Antibody Type: Rabbit Polyclonal

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

Reactivity: Human,Rat

IgG

Human,Rat

1. Sample:
Lane 1: HepG2 (Human) Cell Lysate at 30 μg
Lane 2: A549 (Human) Cell Lysate at 30 μg
Lane 3: U251 (Human) Cell Lysate at 30 μg
Primary: Anti-Osteopontin (TMAB-01299) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 60-65 kDa
Observed band size: 60 kDa
2. Blank control: HepG2. Primary Antibody (green line): Rabbit Anti-Osteopontin antibody (TMAB-01299)
Dilution: 1 μg/10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG.
Secondary Antibody: Goat anti-rabbit IgG-AF647
Dilution: 1 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.
Lane 1: Human HepG2 cell lysates
Lane 2: Human U87MG cell lysates
Lane 3: Human MG63 cell lysates
Lane 4: Human U2OS Cell Lysates
Primary: Anti-Osteopontin (TMAB-01299) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 60 kDa
Observed band size: 60 kDa
4. Paraformaldehyde-fixed, paraffin embedded (Rat lung); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 min; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (osteopontin) Polyclonal Antibody, Unconjugated (TMAB-01299) at 1:400 overnight at 4°C, followed by a conjugated secondary for 20 min and DAB staining.

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

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

Polyclonal

KLH conjugated synthetic peptide: rat osteopontin

Rat

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