Whole protein delivery to B-cells by cell squeezing enables robust MHC class I antigen presentation and antigen-specific CD8+T-cell priming in vitro.
A) Experimental timeline for antigen loading (endocytosis, peptide, or squeezed) on day 0 followed by co-culture with purified, CFSE-labelled OT-I CD8+T-cells. B) Representative histogram overlay showing data from day 4 proliferation of endocytosis+CpG or SQZ+CpG co-cultures; green gates were used to calculate percent of divided input cells and proliferation indices as described in Methods. C,D) Quantitative analysis of percent divided input OT-I CD8+(C) and OT-II CD4+(D) T-cells at day 4 of co-culture. E) Normalized total OT-I CD8+T-cell counts on day 4 of co-cultures were also calculated as described in Methods. All data were shown as means±standard deviation (n = 7 independent experiments for B & D; n = 3 independent experiments for D). Pairs of conditions were tested in C), D) and E) for statistically significant pairwise differences with ordinary 1-way ANOVA followed by Holm Sidak multiple comparisons test; multiplicity adjusted p-values < 0.05 were considered significant and exact p-values were shown where significant. All conditions in D) were significantly lower than positive controls (αCD3/28 beads, peptide).