Fabrication of a Novel Conductometric Biosensor for Detecting Mycobacterium avium subsp. paratuberculosis Antibodies

Abstract

Johne’s disease (JD) is one of the most costly bacterial diseases in cattle. In the U.S., economic losses from the disease have been estimated to exceed $1,500,000,000 per year, mainly from the effects of reduced milk production. Current diagnostic tests for JD are laboratory based and many of those tests require specialized equipment and training. Development of rapid and inexpensive diagnostic assays, which are adapted for point-ofcare applications, would aid in the control of JD. In this study, a polyaniline (Pani)-based conductometric biosensor, in an immunomigration format, was fabricated for the detection of serum antibody (IgG) against the causal organism of JD, Mycobacterium avium subsp. paratuberculosis (MAP). Immobilized Mycobacterium avium purified proteins in the capture membrane were used to detect MAP IgG, previously bound with Pani/anti-bovine IgG* conjugate in the conjugate membrane. After detection, the Pani in the sandwiched captured complex bridges an electrical circuit between the silver electrodes, flanking the capture membrane. The electrical conductance, caused by Pani, was measured as drop in electrical resistance. Testing of the biosensor with known JD positive and negative serum samples demonstrated a significant difference in the mean resistance observed between the groups. This proof-of-concept study demonstrated that a conductometric biosensor could detect MAP IgG in 2 minutes. The biosensor’s speed of detection and the equipment involved would, among other things, support its application towards the various point-ofcare opportunities aimed at JD management and control.

Keywords

• Conductometric
• biosensor
• immunoassay
• electrochemical immunosensor
• polyaniline
• paratuberculosis
• Johne’s disease
• on-site diagnosis