Antibody Type: Recombinant Rabbit Monoclonal

Application: WB,IHC-P,IHC-Fr,ICC/IF,IF,FCM

Reactivity: Human,Mouse,Rat

IgG

8P334

Human,Mouse,Rat

1. HT-1080 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 (Smad3) monoclonal Antibody, Unconjugated (TMAB-01740) 1:100, 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 nucleus.
2. Blank control: HUVEC.
Primary Antibody (green line): Rabbit Anti-Smad3 antibody (TMAB-01740)
Dilution: 1 μg/Test;
Secondary Antibody: Goat anti-rabbit IgG-FITC
Dilution: 0.5 μg/Test.
Protocol
The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 0.1% PBST for 20 min at room temperature. The cells were then incubated in 5% 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.
3. Paraformaldehyde-fixed, paraffin embedded Mouse Liver; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Antibody incubation with Smad3 Monoclonal Antibody, Unconjugated (TMAB-01740) at 1:200 overnight at 4°C, followed by conjugation to the secondary antibody and DAB staining.
4. Paraformaldehyde-fixed, paraffin embedded Rat Liver; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Antibody incubation with Smad3 Monoclonal Antibody, Unconjugated (TMAB-01740) at 1:200 overnight at 4°C, followed by conjugation to the secondary antibody and DAB staining.
5. Paraformaldehyde-fixed, paraffin embedded Human Liver; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Antibody incubation with Smad3 Monoclonal Antibody, Unconjugated (TMAB-01740) at 1:200 overnight at 4°C, followed by conjugation to the secondary antibody and DAB staining.
6. Paraformaldehyde-fixed, paraffin embedded Human Colon Cancer; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Antibody incubation with Smad3 Monoclonal Antibody, Unconjugated (TMAB-01740) at 1:200 overnight at 4°C, followed by conjugation to the secondary antibody and DAB staining.
7. Paraformaldehyde-fixed, paraffin embedded Human Pancreatic Cancer; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Antibody incubation with Smad3 Monoclonal Antibody, Unconjugated (TMAB-01740) at 1:200 overnight at 4°C, followed by conjugation to the secondary antibody and DAB staining.
8. Paraformaldehyde-fixed, paraffin embedded Human Kidney; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Antibody incubation with Smad3 Monoclonal Antibody, Unconjugated (TMAB-01740) at 1:200 overnight at 4°C, followed by conjugation to the secondary antibody and DAB staining.
9. 25 μg total protein per Lane of various lysates probed with Smad3 monoclonal antibody, unconjugated (TMAB-01740) at 1:1000 dilution and 4°C overnight incubation. Followed by conjugated secondary antibody incubation at RT for 60 min.

WB,IHC-P,IHC-Fr,ICC/IF,IF,FCM

WB: 1:500-2000; IHC-P: 1:100-500; IHC-Fr: 1:100-500; ICC/IF: 1:50-200; IF: 1:100-500; FCM: 1ug/Test

Monoclonal

A synthesized peptide: human SMAD3

Human

4088

Response to hypoxia,SMAD family,SMADs,Hypoxia,Cancer,SMADs,Cytoplasmic