Photochemical Treatment of Drosophila APCs Can Eliminate Associated Viruses and Maintain the APC Function for Generating Antigen-Specific CTLs Ex Vivo

Figure 3

Effect of UVADEX on Drosophila cell growth and surface marker expression. (a) Drosophila cell line 668 was treated with UVADEX (5 μg/ml) at 4°C for 30 min following UV treatment for 0 min, 2 min, 10 min, and 20 min, respectively. The treated cells were washed completely to remove residual UVADEX and seeded in a 6-well plate at 1 × 10⁶/ml and continually grown for 16 days. Cell survival was monitored by trypan-blue staining. (b) UVADEX treatment completely inactivates Drosophila cells. Drosophila cell 668 was treated with UVADEX (5 μg/ml) at 4°C for 30 min or γ-irradiation for 45 min. The UVDEX-treated cells and γ-irradiated cells were continually grown for 16 days, respectively. The cells from two different treatments were collected and stained with FITC-conjugated mAb against HLA-ABC, CD80, and CD54 isotype control antibody and PI at 4°C for 30 min, respectively. Flow cytometry analysis was used to analyze the live cells to determine the expression of HLA-ABC, CD80, and CD54. (c) RNA was isolated from each group of cells above, and RT/PCR was performed using Platinum Taq Polymerase and specific primers for human beta-2-microglobulin, human LFA-3, human CD80, human CD86, and human A2.1. PCR products in each group run on 1% agar. Lane 1, MW marker; lane 2, DNA from Drosophila 668 cell; lane 3, DNA from γ-irradiated 668 cells; lane 4, DNA from UVADEX-treated 668 cells for 2 min; lane 5, DNA from UVADEX-treated 668 cells for 10 min; and lane 6, DNA from UVADEX-treated 668 cells for 20 min.

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