Effect of antitetraspanin Mabs on the migration of immature DCs: association between CD63 and integrins. (A) Immature DCs were prepared as described in “Materials and methods,” and their migration in the presence of 100 ng/mL MIP-1α (□), MIP-5 (▪), and RPMI-0.1% BSA alone (▦) was performed as described in “Materials and methods.” An average of 5 experiments is shown. Error bars show standard deviation. Statistically significant differences between DCs incubated with the corresponding Mabs and DCs incubated with isotype-matched control immunoglobulins (IMCs) are indicated (^*P < .05, according to a 2-tailed Student t test). (B) Expression of CD29, CD11b, CD18, α₅, β₃, and MHC class II before and after CD63 internalization (shaded histograms). Isotype-matched control immunoglobulin labeling is shown in open histograms. The histograms shown are representative of at least 3 experiments. (C) Immunoprecipitation of CD63 and CD11b was performed as described in “Materials and methods.” The immunoprecipitates were analyzed by 10% SDS-PAGE under nonreducing conditions and revealed with anti-CD63 (broad band between 40 kDa and 70 kDa¹⁹ ) and anti-CD18 Mabs followed by GAM-PA and BCIP/NBT. Lanes 1 and 5: DC extract. Lanes 2 and 6: CD63 immunoprecipitates (IP 5.01). Lane 3: CD11b immunoprecipitate (IP CD11b). Lanes 4 and 7: control immunoprecipitations performed using isotype-matched control IgG2a (IP IgG2a).