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Western blotAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [2]
- Immunocytochemistry [1]
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- Product number
- 711782 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- LDHA Recombinant Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 100 µg
- Concentration
- 0.5 mg/mL
- Storage
- Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of LDHA was achieved by transfecting PC3 cells with LDHA specific validated siRNA (Silencer® select Product # s350). Western blot analysis was performed using membrane extracts from the LDHA knock down cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with Anti-LDHA Recombinant Rabbit Polyclonal Antibody (Product # 711782, 1-3 µg/mL) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to LDHA.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis was performed on Membrane enriched extracts (30 µg lysate) of PC-3 (Lane 1), DU 145 (Lane 2), LNCaP (Lane 3), MCF-7 (Lane 4), MDA-MB-231 (Lane 5), PANC-1 (Lane 6), SK-OV-3 (Lane 7), HeLa (Lane 8) and Hep G2 (Lane 9). The blots were probed with Anti-LDHA Recombinant Rabbit Polyclonal Antibody (Product # 711782, 2.5 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). A 35 kDa band corresponding to LDHA was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®4-12% Bis-Tris gel (Product # NP0322BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- For immunofluorescence analysis, PC3 cells were fixed and permeabilized for detection of endogenous LDHA using Anti- LDHA Recombinant Rabbit Polyclonal Antibody (Product # 711782, 5 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Panel a) shows representative cells that were stained for detection and localization of LDHA protein (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). Panel c) represents cytoskeletal F-actin staining using Rhodamine Phalloidin (Product # R415, 1:300). Panel d) is a composite image of Panels a, b and c clearly demonstrating cytoplasmic localization of LDHA. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
