Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [2]
- Immunocytochemistry [1]
- Immunohistochemistry [1]
- Chromatin Immunoprecipitation [1]
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Validation data
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- Product number
- PA5-18498 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Ku80 Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- This antibody is tested in Peptide ELISA: antibody detection limit dilution 16,000.
- Reactivity
- Human
- Host
- Goat
- Isotype
- IgG
- Vial size
- 100 µg
- Concentration
- 0.5 mg/mL
- Storage
- -20° C, Avoid Freeze/Thaw Cycles
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western Blot staining of HeLa cell lysate using Product # PA5-18498 at a concentration of 0.1 µg/mL, the primary antibody incubation was 1 hour and the detection method was chemiluminescence.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-Ku80 Polyclonal Antibody (Product # PA5-18498) and a 80 kDa band corresponding to Ku80 was observed in the cell lines tested along with very faint uncharacterized bands (*) at 50 kDa and 38 kDa. 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 (0.5 ug/ml) and detected by chemiluminescence with Rabbit anti-Goat IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP conjugate (Product # A27014, 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 Polyclonal Antibody (Product # PA5-18498) at 5 µg/mL in 0.1% BSA, incubated at 4 degree Celsius overnight and then with Rabbit anti-Goat IgG (H+L), Superclonal™ Recombinant Secondary Antibody, Alexa Fluor 488 conjugate (Product # A27012) 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
- Immunohistochemical analysis of Ku80 in paraffin embedded human spleen using a Ku80 polyclonal antibody (Product #PA5-18498) at a concentration of 5 µg/mL. Steamed antigen retrieval was performed with pH 6 buffered citrate. Samples were then stained with alkaline phosphatase.
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 Polyclonal Antibody (Product # PA5-18498, 2.5 µg) on sheared chromatin from camptothecin treated HeLa cells using the MAGnify ChIP System kit (Product # 49-2024). Normal Goat 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.