702773

antibody from Invitrogen Antibodies

Targeting: DNAJC13 KIAA0678, RME8

Antibody data

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Validation data

Product number: 702773 - Provider product page
Provider: Invitrogen Antibodies
Product name: DNAJC13 Recombinant Rabbit Monoclonal Antibody (11H13L5)
Antibody type: Monoclonal
Antigen: Other
Description: This antibody is predicted to react with Monkey, Cat, Goat, Sheep
Antibody clone number: 11H13L5
Concentration: 0.5 mg/mL

DNAJC13 protein structure - 702773 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on Whole cell extracts (30 µg lysate) of U-87 MG (Lane 1), HeLa (Lane 2), U-2 OS (Lane 3), MCF-7 (Lane 4), Jurkat (Lane 5), NIH/3T3 (Lane 6), PC-3 (Lane 7) and A-431 (Lane 8). The blots were probed with Anti-RME-8 Recombinant Rabbit Monoclonal Antibody (Product # 702773, 2.5 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:4000 dilution). A ~254 kDa band corresponding to RME-8 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 by wet transfer method. 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: Western blot analysis was performed on Whole cell extracts (30 µg lysate) of U-87 MG (Lane 1), HeLa (Lane 2), U-2 OS (Lane 3), MCF-7 (Lane 4), Jurkat (Lane 5), NIH/3T3 (Lane 6), PC-3 (Lane 7) and A-431 (Lane 8). The blots were probed with Anti-RME-8 Recombinant Rabbit Monoclonal Antibody (Product # 702773, 2.5 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:4000 dilution). A ~254 kDa band corresponding to RME-8 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 by wet transfer method. 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: Knockdown of RME8 was achieved by transfecting MCF7 cells with RME8 specific siRNA (Silencer® select Product # s23548 and s23550). Western blot analysis (Fig a) was performed using whole cell extract from the RME8 knock down cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with Anti-RME8 Recombinant Rabbit Monoclonal Antibody (Product # 702773, 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). Densitometric analysis of this Western blot is shown in histogram (Fig b). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to RME8.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: For immunofluorescence analysis, U-87 MG cells were fixed and permeabilized for detection of endogenous RME-8 using Anti- RME-8 Recombinant Rabbit Monoclonal Antibody (Product # 702773, 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 RME-8 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 endosome localization of RME-8. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of DNAJC13 was achieved by transfecting A-431 cells with specific siRNA (Silencer® select Product # s23548 + s23550). Immunofluorescence analysis was performed on A-431cells (untransfected, panel a-d), transfected with DNAJC13 specific siRNA (panel i-l) or non-specific scrambled siRNA (panels e-h). Cells were fixed, permeabilized, and labelled with Anti-DNAJC13 Recombinant Rabbit Monoclonal Antibody (Product # 702773, 1:100 dilution), followed by Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Nuclei (blue) were stained using ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962), and Rhodamine Phalloidin (Product # R415, 1:300) was used for cytoskeletal F-actin (red) staining. Significant reduction of signal was observed upon siRNA mediated knockdown (panel i-l) confirming specificity of the antibody to DNAJC13 (green). The images were captured at 60X magnification.