Alexa flour 488 anti rabbit igg antibodies

Immunofluorescence staining was conducted as previously described [32 ]. Cells were incubated with anti-PHB2 mouse antibodies (1:50, Proteintech) and anti-NDUFS1 rabbit antibodies (1:50, Proteintech) overnight at 4 °C. Then, Cells were incubated at room temperature with Alexa Flour 488 anti-rabbit IgG antibodies (1:100, Proteintech) and Alexa Flour 405 anti-mouse IgG antibodies (1:100, Proteintech) for 1 h protected from light. The co-localization of PHB2 and NDUFS1 was detected by a confocal laser scanning microscope (Carl Zeiss, Oberkochene, Germany).

HGC-27 cells were fixed with methanol at 4 ℃ for 30 min followed by incubation with anti-PHB2 mouse antibodies (1:50, Proteintech, Wuhan, China) and anti-SHIP2 rabbit antibodies (1:50, CST, Beverly, MA, USA) at 4 ℃ overnight. After blocking the cells with goat serum at room temperature for 30 min, samples were then incubated with Alexa Flour 488 anti-rabbit IgG antibodies and Alexa Flour594 anti-mouse IgG antibodies (1:200, Proteintech, Wuhan, China) for 2 h. Finally, DAPI staining was used to determine the nuclei (Beyotime Biotechnology, Shanghai, China). Images were captured using the ZEISS LSM880 confocal laser scanning microscope (Carl Zeiss, Oberkochen, Germany).

MGC-803 cells were fixed with 4% paraformaldehyde for 10 min at room temperature and then treated with phosphate-buffered saline (PBS) containing 0.1% Triton X-100. After blocking with goat serum for 30 min at room temperature, the cells were incubated with anti-SHIP2 rabbit antibodies (1:50, Abcam, Cambridge, UK) and anti-IQGAP2 mouse antibodies (1:50, Santa Cruz, Santa Cruz, CA, USA) at 4 °C overnight. After incubation with Alexa Flour 488 anti-rabbit IgG antibodies and Alexa Flour 594 anti-mouse IgG antibodies (1:200, Proteintech, Wuhan, China) for 2 h and then nuclei staining with 4’, 6-diamidino-2-phenylindole (DAPI, Beyotime Biotechnology, Shanghai, China), specimens were observed under a confocal laser scanning microscope (Carl Zeiss, Oberkochene, Germany).