MA5-13110

antibody from Invitrogen Antibodies

Targeting: XRCC6 D22S671, D22S731, G22P1, KU70, ML8

Provider product page for MA5-13110

Western blot

Immunocytochemistry

Immunoprecipitation

Immunohistochemistry

Flow cytometry

Chromatin Immunoprecipitation

Other assay

Antibody data

Antibody Data
Antigen structure
References [49]
Comments [0]
Validations
Western blot [3]
Immunohistochemistry [1]
Flow cytometry [3]
Chromatin Immunoprecipitation [1]
Other assay [4]

Submit

Product number: MA5-13110 - Provider product page
Provider: Invitrogen Antibodies
Product name: Anti-Ku70 Monoclonal Antibody (N3H10)
Antibody type: Monoclonal
Antigen: Other
Description: MA5-13110 targets Ku (p70) in immunofluorescence, immunohistochemistry (paraffin), immunoprecipitation, FACS and Western blot applications and shows reactivity with Human, Non-human primate, and Xenopus laevis samples. Reacts with mouse and hamster in Western blot applications only. The MA5-13110 immunogen is human placental extract designated as PSE1-PL. Antibodies to this protein (and modification) were previously sold as part of a Thermo Scientific Cellomics High Content Screening Kit. This replacement antibody is now recommended for researchers who need an antibody for high content cell based assays. It has been thoroughly tested and validated for cellular immunofluorescence (IF) applications. Further optimization including the selection of the most appropriate fluorescent Dylight conjugated secondary antibody may have to be performed for your high content assay.
Reactivity: Human, Mouse, Hamster, Xenopus
Host: Mouse
Isotype: IgG
Antibody clone number: N3H10
Vial size: 500 µL
Concentration: 0.2 mg/mL
Storage: 4° C

XRCC6 protein structure - MA5-13110 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of Ku (P70) was performed by loading 50 µg of HepG2 whole cell lysate per well onto a SDS-PAGE gel. Proteins were transferred to a membrane and blocked with 5% nonfat dry milk in PBS-0.1% Tween-20 for 30 minutes. The membrane was probed with a Ku (P70) monoclonal antibody (Product # MA5-13110) at a concentration of 0.5 µg/mL overnight at 4°C on a rocking platform, washed in PBS-0.1%Tween-20, and probed with an anti-mouse IgG-HRP secondary antibody for 1 hour. Detection was performed using a chemiluminescent substrate.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of Ku70 was performed by loading whole cell lysate from 2.5 x 105 mouse embryonic fibroblasts (in 1XSDS sample buffer with 2-ME) onto a 4-12 % Bis-Tris polyacrylamide gel (Product # WG1402BOX). Proteins were transferred to PVDF membrane with wet/tank transfer. Membrane was blocked in 5% milk/TBST. Ku70 was detected at approximately 76 kDa using a monoclonal anti-Ku70 antibody (Product # MA5-13110) at a dilution of 1:1000 in 5% milk/TBST at 4°C overnight, followed by a secondary antibody HRP-anti-mouse at a dilution of 1:5000 at room temperature for 1 hour. Chemiluminescent detection was performed using Pierce ECL Western Blotting Substrate (Product # PI-32209). Data courtesy of Antibody Data Exchange Program.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of Ku70 was achieved by transfecting HeLa cells with Ku70 specific siRNAs (Silencer® select Product # s5457, s5455). Western blot analysis (Fig. a) was performed using whole cell extracts from the Ku70 knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with Ku70 Monoclonal Antibody (N3H10) (Product # MA5-13110, 1 µg/ml) and Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25 µg/ml, 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 Ku70.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of Ku (p70) in Hela cells compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/mL, fixed with 2% paraformaldehyde, washed with PBS, and incubated with Ku (p70) monoclonal antibody (Product # MA5-13110) at a dilution of 0.25 µg/test for 60 min at room temperature. Cells were then blocked in a solution of 2% BSA-PBS for 30 min at room temperature, incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated goat anti-mouse IgG (H+L) secondary antibody, and re-suspended in PBS for FACS analysis.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of Ku (p70) in HepG2 cells compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/mL, fixed with 2% paraformaldehyde, washed with PBS, and incubated with Ku (p70) monoclonal antibody (Product # MA5-13110) at a dilution of 0.25 µg/test for 60 min at room temperature. Cells were then blocked in a solution of 2% BSA-PBS for 30 min at room temperature, incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated goat anti-mouse IgG (H+L) secondary antibody, and re-suspended in PBS for FACS analysis.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of Ku (p70) in C2C12 cells compared to an isotype control (blue). Cells were harvested, adjusted to a concentration of 1-5x10^6 cells/mL, fixed with 2% paraformaldehyde, washed with PBS, and incubated with Ku (p70) monoclonal antibody (Product # MA5-13110) at a dilution of 0.5 µg/test for 60 min at room temperature. Cells were then blocked in a solution of 2% BSA-PBS for 30 min at room temperature, incubated for 40 min at room temperature in the dark using a Dylight 488-conjugated goat anti-mouse IgG (H+L) secondary antibody, and re-suspended in PBS for FACS analysis.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunoprecipitation of Ku (P70) using Ku (P70) Monoclonal Antibody (Product # MA5-13110) on denatured Human BT474 Cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunoprecipitation of Ku (P70) using Ku (P70) Monoclonal Antibody (Product # MA5-13110) on Native Human T47D Cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of Ku (green) in HeLa cells either left untreated (left panel) or treated with 50uM etoposide (right panel) for 3 hours. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA (Product # 37525) for 15 minutes at room temperature. Cells were probed with a Ku (p70) monoclonal antibody (Product # MA5-13110) at a dilution of 1:100 for at least 1 hour at room temperature, washed with PBS, and incubated with DyLight 488 goat anti-mouse IgG secondary antibody (Product # 35502) at a dilution of 1:400 for 30 minutes at room temperature. F-Actin (red) was stained with DyLight 554 Phalloidin (Product # 21834) and nuclei (blue) were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Thermo Scientific ArrayScan or ToxInsight Instrument at 20X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of Ku70 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 1% BSA for 1 hour at room temperature. The cells were labeled with Ku70 Monoclonal Antibody (N3H10) (Product # MA5-13110) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:2000 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.