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A protein complex that is thought to mediate chromatin assembly in DNA replication and DNA repair. Assembles histone octamers onto replicating DNA in vitro. CAF-1 performs the first step of the nucleosome assembly process, bringing newly synthesized histones H3 and H4 to replicating DNA; Histones H2A/H2B can bind to this chromatin precursor subsequent to DNA replication to complete the histone octamer. P48 can bind to histone H4 in the absence of CAF-1 P150 and P160. Binds directly to helix 1 of histone H4, a region that is not accessible when H4 is in chromatin. SUBUNIT: CAF-1 is composed of three subunits, P48, P60 AND P150. Only minor amounts of P48 are complexed with P60 AND P150 in G1 phase. P48/RBBP4 is also part of the core histone deacetylase (HDAC) complex composed of HDAC1, HDAC2, RBBP4 and RBBP7. The core complex associates with MTA2, MBD3, MTA1L1, CHD3 and CHD4 to form the nucleosome remodelling and histone deacetylation (NuRD) complex, or with SIN3, SAP18 and SAP30 to form the SIN3 HDAC complex. Interacts with the viral protein-binding domain of the retinoblastoma protein (RB1). Interacts with SUV39H1 and HDAC7 (By similarity).
Our Abpromise guarantee covers the use of ab1765 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|IHC-P||Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.|
|IP||Use a concentration of 20 µg/ml.|
|WB||1/1000. Detects a band of approximately 55 kDa (predicted molecular weight: 47.6 kDa).|
ab1765 staining RbAp48 in MDCK cell pellets by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Pellets were fixed with paraformaldehyde and blocked with 1% BSA for 3 hours at 22°C; antigen retrieval was by heat mediation in a citrate buffer, permeabilization was with Tween-20. Samples were incubated with primary antibody (1/100 in TBST) for 16 hours. A HRP-conjugated Goat anti-rabbit IgG polyclonal was used as the secondary antibody.
Rabbit polyclonal to CAF 1 p48 (ab1765)
Each lane contains 20 ug of HeLa whole cell lysate.
Lane 1: ab1765 1/500
Lane 2: ab1765 1/1000
Secondary antibody - Goat anti-Rabbit (HRP) (ab6721) at 1/2000.
ab1765 used in Immunoprecipitation of HeLa whole cell lysate.
Whole cell extract (containing 100µg of soluble protein) was incubated with 20µg of antibody overnight. Protein A-Agarose was added to the immunocomplexes and incubated at room temperature for three hours. Immunocomplexes were eluted and then resolved using 7.5% SDS-PAGE under reducing conditions. For the detection of HDAC1, 2 and 3 a non-Abcam mouse monoclonal antibody was used at a dilution of 1/800. Results support the understanding that HDAC1 and 2 are in the same complex together with p48. Note that HDAC1 does not coprecipitate HDAC2, probably displaced by the antibody during the experiment.
Lane 1: Input
Lane 2: HDAC1 (ab1769)
Lane 3: p48 (ab1765)
Lane 4: No antibody control
This image is courtesy of an Abreview by Hugh Spotswood submitted on 9 February 2006.
ab1765 staining cultured human HeLa cells by ICC/IF. Cells were PFA fixed and permeabilized in 0.5% Triton X100 prior to blocking in 5% BSA for 1 hour at 20°C. The primary antibody was diluted 1/50 and incubated with the sample for 1 hour at 20°C. A Cy3® conjugated donkey anti-rabbit antibody was used as the secondary.