Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [3]
- Immunocytochemistry [4]
- Immunohistochemistry [3]
- Flow cytometry [1]
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- Product number
- MA5-34711 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- MBD3 Recombinant Rabbit Monoclonal Antibody (JB61-47)
- Antibody type
- Monoclonal
- Antigen
- Recombinant full-length protein
- Description
- Positive Control: U937, A549, NIH-3T3, SH-SY-5Y, rat testis tissue, human placenta tissue, mouse brain tissue.
- Reactivity
- Human, Mouse
- Host
- Rabbit
- Isotype
- IgG
- Antibody clone number
- JB61-47
- Vial size
- 100 µL
- Concentration
- 1 mg/mL
- Storage
- -20° C, Avoid Freeze/Thaw Cycles, store in dark
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of MBD3 in U937 cell lysate. Samples were incubated with MBD3 monoclonal antibody (Product # MA5-34711), at a dilution of 1:500.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of Methyl-CpG-binding protein MBD3 was achieved by transfecting MCF7 with MBD3 specific siRNAs (Silencer® select Product # S28737, S28736). Western blot analysis (Fig. a) was performed using nuclear enriched extracts from the Methyl-CpG-binding protein MBD3 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with MBD3 Recombinant Rabbit Monoclonal Antibody (JB61-47) (Product # MA5-34711, 1:500 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20,000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to MBD3.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using MBD3 Recombinant Rabbit Monoclonal Antibody (JB61-47) (Product # MA5-34711) and a 32 kDa band corresponding to Methyl-CpG-binding protein MBD3 was observed across the cell panel tested. Fresh nuclear enriched extracts (20 µg lysate) of A-673 (Lane 1), HEK-293 (Lane 2), MCF7 (Lane 3), A549 (Lane 4), Mouse Spleen (Lane 5), Mouse Testis (Lane 6) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX), 12 well. 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 with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20,000 dilution) using the iBright™ FL1500 Imaging System (Product # A44115). Chemiluminescent detection was performed using SuperSignal™ West Pico PLUS Chemiluminescent Substrate (Product # 34580).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Methyl-CpG-binding protein MBD3 was performed using 70% confluent log phase MCF7 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with MBD3 Recombinant Rabbit Monoclonal Antibody (JB61-47) (Product # MA5-34711) at 1:200 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790, 1:3000 dilution), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b: Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing nuclear localization. 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
- Immunofluorescent analysis of MBD3 in SH-SY-5Y cells (green). Samples were fixed in paraformaldehyde and permeabilised with 0.25% Triton X100/PBS, incubated with MBD3 monoclonal antibody (Product # MA5-34711), followed by DAPI (blue).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescent analysis of MBD3 in NIH-3T3 cells (green). Samples were fixed in paraformaldehyde and permeabilised with 0.25% Triton X100/PBS, incubated with MBD3 monoclonal antibody (Product # MA5-34711), followed by DAPI (blue).
- Submitted by
- Invitrogen Antibodies (provider)
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- Experimental details
- Immunofluorescent analysis of MBD3 in A549 cells (green). Samples were fixed in paraformaldehyde and permeabilised with 0.25% Triton X100/PBS, incubated with MBD3 monoclonal antibody (Product # MA5-34711), followed by DAPI (blue).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
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- Experimental details
- Immunohistochemistry analysis of MBD3 in paraffin-embedded rat testis tissue. Samples were incubated with MBD3 monoclonal antibody (Product # MA5-34711), and followed by hematoxylin.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry analysis of MBD3 in paraffin-embedded mouse brain tissue. Samples were incubated with MBD3 monoclonal antibody (Product # MA5-34711), and followed by hematoxylin.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry analysis of MBD3 in paraffin-embedded human placenta tissue. Samples were incubated with MBD3 monoclonal antibody (Product # MA5-34711), and followed by hematoxylin.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Flow cytometry of MBD3 in SH-SY-5Y cells (red) compared with an unlabelled control (cells without incubation with primary antibody; black). Samples were incubated with MBD3 monoclonal antibody (Product # MA5-34711) at a dilution of 1:100, followed by Alexa Fluor 488-conjugated goat anti-rabbit IgG.