Sample:
Lane 1: Human SW480 cell lysates
Primary: Anti- TIMP-2 (bsm-51320M) at 1/500 dilution
Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution
Predicted band size: 24 kD
Observed band size: 24 kD

Sample:
Lane 1: Human HT-1080 cell lysates
Primary: Anti- TIMP-2 (bsm-51320M) at 1/2000 dilution
Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution
Predicted band size: 24 kD
Observed band size: 24 kD

Paraformaldehyde-fixed, paraffin embedded (human kidney sections); 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 (TIMP-2) Monoclonal Antibody, Unconjugated (bsm-51320M) at 1:25 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.

Blank control:K562.
Primary Antibody (green line): Mouse Anti-TIMP-2 antibody (bsm-51320M)
Dilution:1:25;
Secondary Antibody : Goat anti-mouse IgG-AF488
Dilution:1:400.
Protocol
The cells were fixed with 2% PFA (10min at room temperature)and then permeabilized with 90% methanol for 10 min.The cells were then incubated in 2%BSA to block non-specific protein-protein interactions for 30 min at room temperature .Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.