Application of Recombinant Antibody Technology for the Development of Anti-Lipid antibodies for Tuberculosis Diagnosis

Abstract

Tuberculosis (TB) is the most significant infectious disease globally and is caused by the bacterium Mycobacterium tuberculosis. The burden of disease falls disproportionately on third world countries and the paucity of diagnostic resources and consequent misdiagnosis has resulted in significant morbidity and mortality despite the availability of effective treatment. There is thus an urgent need for a low cost point-of-care diagnostic. Antibody-based diagnostics are ideal but a suitable biomarker has yet to be found despite intensive research on protein targets. Lipid biomarkers however remain relatively unexplored due to the lack of suitable antibodies arising from their poor immunogenicity. However, the advent of recombinant antibody phage display allows for the selection of such antibodies in vitro without an immune response. We have therefore applied this technique to generate high affinity, highly specific antibodies against two TB lipid biomarkers: lipoarabinomannan, a soluble glycolipid found in urine; and mycolic acid, an insoluble fatty acid present at high levels in sputum. These antibodies were thoroughly characterized and testing against clinical samples indicated that our anti-lipoarabinomannan antibody had significant sensitivity in the smear-negative, HIV negative cohort. We also optimized a novel simplified method for expression of these antibodies in E. coli as current methods rely primarily upon expensive mammalian expression.