Conjugate Vaccines from Bacterial Antigens by Squaric Acid Chemistry: A Closer Look

Abstract By using O‐SP‐core (O‐SPcNH 2 ) polysaccharide, isolated from Vibrio cholera O1 lipopolysaccharide (LPS) and related synthetic substances, a detailed study of factors that affect conjugation of bacterial polysaccharides to protein carriers through squaric acid chemistry to form conjugate vaccines has been carried out. Several previously unrecognized processes that take place during the squarate labeling of the O‐SPcNH 2 and subsequent conjugation of the formed squarate (O‐SPcNH‐SqOMe) have been identified. The efficiency of conjugation at pH 8.5, 9.0, and 9.5 to bovine serum albumin (BSA) and to the recombinant tetanus toxin fragment C (rTT‐Hc) has been determined. The study led to a protocol for more efficient labeling of O‐SPcNH 2 antigen with the methyl squarate group, to yield a…