Sample:
Spinal cord (Mouse) Lysate at 40 ug
Primary: Anti- GPR125 (bs-12021R) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 143 kD
Observed band size: 183 kD

Paraformaldehyde-fixed, paraffin embedded (mouse brain tissue); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (GPR125) Polyclonal Antibody, Unconjugated (bs-12021R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Blank control (blue line): Mouse colon (blue).
Primary Antibody (green line): Rabbit Anti-GPR125 antibody (bs-12021R)
Dilution: 1μg /10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG .
Secondary Antibody (white blue line): F(ab’)2 fragment goat anti-rabbit IgG-FITC
Dilution: 1μg /test.
Protocol
The cells were fixed with 2% paraformaldehyde for 10 min at room temperatureCells stained with Primary Antibody for 30 min at room temperature. The cells were then incubated in 1 X PBS/2%BSA/10% goat serum to block non-specific protein-protein interactions followed by the antibody for 15 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.