Fig. 5

Co-immunoprecipitation and immunocolocalization to analyze the interaction between TvAP33 and BNIP3. a Western blot analysis of the expression of TvAP33 and BNIP3 in 293 T cells. Lanes: M Protein molecular weight marker (ordinate values in kDa), 1 after transfection of 293 T cells with pCMV-3HA-BNIP3, the primary antibody of anti-BNIP3 was used to recognize the expression of BNIP3 in cells, 2 after transfection of 293 T cells with pDsRed-N1-TvAP33, the primary antibody of anti-TvAP33 was used to recognize the expression of TvAP33 in cells. b Co-immunoprecipitation to verify the interaction between TvAP33 and BNIP3. After cotransfection of 293 T cells with pDsRed-N1-TvAP33 and pCMV-3HA-BNIP3, the immune precipitates were obtained from cellular lysis by the AminoLink Plus Coupling Resin immobilized with affinity-purified anti-Flag antibody or mouse IgG as control. The primary antibodies of anti-Flag and anti-HA were used to confirm TvAP33 and BNIP3 in precipitates and cellular lysis, respectively. c Immunocolocalization to verify the interaction between TvAP33 and BNIP3 in VK2/E6E7 cells. After transfection of VK2/E6E7 cells with pDsRed-N1-TvAP33, the primary antibodies of anti-Flag and anti-BNIP3 were used to confirm TvAP33 and BNIP3 in VK2/E6E7 cells, respectively. The Cy3 (red) and FITC (green) was used to label TvAP33 and BNIP3, and the DAPI (blue) was used to stain the nuclei. Blank VK2/E6E7 cells were used as controls