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Western blot
ImmunocytochemistryAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [1]
- ELISA [1]
- Chromatin Immunoprecipitation [1]
- Other assay [1]
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- Product number
- NBP2-59261 - Provider product page
- Provider
- Novus Biologicals
- Product name
- Rabbit Polyclonal Histone H2a Antibody
- Antibody type
- Polyclonal
- Description
- Peptide affinity purified.
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 50 ug
- Storage
- Store at -20C. Avoid freeze-thaw cycles.
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Supportive validation
- Submitted by
- Novus Biologicals (provider)
- Main image
- Experimental details
- Western Blot: Histone H2a [ac Lys5] Antibody [NBP2-59261] - Western blot was performed on whole cell (25 ug, lane 1) and histone extracts (15 ug, lane 2) from HeLa cells, and on 1 ug of recombinant histone H2A, H2B, H3 and H4 (lane 3, 4, 5 and 6, respectively) using the antibody against H2AK5ac. The antibody was diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The marker (kDa) is shown on the left
Supportive validation
- Submitted by
- Novus Biologicals (provider)
- Main image
- Experimental details
- ELISA: Histone H2a [ac Lys5] Antibody [NBP2-59261] - To determine the titer of the antibody, an ELISA was performed using a serial dilution of the antibody directed against H2AK5ac in antigen coated wells. The antigen used was a peptide containing the histone modification of interest. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:25,000.
Supportive validation
- Submitted by
- Novus Biologicals (provider)
- Main image
- Experimental details
- Chromatin Immunoprecipitation: Histone H2a [ac Lys5] Antibody [NBP2-59261] - ChIP assays were performed using human HeLa cells, the antibody against H2AK5ac and optimized PCR primer sets for qPCR. ChIP was performed using sheared chromatin from 1.5 million cells. A titration of the antibody consisting of 0.5, 1, 2 and, 5 ug per ChIP experiment was analysed. IgG (1 ug/IP) was used as negative IP control. QPCR was performed with primers for a region approximately 1 kb upstream of the GAPDH and ACTB promoters, used as positive controls, and for the coding region of the inactive MYOD1 gene and the Sat2 satellite repeat, used as negative controls. Figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).
Supportive validation
- Submitted by
- Novus Biologicals (provider)
- Main image
- Experimental details
- Dot Blot: Histone H2a [ac Lys5] Antibody [NBP2-59261] - To test the cross reactivity of the antibody against H2AK5ac, a Dot Blot analysis was performed with peptides containing other histone modifications and the unmodified H2AK5. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:5,000. Figure shows a high specificity of the antibody for the modification of interest.
