Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free

Novus Biologicals | Catalog # NBP1-30141

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Key Product Details

Species Reactivity

Validated:

Human, Mouse, Rat, Porcine, C. elegans, Drosophila, Invertebrate, Mammal, Yeast

Cited:

Human, Mouse, Porcine, Invertebrate, Yeast

Applications

Validated:

Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, ELISA, Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, Chromatin Immunoprecipitation (ChIP)

Cited:

Immunohistochemistry-Paraffin, Western Blot, Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, Chemotaxis, Proximity Ligation Assay, IF/IHC

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG1 kappa Clone # 6F12-H4

Format

BSA Free
View all Histone H3 Primary Antibodies »

Product Specifications

Immunogen

This Histone H3 [Trimethyl Lys9] antibody (6F12-H4) was raised against a synthetic peptide made to an N-terminal region of Histone H3 (between amino acids 1-50). [UniProt# P84243]

Reactivity Notes

Predicted to react with most mammalian species. Invertebrate / Blattella germanica (German cockroach) reactivity reported in scientific literature (PMID: 23872316). Porcine reactivity reported in scientific literature (PMID: 25736622)

Modification

Trimethyl Lys9

Localization

Nucleus

Clonality

Monoclonal

Host

Mouse

Isotype

IgG1 kappa

Theoretical MW

15 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free

Western Blot: Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) [NBP1-30141] - Whole cell protein from human HeLa, A431, mouse Neuro2A and rat PC12 cells was separated on a 4-20% gel by SDS-PAGE, transferred to 0.2 um PVDF membrane and blocked in 5% non-fat milk in TBST. The membrane was probed with 1.0 ug/ml anti-Histone h3 in block buffer and detected with an anti-mouse HRP secondary antibody using chemiluminescence. Observed molecular weight is ~15 kDa.
Immunohistochemistry-Paraffin: Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) [NBP1-30141] - Histone 3 was detected in immersion fixed paraffin-embedded sections of human colon cancer using anti-human mouse monoclonal antibody (Catalog # NBP1-30141) at 1:1000 dilution overnight at 4C. Tissue was stained using the VisuCyte anti-mouse HRP polymer detection reagent (Catalog # VC001) with DAB chromogen (brown) and counterstained with hematoxylin (blue). Images may not be copied, printed or otherwise disseminated without express written permission of Novus Biologicals a bio-techne brand.
Immunocytochemistry/Immunofluorescence: Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) [NBP1-30141] - HeLa cells were fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X PBS + 0.5% Triton X-100. The cells were incubated with anti-Histone H3 [Trymethyl Lys9] (6F12-H4) at 2 ug/ml overnight at 4C and detected with an anti-mouse DyLight 488 (green) at a 1:500 dilution. Nuclei were counterstained with DAPI (blue). Cells were imaged using a 40X objective.
Western Blot: Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) [NBP1-30141] - Analysis of Histone H3 (NBP1-30141) using HeLa nuclear lysate [NB800-PC9]. Observed molecular weight ~15 kDa.
Immunocytochemistry/Immunofluorescence: Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) [NBP1-30141] - Analysis of A549 cells (fixed with methanol) using Histone H3 [Trimethyl Lys9] antibody. Image from verified customer review.
Chromatin Immunoprecipitation: Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) [NBP1-30141] - Used to perform ChIP on HeLa cells.
Western Blot: Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) [NBP1-30141] - H3K9Me3 analysis of MCF7 and HCT116. Theorectical molecular weight is ~15 kDa. Image from verified customer review.
Histone-H3-Trimethyl-Lys9-Antibody-6F12-H4-Western-Blot-NBP1-30141-img0013.jpg
Western Blot: Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) [NBP1-30141] - Analysis of Histone H3 K9-me3 in HeLa histone lysates. Observed molecular weight is ~15 kDa.
Histone H3 [Trimethyl Lys9] Antibody (6F12-H4)

Histone H3 [Trimethyl Lys9] (6F12-H4) in MCF7 Human Cell Line.

Histone H3 [Trimethyl Lys9] (6F12-H4) was detected in immersion fixed MCF7 human breast cancer cell line using Mouse anti- Histone H3 [Trimethyl Lys9] (6F12-H4) Protein G Purified Monoclonal Antibody conjugated to DyLight 550 (Catalog # NBP1-30141R) (red) at 5 µg/mL overnight at 4C. Cells were counterstained with DAPI (blue). Cells were imaged using a 100X objective and digitally deconvolved.

Applications for Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free

Application
Recommended Usage

Chromatin Immunoprecipitation (ChIP)

1:10-1:500

ELISA

1:100-1:2000

Immunocytochemistry/ Immunofluorescence

1:200

Immunohistochemistry

1:1000. Use reported in scientific literature (PMID 23872316)

Immunohistochemistry-Paraffin

1:1000

Immunoprecipitation

1:10-1:500. Use reported in scientific literature (PMID 28270554)

Western Blot

1:2000
Application Notes
In Western blot, a band is seen at ~15 kDa.

Reviewed Applications

Read 2 reviews rated 5 using NBP1-30141 in the following applications:

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

Tris-Glycine and 0.15M NaCl

Format

BSA Free

Preservative

0.05% Sodium Azide

Concentration

1 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: Histone H3

Chromatin is an arrangement of DNA and proteins which form chromosomes. Chromatin is formed from nucleosomes, which are comprised of a set of four histone proteins (H2A, H2B, H3, H4) wrapped with DNA. Chromatin is very dynamic, where post-translational modifications can regulate DNA to be copied, transcribed, or repaired.

