need special tools. We developed a rapid, inexpensive, sensitive, and straightforward approach to address these problems using an Antihistidine biosensor. Colloidal gold nanoparticles (GNPs) were synthesized by chitosan as reducer and stabilizer via the green synthesis method. Then, anti-His tag antibody was immobilized on the Chitosan-gold nanoparticles (CS-GNPs) surface to visually detect spotted target protein on nitrocellulose (NC) membrane. Our results showed that the designed dot-blot immunoassay can detect histidine-tagged recombinant proteins with the limit of detection (LOD) of 1µg/ml without any signal enhancement directly from cell lysate within 5 minutes. In an extra step, we applied HAuCl4 and NH2OH\ HCl as a gold enhancement system and increased the detection sensitivity to 0.1 µg/ml. The results showed that the developed assay can rapidly detect production of recombinant proteins, and it can be used as a screening method in low-resource laboratories.