Figure 5.

Adoptive cell transfer experiment with and without AMD3100 treatment of the transferred cells. (A) Bursae of the embryos were tested for BCR^pos, GFP^neg cells after wt cell transfer into BCR^neg embryos on ED19. The detection was performed by staining with the anti-chicken IgM antibody. In the bursae of BCR^neg embryos in which wt splenocytes were injected (A, top panel), up to 17% BCR^pos/GFP^neg cells were detected (A, top panel, population circled in red). In the recipients of wt cells after AMD3100 treatment this population was limited to 1% (A, bottom panel, population circled in red). (B) After detection of wt IgM^pos cells in the bursae of the recipient BCR^neg embryos per flow cytometry and absolute counting, the amount of the transferred cells detected in the bursa were compared. In the bursae of the embryos, in which AMD3100 treated cells (n = 3) were transferred were significantly less IgM^pos cells detected than in the bursae of the embryos in which splenocytes were transferred without AMD3100 treatment (n = 6) (normally distributed per Shapiro-Wilk and Kolmogorov-Smirnov tests, independent t-test, *p < 0.05). (C) Fluorescence histology staining of the nuclei with DAPI (in blue) and the B cell receptor with the antiIgM antibody (in red) of a wt embryonic bursa (a), of a BCR^neg embryonic bursa (b) and of a BCR^neg embryonic bursa after adoptive cell transfer of wt splenocytes (c). The presence of BCR^pos B cells is clearly demonstrated in the BCR^neg embryonic bursa. B cells are organized in bursal follicles as seen in the wt bursa. Representative results out of at least three examined animals per group are shown.