The antigenicity and cholesteroid nature of mycolic acids determined by recombinant chicken antibodies

Abstract

Mycolic acids (MA) are major, species-specific lipid components of Mycobacteria and related genera. In Mycobacterium tuberculosis, it is made up of alpha-, methoxy- and keto-MA, each with specific biological functions and conformational characteristics. Antibodies in tuberculosis (TB) patient sera respond differently towards the three MA classes and were reported to cross-react with cholesterol. To understand the antigenicity and cholesterol cross-reactivity of MA, we generated three different chicken -derived phage-displayed single-chain variable fragments (scFv) that reacted similarly towards the natural mixture of MA, but the first recognized all three classes of chemically synthetic MAs, the second only the two oxygenated types of MAs and the third only methoxy MA. The cholesterol cross-reactivity was investigated after grafting each of the three scFv types onto two configurations of constant chain domains–CH1-4 and CH2-4. Weak but significant cross-reactivity with cholesterol was found only with CH2-4 versions, notably those two that were also able to recognize the trans-keto MA. The cholesteroid nature of mycobacterial mycolic acids therefore seems to be determined by the trans-keto MA subclass. The significantly weaker binding to cholesterol in comparison to MA confirms the potential TB diagnostic application of these antibodies.

Synthesis and characterisation of quantum dots coupled to mycolic acids as a water‐soluble fluorescent probe for potential lateral flow detection of antibodies and diagnosis of tuberculosis

“…Attached at the end of the strip is an absorbent pad used as a reservoir for waste and preventing the backflow of the liquid. [23,24] To develop such a test based on the MA-anti-MA (antigen-antibody) interaction one could make use of anti-MA antibodies that have been developed, [25] but it would be necessary to add a fluorescent tag to the MAs in order to visualise the antibody-antigen-antibody aggregate and it would be necessary to solubilize the hydrophobic MAs in order for it to interact with the aqueous serum.…”