4. 1 mL syringe with 27 G needle.
5. C. trachomatis elementary bodies, stored at À80
C in SPG
buffer.
2.3 Cell Sorting
1. RPMI medium (R0).
2. Amine-reactive viability dye [e.g., Zombie Fixable Viability
Dye from BioLegend].
3. Fluorescent antibodies against CD3, CD14, CD19, CD27,
CD38, IgG, IgM, IgD, and IgA.
4. 5 mL Polystyrene tubes for FACS.
Fig. 1 Schematic showing process of antibody isolation and vaccine development. Following vaccination with
a novel immunogen, PBMCs are isolated from blood taken from the vaccine recipients and are stained using
fluorescent elementary bodies (EBs) that are prepared concurrently with the PBMCs. This staining allows for
antigen-specific B cells to be sorted and the immunoglobulin genes cloned and produced recombinantly,
where they can be analyzed for affinity, specificity, and neutralization potential. This data can then be used to
design a new immunogen better able to push the humoral immune response toward epitopes that were shown
to be beneficial
Isolation of Chlamydia-Specific B Cells
21