Tissue/cell: mouse heart tissue; 4% Paraformaldehyde-fixed and paraffin-embedded; Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum) at 37℃ for 20 min; Incubation: Anti-NME1/Nm23-H1/NDKA Polyclonal Antibody, Unconjugated(PC1066) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(A10026) and DAB staining

Blank control: RSC96(blue). Primary Antibody:Rabbit Anti-NME1 antibody(PC1066), Dilution: 1μg in 100 μL 1X PBS containing 0.5% BSA; Isotype Control Antibody: Rabbit IgG(orange) ,used under the same conditions ); Secondary Antibody: Goat anti-rabbit IgG-PE(white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA. Protocol The cells were fixed with 2% paraformaldehyde (10 min) , then permeabilized with 90% ice-cold methanol for 30 min on ice. Antibody (PC1066, 1μg /1x10^6 cells) were incubated for 30 min on the ice, followed by 1 X PBS containing 0.5% BSA + 1 0% goat serum (15 min) to block non-specific protein-protein interactions. Then the Goat Anti-rabbit IgG/PE antibody was added into the blocking buffer mentioned above to react with the primary antibody of PC1066 at 1/200 dilution for 30 min on ice. Acquisition of 20,000 events was performed.

Blank control (blue line): A549 (blue). Primary Antibody (green line): Rabbit Anti-NME1 antibody (PC1066) Dilution: 1μg /10^6 cells; Isotype Control Antibody (orange line): Rabbit IgG . Secondary Antibody (white blue line): Goat anti-rabbit IgG-PE Dilution: 1μg /test. Protocol The cells were fixed with 2% paraformaldehyde (10 min) , then permeabilized with 90% ice-cold methanol for 30 min on ice. Cells 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.

Sample: Hela(Human) Cell Lysate at 30 ug Primary: Anti-NM23A (PC1066) at 1/1000 dilution Secondary: Goat Anti-Rabbit IgG at 1/20000 dilution Predicted band size: 17 kD Observed band size: 18 kD

Hela cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum) at 37°C for 20 min; Antibody incubation with (NM23A) polyclonal Antibody, Unconjugated (PC1066) 1:25, 90 minutes at 37°C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue) was used to stain the cell nuclei.