Coralite594 conjugated goat anti rabbit igg

PC9 cells were seeded on glass coverslips and incubated for 48 h. After being fixed with 4% paraformaldehyde, cells were treated with 0.5% Triton X-100, then sealed with 5% goat serum at room temperature for 1 h. The primary antibody was incubated at 4 °C overnight. The anti-HOXC11 mouse antibody was purchased from Novus (1:100, NBP2-00499). The anti-IKKα rabbit antibody and Myc-Tag mouse antibody were purchased from CST (1:3000, 61294; 1:8000, 2276). The fluorescent secondary antibody was diluted with 5% BSA at the ratio of 1:200 and incubated at room temperature for 1 h under dark conditions. The CoraLite 594-conjugated Goat Anti-Rabbit IgG and CoraLite 488-conjugated Affinipure Goat Anti-Mouse IgG were purchased from Proteintech (SA00013-4; SA00013-1). Hoechst stained the nuclei. The photos were caught by laser confocal scanning microscopy (LSM700, ZEISS, Jena, Germany).

The suspended cells were fixed with 4% paraformaldehyde for 30 min and washed by centrifugation in PBS. Then cells were resuspended in PBS containing 0.3% Triton X-100 and 5% BSA, and blocked for 1 h. After centrifugation and washing, the cell pellet was resuspended in 3% BSA solution with primary antibodies at room temperature for 1 h. After that, cells were washed twice with PBS. Then the cell pellet was resuspended in 3% BSA solution with the secondary antibodies at room temperature for 1 h. After centrifugation and washing, the resuspended cells were stained with a 1:1 mixture of DAPI solution and 3% BSA solution for 10 min. After washing with the PBS, the sample was standby for observation. The primary antibodies used in this study were CD45 antibody (Cell Signaling Technology, Danvers, MA, USA), EpCAM antibody (Cell Signaling Technology), and cytokeratin 19 antibody (Abcam, Cambridge Science Park, UK). The secondary antibodies were coraLite488 conjugated goat anti-mouse IgG (Proteintech, Chicago, IL, USA), and coraLite594 conjugated goat anti-rabbit IgG (Proteintech). Fluorescence images were taken using a fluorescence microscopy (Olympus, Tokyo, Japan) with a 40× objective lens.

Anti-Flag, anti-HA, anti-Myc, anti-GST tag antibodies, anti-Phospho-IRF3 (S396) antibody, anti-GAPDH and anti-actin antibodies, CoraLite594-conjugated goat anti‐rabbit IgG, and CoraLite488-conjugated Goat Anti-Rabbit IgG antibodies were purchased from Proteintech, Wuhan, China; anti-IRF3 antibody was obtained from the Cell Signaling, Danvers, USA. MG132 was purchased from Sigma, St Louis, USA. Z-VAD-FMK was obtained from Promega, Madison, USA. Chloroquine was purchased from MCE, Monmouth, USA.