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Western blotAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [2]
- Immunocytochemistry [2]
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- Product number
- 703370 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- MBD1 Recombinant Rabbit Monoclonal Antibody (12H8L4)
- Antibody type
- Monoclonal
- Antigen
- Other
- Description
- This antibody is predicted to react with Monkey, Bovine.
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Antibody clone number
- 12H8L4
- 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 MBD1 was achieved by transfecting HeLa cells with MBD1 specific siRNA (Silencer® select Product # s8547 & s8549). Western blot analysis (Fig a) was performed using modified whole cell extracts (1% SDS) from MBD1 knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with Anti-MBD1 Recombinant Rabbit Monoclonal Antibody (Product # 703370, 1:500 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in the histogram (Fig b). Loss of signal upon siRNA mediated knockdown confirms that antibody is specific to MBD1.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-MBD1 Recombinant Rabbit Monoclonal Antibody (Product # 703370) and a 67 kDa band corresponding to MBD1 was observed across the cell lines tested. Modified whole cell extracts (1% SDS) (30 µg lysate) of HeLa (Lane 1), HEK-293 (Lane 2), Hep G2 (Lane 3), MCF7 (Lane 4), U-2 OS (Lane 5), HCT 116 (Lane 6) and PC-3 (Lane 7) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:500 dilution) and detected by chemiluminescence Goat Anti-Rabbit IgG Secondary Antibody, HRP conjugate (Product # A27036, 1:4000 dilution). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- For immunofluorescence analysis, HeLa cells were fixed and permeabilized for detection of endogenous MBD1 using Anti-MBD1 Recombinant Rabbit Monoclonal Antibody (Product # 703370, 1:100 dilution) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # 703404, 1:2000). Panel a) shows representative cells that were stained for detection and localization of MBD1 protein (green), Panel b) is stained for nuclei (blue) using ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). 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 nuclear localization of MBD1. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- For immunocytochemistry analysis, HeLa cells were fixed and permeabilized for detection of endogenous MBD1 using Anti-MBD1 Recombinant Rabbit Monoclonal Antibody (Product # 703370, 1:100 dilution) 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 MBD1 protein (green), Panel b) is stained for nuclei (blue) using ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). 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 nuclear localization of MBD1. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
