Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [6]
- Immunocytochemistry [5]
- Immunohistochemistry [3]
- Flow cytometry [1]
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- Product number
- MA5-32627 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- TDP-43 Recombinant Rabbit Monoclonal Antibody (JM51-10)
- Antibody type
- Monoclonal
- Antigen
- Synthetic peptide
- Description
- Recombinant rabbit monoclonal antibodies are produced using in vitro expression systems. The expression systems are developed by cloning in the specific antibody DNA sequences from immunoreactive rabbits. Then, individual clones are screened to select the best candidates for production. The advantages of using recombinant rabbit monoclonal antibodies include: better specificity and sensitivity, lot-to-lot consistency, animal origin-free formulations, and broader immunoreactivity to diverse targets due to larger rabbit immune repertoire.
- Reactivity
- Human, Mouse, Rat, Zebrafish
- Host
- Rabbit
- Isotype
- IgG
- Antibody clone number
- JM51-10
- Vial size
- 100 µL
- Concentration
- 1 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
- Western blot analysis of TDP-43 in different lysates using a Monoclonal antibody (Product #MA5-32627) at a dilution of 1:500. Positive control: Line 1: Hela, Line 2: K563.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot of TDP-43 was performed by loading 50 µg of WT (lane 1) and TARDBP CRISPR KO (lane 2) HAP1 cell lysates in RIPA buffer onto a 5-16% gradient polyacrylamide gel. Proteins on the blots were visualized with Ponceau staining (below immunoblot). Proteins were transferred to nitrocellulose membrane and blocked in 5% milk for 1 hr. TDP-43 was detected at approximately 45 kDa using a TDP-43 recombinant monoclonal antibody (Product # MA5-32627) at a dilution of 1:1000 in 5% BSA in TBS with 0.1% Tween 20 (TBST) overnight at 4 deg. The peroxidase-conjugated secondary antibody (Product # 65-6120) was diluted to 0.2 µg/mL in TBST with 5% milk for 1 hr. Chemiluminescent detection was performed using Pierce ECL Western Blotting Substrate (Product # 32106). Data courtesy of YCharOS Inc., an open science company with the mission of characterizing commercially available antibodies using knockout validation.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of TDP-43 in different lysates using a Monoclonal antibody (Product #MA5-32627) at a dilution of 1:500. Positive control: Line 1: Hela, Line 2: K563.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- CRISPR-Cas9 mediated genome editing of TARDBP was achieved by using LentiArray™ Lentiviral sgRNA (Product # A32042) (Assay ID CRISPR1087594_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Fig (a) Western blot analysis of TARDBP was performed by loading 30 µg of HeLa CAS9 (Lane 1) and HeLa CAS9 cells transduced with TARDBP Lentiviral sgRNA (Lane 2) whole cell extracts. The blot was probed with Anti-TDP-43 Recombinant Rabbit Monoclonal Antibody (JM51-10) (Product # MA5-32627) using 1:1000 dilution and Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036). A reduced signal in sgRNA transduced cells using the LentiArray™ CRISPR product line confirms that antibody is specific to TARDBP (Fig (b)). An uncharacterized band was observed at ~80 kDa.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of TDP-43 was achieved by transfecting HeLa with TDP-43 specific siRNAs (Silencer® select Product # S23829, S23830). Western blot analysis (Fig. a) was performed using Whole cell extracts from the TDP-43 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with TDP-43 Recombinant Rabbit Monoclonal Antibody (JM51-10) (Product # MA5-32627, 1:5000 dilution ) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 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 TDP-43.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-TDP-43 Recombinant Rabbit Monoclonal Antibody (JM51-10) (Product # MA5-32627) and a 43kDa band corresponding to TDP-43 was observed across all cell lines and tissues tested. Whole cell extracts (30 µg lysate) of HeLa (Lane 1), HEK-293 (Lane 2), Caco-2 (Lane 3), MCF7 (Lane 4), Neuro-2a (Lane 5), PC-12 (Lane 6) and Mouse Brain (Lane 7) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein 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 (1:5000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036,1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005). A 35kDa cleaved fragment of TDP-43 was also observed.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunocytochemical analysis of TDP-43 in 293T cells using a TDP-43 Monoclonal antibody (Product # MA5-32627) as seen in red. The nuclear counter stain is DAPI (blue). Cells were fixed in paraformaldehyde, permeabilised with 0.25% Triton X100/PBS.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunocytochemical analysis of TDP-43 in Hela cells using a TDP-43 Monoclonal antibody (Product # MA5-32627) as seen in red. The nuclear counter stain is DAPI (blue). Cells were fixed in paraformaldehyde, permeabilised with 0.25% Triton X100/PBS.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunocytochemical analysis of TDP-43 in SH-SY5Y cells using a TDP-43 Monoclonal antibody (Product # MA5-32627) as seen in red. The nuclear counter stain is DAPI (blue). Cells were fixed in paraformaldehyde, permeabilised with 0.25% Triton X100/PBS.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence of TDP-43 was performed using HAP1 wild-type and TARDBP KO cells that were transfected with a green or a far-red fluorescent dye, respectively. Post-transfection, WT and KO cells were mixed and plated to a 1:1 ratio on coverslips as a mosaic and incubated for 24 hrs. Cells were fixed in 4% PFA (in PBS) or methanol for 15 min; cells were permeabilized with 0.1% Triton X-100 for 10 min at RT and blocked with PBS with 5% BSA, 5% goat serum, and 0.01% Triton X-100 for 30 min. Cells were stained with the TDP-43 recombinant monoclonal antibody (Product # MA5-32627) at a 1:1000 dilution overnight at 4 deg. Secondary antibody incubation was performed using 1 µg/mL of Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor 555 antibody (Product # A21429) together with DAPI for 1 hr. Imaging was performed with a 40X oil objective and analysis was performed using Image J. Cell image represents a single focal plane; WT and KO cells are outlined with a yellow (WT) or magenta (KO) dashed line. Data courtesy of YCharOS Inc., an open science company with the mission of characterizing commercially available antibodies using knockout validation.
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- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of TDP-43 was performed using 70% confluent log phase HeLa 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 TDP-43 Recombinant Rabbit Monoclonal Antibody (JM51-10) (Product # MA5-32627) 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), (1:2000 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.
Supportive validation
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- Invitrogen Antibodies (provider)
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- Experimental details
- Immunohistochemical analysis of TDP-43 of paraffin-embedded Human spleen tissue using a TDP-43 Monoclonal antibody (Product #MA5-32627). Counter stained with hematoxylin.
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- Invitrogen Antibodies (provider)
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- Experimental details
- Immunohistochemical analysis of TDP-43 of paraffin-embedded Human pancreas tissue using a TDP-43 Monoclonal antibody (Product #MA5-32627). Counter stained with hematoxylin.
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- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemical analysis of TDP-43 of paraffin-embedded Mouse placenta tissue using a TDP-43 Monoclonal antibody (Product #MA5-32627). Counter stained with hematoxylin.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Flow Cytometric analysis of TDP-43 in Hela cells using a TDP-43 Monoclonal Antibody (Product # MA5-32627) at a dilution of 1:50, as seen in red compared with an unlabelled control (cells without incubation with primary antibody; black). Alexa Fluor 488-conjugated goat anti rabbit IgG was used as the secondary antibody.