Anti-Legumain Polyclonal Antibody is a Rabbit antibody targeting Legumain. Anti-Legumain Polyclonal Antibody can be used in WB,IHC-P,IHC-Fr,IF,FCM.

IgG

Human,Mouse (predicted:Rat,Dog,Cow,Horse)

1. Paraformaldehyde-fixed, paraffin embedded (mouse placenta); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (Legumain) Polyclonal Antibody, Unconjugated (TMAB-08270) at 1: 200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
2. Sample: Kidney (Mouse) Lysate at 40 μg
Primary: Anti-Legumain (TMAB-08270) at 1/300 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 46 kD
Observed band size: 55 kD
3. Sample: Placenta (Mouse) Lysate at 40 μg
Primary: Anti-Legumain (TMAB-08270) at 1/300 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 46 kD
Observed band size: 55 kD
4. Blank control: HL60.
Primary Antibody (green line): Rabbit Anti-Legumain antibody (TMAB-08270)
Dilution: 3 μg /10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG.
Secondary Antibody: Goat anti-rabbit IgG-PE
Dilution: 1 μg /test.
Protocol
The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 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 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.
5. Sample:
Lane 1: Human JAR cell lysates
Lane 2: Human MCF-7 cell lysates
Primary: Anti-Legumain (TMAB-08270) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 46 kDa
Observed band size: 55 kDa

WB: 1:500-2000; IHC-P: 1:100-500; IHC-Fr: 1:100-500; IF: 1:100-500; FCM: 3μg/Test

Polyclonal

KLH conjugated synthetic peptide: human Legumain

Human

LGMN

Legumain

Has a strict specificity for hydrolysis of asparaginyl bonds. Can also cleave aspartyl bonds slowly, especially under acidic conditions. May be involved in the processing of proteins for MHC class II antigen presentation in the lysosomal/endosomal system.