Mouse anti egfp

Bacillus subtilis-gCa and B. subtilis-gDa were grown in 5 ml LB medium at 37 °C, 220 rpm for 24 h, 48 h and 60 h. Then, the two strains of bacteria were washed three times by sterile phosphate-buffered saline (PBS). After washed, these were centrifuged for 5 min at 12,000×g at 4 °C and re-suspended in 500 μl PBS. And these bacteria were disrupted to release the target proteins by Ultrasonic Cell Crusher (Xianou Tech, China) as previously described [29 (link)]. Furthermore, the proteins were detected by western blotting (Fig. 1b, c), in which the mouse anti-EGFP (Sigma) and HRP-conjugated goat anti-mouse IgG (Sigma) were used. Then the blots were developed by ECL enhanced chemiluminescence. The images were captured using a 4600SF Gel Image System (Tanon, China).
Besides, the re-suspended bacteria were observed under Zeiss LSM710 confocal microscope (Zeiss, Germany). The images were analyzed using ZEN 2012 (Carl Zeiss) (Fig. 1d, e).