MA5-14953
antibody from Invitrogen Antibodies
Targeting: XRCC5
KARP-1, KU80, Ku86, KUB2
Antibody data
- Antibody Data
- Antigen structure
- References [1]
- Comments [0]
- Validations
- Western blot [2]
- Immunocytochemistry [1]
- Chromatin Immunoprecipitation [1]
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Validation data
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- Product number
- MA5-14953 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Ku80 Monoclonal Antibody (S.669.4)
- Antibody type
- Monoclonal
- Antigen
- Synthetic peptide
- Description
- It is not recommended to aliquot this antibody.
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Antibody clone number
- S.669.4
- Vial size
- 100 µL
- Concentration
- 7 µg/mL
- Storage
- -20°C
Submitted references A Fibrinogen Alpha Fragment Mitigates Chemotherapy-Induced MLL Rearrangements.
Eberle J, Wiehe RS, Gole B, Mattis LJ, Palmer A, Ständker L, Forssmann WG, Münch J, Gebhardt JCM, Wiesmüller L
Frontiers in oncology 2021;11:689063
Frontiers in oncology 2021;11:689063
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of Ku80 was achieved by transfecting HeLa with Ku80 specific siRNAs (Silencer® select Product # s14952, s14953). Western blot analysis (Fig. a) was performed using whole cell extracts from the Ku80 knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with Ku80 Monoclonal Antibody (Product # MA5-14953, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 0.25µg/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 Ku80.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-Ku80 Monoclonal Antibody (S.669.4), (Product # MA5-14953) and a 80 kDa band corresponding to Ku80 was observed in the cell lines tested. Modified whole cell extracts (1%SDS) (30 µg lysate) of HeLa (Lane 1), MCF7 (Lane 2), K-562 (Lane 3), Jurkat (Lane 4), COS-7 (Lane 5), A549 (Lane 6) and HEK-293 (Lane 7) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX). 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 (1:1000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP conjugate (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).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Ku80 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 1 hour at room temperature. The cells were labeled with Ku80 Monoclonal Antibody (Product # MA5-14953) at 1:200 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then with Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, Alexa Fluor 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing staining in nucleus. 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
- Chromatin Immunoprecipitation (ChIP) assay of endogenous Ku80 protein using Anti-Ku80 Antibody: ChIP was performed using Anti-Ku80 Rabbit Monoclonal Antibody (Product # MA5-14953, 5 µg) on sheared chromatin from Camptothecin treated HeLa cells using the MAGnify ChIP System kit (Product # 49-2024). Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by qPCR using primers binding to GAPDH transcriptional start site, GAPDH gene body (+2Kb), CDKN1A intron 1 and SAT2 satellite repeats. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.