Legumain Rabbit Polyclonal Antibody

• Catalog Number: orb5765
• Category: Antibodies
• Description: Legumain Rabbit Polyclonal Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: FC, IF, IHC-Fr, IHC-P, WB
• Predicted Reactivity: Bovine, Canine, Equine
• Reactivity: Human, Mouse, Rat
• Isotype: IgG
• Immunogen: KLH conjugated synthetic peptide derived from human Legumain (201-300/433aa)
• Antibody Type: Primary Antibody
• Concentration: 1mg/ml
• Dilution range: WB=1:500-2000, IHC-P=1:100-500, IHC-F=1:100-500, IF=1:100-500, Flow-Cyt=3ug/Test
• Form/Appearance: Liquid
• Conjugation: Unconjugated
• MW: 46 kDa
• Target: LGMN
• UniProt ID: Q99538
• RRID: AB_10925536
• Storage: Maintain refrigerated at 2-8°C for up to 2 weeks. For long term storage store at -20°C in small aliquots to prevent freeze-thaw cycles.
• Buffer/Preservatives: 0.01M TBS (pH7.4) with 1% rAlbumin, 0.02% Proclin300 and 50% Glycerol.
• Alternative names: AEP; Asparaginyl endopeptidase; EC 3.4.22.34; LGMN
• Note: For research use only
• Expiration Date: 12 months from date of receipt.

Blank control: HL60. Primary Antibody (green line): Rabbit Anti-Legumain antibody (orb5765), 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 20000 events was performed.

Overlay histogram showing Mouse spleen cells (red line). The cells were fixed with 1% paraformaldehyde (10 min). The cells were then incubated with the antibody (2 ug/1x106 cells) for 30 min at 22-25°C. Isotype control antibody (black line) was rabbit IgG (2 ug/1x106 cells) used under the same conditions. Acquisition of events were used for analysis.

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 (orb5765) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructionsand DAB staining.

Sample: Kidney (Mouse) Lysate at 40 ug, Primary: Anti-Legumain (orb5765) at 1/300 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 46 kD, Observed band size: 55 kD.

Sample: Lane 1: Human JAR cell lysates, Lane 2: Human MCF-7 cell lysates, Primary: Anti-Legumain (orb5765) at 1/1000 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 46 kDa, Observed band size: 55 kDa.

Sample: Placenta (Mouse) Lysate at 40 ug, Primary: Anti-Legumain (orb5765) at 1/300 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 46 kD, Observed band size: 55 kD.