Anti-TATA binding蛋白TBP抗体-核Loading Control and ChIP Grade

• IHC-P

AbReview43883****

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (AB63766)

ab63766 staining TAT binding protein TBP in human infantile fibromatosis tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% FBS/BSA for 3 hours at room temperature; antigen retrieval was by heat mediation in Tris pH9. Samples were incubated with primary antibody (1/100 in TBS + 1% BSA + 1% FBS) for 16 hours. An undiluted HRP-conjugated goat anti-rabbit IgG polyclonal was used as the secondary antibody.

This image is courtesy of an anonymous Abreview

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (AB63766)

IHC image of TATA binding protein TBP staining in Human breast ductal carcinoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab63766, 10μg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

• ChIP

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ChIP - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (AB63766)

Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 5µg of ab63766 (blue), and 20µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach for active loci and Taqman approach for inactive loci). Primers and probes are located in the first kb of the transcribed region.

• IP

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Immunoprecipitation - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (AB63766)

TATA binding protein TBP was immunoprecipitated using 0.5mg HepG2 whole cell extract, 5µg of Rabbit polyclonal to TATA binding protein TBP and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
The antibody was incubated under agitation with Protein G beads for 10min, HepG2 whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70^oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab63766.
Secondary : Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
Band : 45kDa : TATA binding protein TBP.

All lanes:

Immunoprecipitation - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (ab63766)

Predicted band size: 38 kDa

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• WB

Project5081****

Western blot - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (AB63766)

All lanes:

Western blot - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (ab63766) at 1 µg/mL

Lane 1:

HeLa Whole Cell Lysate at 10 µg

Lane 2:

HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate at 10 µg

Secondary

All lanes:

Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

Predicted band size: 38 kDa

Observed band size: 45 kDa,60 kDa

false

• WB

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Western blot - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (AB63766)

All lanes:

Western blot - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (ab63766) at 1 µg/mL

Lane 1:

Testis (Mouse) Tissue Lysate at 10 µg

Lane 2:

NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate at 10 µg

Lane 3:

Testis (Rat) Tissue Lysate at 10 µg

Lane 4:

Western blot - Hep G2 nuclear extract lysate (<a href='/products/tissue-lysates/hep-g2-nuclear-extract-lysate-ab14660'>ab14660</a>) at 10 µg

Secondary

All lanes:

Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

Predicted band size: 38 kDa

Observed band size: 38 kDa,45 kDa,55 kDa

true

Exposure time: 3min

• WB

Unknown

Western blot - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (AB63766)

All lanes:

Western blot - Anti-TATA binding protein TBP antibody - Nuclear Loading Control and ChIP Grade (ab63766) at 1 µg/mL

Lane 1:

Western blot - Recombinant Human TATA binding protein TBP (<a href='/products/proteins-peptides/recombinant-human-tata-binding-protein-tbp-ab81897'>ab81897</a>) at 0.1 µg

Lane 2:

Western blot - Recombinant Human TATA binding protein TBP (<a href='/products/proteins-peptides/recombinant-human-tata-binding-protein-tbp-ab81897'>ab81897</a>) at 0.01 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-preadsorbed-ab97080'>ab97080</a>) at 1/5000 dilution

Predicted band size: 38 kDa

true

Exposure time: 10s