Figure 3.

Targeted mutation of the integrin‐binding region of P66 significantly reduces binding to cells expressing α_vβ₃ integrin.

A. Up to 0.1 μM recombinant protein was incubated with 293 + α_vβ₃ cells grown to confluence in 96‐well plates for 1 h before washing and fixation of bound protein. Bound protein was quantified by ELISA using anti‐MBP as primary antibody, and anti‐rabbit‐HRP conjugate as secondary. Developed plates were read at 655 nm. ELISA signal was normalized to crystal violet staining of the wells and results were normalized to MBP‐P66M.

B. 1.25 × 10⁶ HB19‐1 (wild‐type) and derivative bacteria per well were incubated with cells for 1 h before completing the assay as above, substituting anti‐Lyme spirochete as primary antibody. Results were normalized to wild type. Mean and standard error of measurement are indicated. Statistical analyses were performed using one‐way ANOVA Kruskal–Wallis test with Dunn's multiple comparison post‐test. *P < 0.05, **P < 0.01, ***P < 0.001.