BBC3 Antibody

• Catalog Number: orb1239911
• Category: Antibodies
• Description: BBC3 Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: ELISA, ICC, IF, WB
• Predicted Reactivity: Rat
• Reactivity: Human, Mouse
• Isotype: IgG
• Immunogen: Anti-PUMA antibody (orb1239911) was raised against a peptide corresponding to 14 amino acids near the carboxyl terminus human PUMA isoform 1. The immunogen is located within the last 50 amino acids of PUMA.
• Concentration: 1 mg/mL
• Dilution range: WB: 1-4 μg/mL; IF: 2 μg/mL; ICC: 1 μg/mL. Antibody validated: Western Blot in human and mouse samples; Immunocytochemistry and Immunofluorescence in human samples. All other applications and species not yet tested.
• Form/Appearance: Liquid
• Conjugation: Unconjugated
• MW: 23 kDa
• Target: BBC3
• UniProt ID: Q96PG8
• NCBI: NP_055232
• Storage: PUMA antibody can be stored at 4°C for three months and -20°C, stable for up to one year. As with all antibodies care should be taken to avoid repeated freeze thaw cycles. Antibodies should not be exposed to prolonged high temperatures.
• Buffer/Preservatives: PUMA Antibody is supplied in PBS containing 0.02% sodium azide.
• Alternative names: PUMA Antibody: JFY1, PUMA, JFY-1, Bcl-2-binding co
• Note: For research use only
• Expiration Date: 12 months from date of receipt.

Western Blot Validation of PUMA in K562 and 3T3/NIH Cells. Loading: 15 µg of lysates per lane. Antibodies: orb1239911 (2 µg/mL), 1 h incubation at RT in 5% NFDM/TBST. Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.

Independent Antibody Validation (IAV) via Protein Expression Profile in Cell Lines. Loading: 20 µg of lysates per lane. Antibodies: orb1239911 (3 µg/mL), orb1239919 (2 µg/mL), beta-actin (1 µg/mL) and GAPDH (0.02 µg/mL), 1 h incubation at RT in 5% NFDM/TBST. Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.

Validation with PUMA siRNA Knockdown in 293 Cells. 293 cells were transfected with control siRNAs (lane 1) or PUMA siRNAs (lane 2) Loading: 15 µg of 293 whole cell lysates per lane. Antibodies: orb1239911 (2 µg/mL), beta-actin (1 µg/mL) and GAPDH (0.02 µg/mL), 1 h incubation at RT in 5% NFDM/TBST. Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.

Sensitivity Test for PUMA in 2983 Cells. Loading: Lysates/proteins at 15 µg per lane. Antibodies: orb1239911 (lane 1-3: 1, 2 and 4 µg/mL). 1 h incubation at RT in 5% NFDM/TBST. Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.

Immunofluorescence Validation of PUMA in K562 Cells. Immunofluorescent analysis of 4% paraformaldehyde-fixed K562 cells labeling PUMA with orb1239911 at 2 µg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (red). Image showing cytosol staining on K562 cells.

Immunocytochemistry Validation of PUMA in K562 Cells. Immunocytochemical analysis of K562 cells using anti-PUMA antibody (orb1239911) at 1 µg/ml. Cells was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4°C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin.

KD Validation of PUMA in HOSE-RasV12 Cells (Elgendy et al., 2011). HOSE-RasV12 cells were transfected with control shRNA plasmid or shRNA plasmids (KD) targeted against Noxa or Puma, as indicated. PUMA expression was not observed in PUMA KD cells detected by anti-PUMA antibodies (orb1239911).

Induction Validation of PUMA in Primary Cortical Neurons (Sabirzhanov et al., 2014). PUMA protein levels were increased in etoposide-treated primary cortical neurons detected by anti-PUMA antibodies (orb1239911).

KD Validation of PUMA PUMA in Tet Cells (Han et al., 2010). Immunoblot analyses of Tet-induced p53 cells treated with NOXA, Puma, Bim or non-targeting siRNAs that were utilized in this experiment. PUMA protein levels were markedly reduced in PUMA KD cells detected by anti-PUMA antibodies (orb1239911).