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. Author manuscript; available in PMC: 2016 Jul 1.
Published in final edited form as: J Immunol Methods. 2015 Apr 9;422:111–117. doi: 10.1016/j.jim.2015.04.001

Figure 2. Effects on 2D6 and ZAC-3 on V. cholerae agglutination and motility.

Figure 2

(A) Bacterial macro-agglutination assays: mid-log phase V. cholerae O395 was mixed with an equal volume of LB medium containing two-fold serial dilutions of Sal4 (control), 2D6 or ZAC-3 IgA or IgG mAbs and then seeded in U-bottom 96-well plates. Microtiter wells were monitored for the appearance of aggregated bacteria over a period of 2 hr. 2D6 IgA promoted the appearance of visible clumps at concentrations ≥18 μg/ml, whereas ZAC-3 IgA promoted agglutination at ≥0.25 μg/ml (arrowheads). Neither 2D6 nor ZAC-3 IgG produced visible agglutination at any concentration tested. (B and C) Semi-solid liquid motility assays: single colonies of V. cholerae O395 were stabbed into 0.3% semi-solid LB agar containing 9 μg/ml of ZAC-3 IgG1 or ZAC-3 Fab fragments. Plates were incubated at 37°C and the diameter of the bacterial spread was measured every 60 min. These data demonstrate that both ZAC-3 IgG and Fab fragments significantly reduced V. cholerae motility (P=0.0003, Student t test), although the Fab fragments effects were only apparent at the earliest time point.