Fig. 4: Migration of moDC toward CCL21 and CXCL12 in the presence of therapeutic protein correlates with its immunogenic potential.

a Schematic of the transwell migration assay capturing the potential for DC migration toward therapeutic protein in the SC space. A concentration gradient of therapeutic protein and chemokines was created across the transwell insert. Immature DCs were plated in the upper chamber and allowed to migrate into the lower chamber. Migrated DCs were counted in the lower chamber by flow cytometry. Schematic created at Biorender.com. b The migration index of moDC along a concentration gradient of therapeutic protein in the presence of CCL21 and CXCL12. Cells were plated in the upper chambers of transwell inserts with 10 μg/mL therapeutic protein. 50 μg/mL therapeutic protein plus 100 ng/mL each CCL21 and CXCL12 were added to lower chambers. Migration index = (% migrated of protein treatment group)/(average % migrated of “media” control), where “media” has only chemokines in the lower chamber. c (Left y-axis) The fold-change in CXCR4⁺ (%) moDC over untreated as a function of KLH concentration for a representative donor: (◆ closed diamond) 236. (Right y-axis) The migration index of moDC as a function of KLH concentration in the lower chamber for representative donors: (□ open square) 919, (△ open triangle) 014. Each dot is a technical replicate (n = 3 wells); error bars are mean ± SD. d, e Cells were plated in the upper chambers of transwell inserts with 100 μg/mL therapeutic protein. 1000 μg/mL therapeutic protein plus 100 ng/mL each CCL21 and CXCL12 were added to lower chambers. In b, d, e treatments were tested in triplicate (n = 3 wells) for each donor, and each dot represents the mean for one donor: (● closed circle) 274, (■ closed square) 926, (□ open square) 919, (△ open triangle) 014, (∇ upside-down triangle) 890, (◆ closed diamond) 236. Error bars are mean ± SEM.