Abstract: The aim of this study was to establish a simple and reliable assay for fluorescence labeling Muscovy duck lymphoeytes and then analyze the effects of Muscovy duck reovirus (MDRV) infection on lymphocytes homing of ileum in Muscovy ducklings. 5(6)-carboxydiacetate fluorescein succinimidyl ester (CFDA-SE, CFSE), a living cell fluorescent reagent, was used to label the peripheral blood lymphocytes of Muscovy ducklings. Flow cytometry was used to detect and analyze the lymphocytes that marked in vitro and traced in vivo. In addition, CFSE⁺ lymphocytes of ileum in MDRV-infected and uninfected Muscovy duck were detected and analyzed by flow cytometry and paraffin section immunofluorescence. Results were as follows:The optimal conditions of labeling lymphocytes by CFSE were PBS as the incubation solution, 10 μmol·L^-1 CFSE, 37℃ and 30 min, respectively. The labeled lymphocytes were injected into the Muscovy duck through the brachial vein, the CFSE⁺ lymphocytes in the peripheral blood were basically stable at 2% to 5%. The order of sequence of peak of CFSE⁺ lymphocytes was spleen, jejunum, ileum, cecum, duodenum, bursa and thymus. CFSE-labeled lymphocytes were injected into brachial veins of the young Muscovy ducks before MDRV infection, and the rate of CFSE⁺ lymphocytes in MDRV group was significantly (P<0.01) higher than that in the MOCK group at 1-10 days post infection. This result was consistent with tests of α4⁺ lymphocytes and paraffin section fluorescence. Overall, these results showed that the CFSE-labeling lymphocytes can be used for lymphocytes trace in vivo, and preliminary application results revealed that MDRV infection promotes the lymphocyte homing of ileum in Muscovy ducklings, which provide a basis for further studying the intestine pathogenic mechanism of MDRV infection.

Key words: CFSE, Muscovy ducklings, lymphocyte, flow cytometry, homing