CFSE⁺ cells enter the paracortex, co-localize with BCG and are closely apposed to P25 TCRTg cells.

<p>P25 TCRTg cells were isolated from LNs of naïve P25 TCRTg RAG–1^-/- ECFP mice using a CD4 negative selection procedure (Miltenyi Biotec), and transferred i.v. into WT recipients. Recipients were then inoculated with BCG-RFP and the CFSE-based migration assay performed. (<b>A</b>) Distribution of CFSE⁺ cells in the paracortex (T-cell area) of the pLN as determined by CD3 staining on pLN sections from BCG-injected mice. (<b>B</b>) Quantification of CFSE⁺ cells within the T-cell area of the pLN from BCG- and PBS-injected mice expressed as the number of CFSE⁺ cells per area of CD3 staining. (<b>C</b>) Micrograph showing distribution of BCG (red), CFSE⁺ cells (green), p25 TCRTg cells (blue) and HEVs (white) in the pLN. Boxed inserts in the micrograph were magnified to show x/y/z projections of two CFSE⁺ cells with intracellular BCG-RFP. BCG were overexposed in these magnified views to facilitate visualization. Infected cell in lower panel is also in contact with a p25TCRTg ECFP cell (blue). (<b>D</b>) Quantification of the number of CFSE⁺ cells containing BCG-RFP (left graph), found in close apposition with P25 TCRTg ECFP cells (center graph), and containing BCG-RFP and in close apposition with P25 TCRTg ECFP cells (right graph). Data samples represent average obtained from individual pLN from which multiple fields of maximum intensity projections were analyzed. Bars indicate standard error of the mean. White bar on micrographs depict 100 microns. *Denotes statistically significant difference between BCG-infected and PBS controls in (<b>B</b>) and between day 1 and 6 in (<b>D</b>).