Histones are nuclear proteins responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Changes in chromatin structure play a large role in the regulation of transcription. The chromatin fibers are compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures.

Common histone modifications include methylation of lysine and arginine, acetylation of lysine, phosphorylation of threonine and serine, and sumoylation, biotinylation, and ubiquitylation of lysine. Posttranslational modifications such as acetylation of core histones regulates gene expression, thus altering protein function and regulation (1). Histone H3 is primarily acetylated at lysines 9, 14, 18, and 23 and have a theoretical molecular weight of 15 kDa. Acetylation at lysine 9 and 14 appears to control histone deposition, chromatin assembly and active transcription. Methylation of arginine residues within histone H3 has also been linked to transcription regulation. Histone H3 has been linked to various types of cancer as a biomarker through the aberrant expression of histone deacetylase (HDAC) enzymes and changes to chromatins (2-4).

References

1. Zhang, Y. X., Akumuo, R. C., Espana, R. A., Yan, C. X., Gao, W. J., & Li, Y. C. (2018). The histone demethylase KDM6B in the medial prefrontal cortex epigenetically regulates cocaine reward memory. Neuropharmacology, 141, 113-125. doi:10.1016/j.neuropharm.2018.08.030

2. Nandakumar, V., Hansen, N., Glenn, H. L., Han, J. H., Helland, S., Hernandez, K,...Meldrum, D. R. (2016). Vorinostat differentially alters 3D nuclear structure of cancer and non-cancerous esophageal cells. Sci Rep, 6, 30593. doi:10.1038/srep30593

3. Zhou, M., Li, Y., Lin, S., Chen, Y., Qian, Y., Zhao, Z., & Fan, H. (2019). H3K9me3, H3K36me3, and H4K20me3 Expression Correlates with Patient Outcome in Esophageal Squamous Cell Carcinoma as Epigenetic Markers. Dig Dis Sci, 64(8), 2147-2157. doi:10.1007/s10620-019-05529-2

4. Li, Y., Guo, D., Sun, R., Chen, P., Qian, Q., & Fan, H. (2019). Methylation Patterns of Lys9 and Lys27 on Histone H3 Correlate with Patient Outcome in Gastric Cancer. Dig Dis Sci, 64(2), 439-446. doi:10.1007/s10620-018-5341-8

Alternate Names

H3F3A, H3K9Me3

Entrez Gene IDs

3020 (Human); 3021 (Human); 287352 (Rat)

Gene Symbol

H3-3A

UniProt

P84243 (Human), P84244 (Mouse)

Additional Histone H3 Products

Product Documents for Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free

Certificate of Analysis

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Product Specific Notices for Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Related Research Areas

Epigenetics
Interferons
Notch Pathway

Citations for Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free

Customer Reviews for Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free (2)

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Customer Images

Showing  1 - 2 of 2 reviews Showing All
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  • Histone H3 [Trimethyl Lys9] Antibody (6F12-H4)
    Name: Anonymous
    Application: Immunocytochemistry
    Sample Tested: A549 cells
    Species: Human
    Verified Customer | Posted 03/11/2022
    Left: NBP1-30141; Right: DAPI-stained nuclei A549 cells, fixed with methanol.
    Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free NBP1-30141
  • Name: Anonymous
    Application: Western Blot
    Sample Tested: H3K9(me)3 immunoblot in MCF7 and HCT116
    Species: Human
    Verified Customer | Posted 09/09/2014
    H3K9(me)3 immunoblot in MCF7 and HCT116
    Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free NBP1-30141

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Protocols

View specific protocols for Histone H3 [Trimethyl Lys9] Antibody (6F12-H4) - BSA Free (NBP1-30141):



Western Blot Protocol
1. Perform SDS-PAGE (4-12% MOPS) on samples to be analyzed, loading 40 ug of total protein per lane.
2. Transfer proteins to Nitrocellulose according to the instructions provided by the manufacturer of the transfer
apparatus.
3. Rinse membrane with dH2O and then stain the blot using Ponceau S for 1-2 minutes to access the transfer of
proteins onto the nitrocellulose membrane. Rinse the blot in water to remove excess stain and mark the lane locations
and locations of molecular weight markers using a pencil.
4. Rinse the blot in TBS for approximately 5 minutes.
5. Block the membrane using 5% BSA in TBS + Tween, 1 hour at RT.
6. Rinse the membrane in dH2O and then wash the membrane in wash buffer [TBS + 0.1% Tween] 3 times for 10
minutes each.
7. Dilute the rabbit anti-Histone H4 [K20-me1]primary antibody (NBP1-30091) in blocking buffer and incubate 1 hour at room
temperature.
8. Rinse the membrane in dH2O and then wash the membrane in wash buffer [TBS + 0.1% Tween] 3 times for 10
minutes each.
9. Apply the diluted rabbit-IgG HRP-conjugated secondary antibody in blocking buffer (as per manufacturers
instructions) and incubate 1 hour at room temperature.
10. Wash the blot in wash buffer [TBS + 0.1% Tween] 3 times for 10 minutes each (this step can be repeated as
required to reduce background).
11. Apply the detection reagent of choice in accordance with the manufacturers instructions (Pierce ECL).
Note: Tween-20 can be added to the blocking or antibody dilution buffer at a final concentration of 0.05-0.2%, provided
it does not interfere with antibody-antigen binding.

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