Immunofluorescence analysis of Histone H3.3 was performed using 70% confluent log phase NIH/3T3 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Histone H3.3 Rabbit Polyclonal Antibody (PA517697) at 1:250 dilution in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (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 (S36938). 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 shows the no primary antibody control. The images were captured at 60X magnification.
|Tested species reactivity||Bovine, Fruit fly, Human, Mouse, Non-human primate, Pig, Rat, Zebrafish|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to the carboxy-terminal sequence of human histone H3|
|Purification||Antigen affinity chromatography|
|Storage buffer||0.01M HEPES, pH 7.5, with 0.15M NaCl, 100µg/ml BSA, 50% glycerol|
|Tested Applications||Dilution *|
|ChIP assay (ChIP)||1.5 µg/10^6 cells|
|Flow Cytometry (Flow)||1:20|
|Western Blot (WB)||1:1,000 - 1:5,000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
It is not recommended to aliquot this antibody.
This antibody is not cross-reactive with other histones.
Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. This structure consists of approximately 146 bp of DNA wrapped around a nucleosome, an octamer composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin fiber is further compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures. This gene is intronless and encodes a member of the histone H3 family. Transcripts from this gene lack polyA tails; instead, they contain a palindromic termination element. This gene is found in the large histone gene cluster on chromosome 6p22-p21.3.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.