Electrochemical immunosensor for determination of LeptospiraL LIPL32 protein

Abstract

LipL32 is a surface membrane protein present only on pathogenic Leptospira species, which are the cause of Leptospirosis disease. As this zoonotic disease is of global importance but current diagnostic tools are still ineffective and/or inaccessible to rural areas which are affected, the need to develop an immunosensor for its detection is clear. In this work, an immunosensor for LipL32 protein was developed. Prussian blue was mixed with Graphene-PEDOT:PSS, creating Graphene-Prussian blue composite (GPB). The GPB was drop-casted on screen-printed carbon electrode (SPCE) which was later coated by Chitosan (CS) and citrate-capped gold nanoparticles (cAuNPs). The modified electrode was then further incubated with Staphylococcal protein A (PrA) and anti-LipL32 antibodies (Ab) for more sensitivity in LipL32 detection. Bovine Serum Albumin (BSA) then was used to coat the topmost layer to prevent non-specific binding of the electrode to unrelated biomolecules. The optimum electrode obtained could be used to detect LipL32 in a linear range of 50 to 200 ng ml-1 and 250 to 600 ng ml-1, lower detection limit of 54.98 ng ml-1 with RSD of reproducibility at 8.254. Furthermore, tests with intact and sonicated Leptospira interrogans cells resulted in linear range of 10 to 100 cells ml-1, lower detection limit of 44.29 cells ml-1 and 77.25 cells ml-1. The stability of the fabricated electrode also satisfactory, with only 6.79% drop in residual response current after three weeks of storage at 4°C in dried form.