Anti-SPT5 antibody [EPR5145(2)]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SPT5 antibody [EPR5145(2)] (AB126592)

ab126592 at 1/100 dilution staining SPT5 in paraffin-embedded Human colon tissue by immunohistochemistry.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-SPT5 antibody [EPR5145(2)] (AB126592)

ab126592 at 1/100 dilution staining SPT5 in HeLa cells by immunofluorescence.

• WB

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Western blot - Anti-SPT5 antibody [EPR5145(2)] (AB126592)

All lanes:

Western blot - Anti-SPT5 antibody [EPR5145(2)] (ab126592) at 1/1000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

HepG2 cell lysate at 10 µg

Lane 3:

RAW 264.7 cell lysate at 10 µg

Lane 4:

PC-12 cell lysate at 10 µg

Lane 5:

NIH/3T3 cell lysate at 10 µg

Lane 6:

Jurkat cell lysate at 10 µg

Lane 7:

293T cell lysate at 10 µg

Secondary

All lanes:

Goat anti-rabbit HRP conjugated at 1/2000 dilution

Predicted band size: 121 kDa

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• OI-RD Scanning

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OI-RD Scanning - Anti-SPT5 antibody [EPR5145(2)] (AB126592)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.

• WB

CiteAb

Western blot - Anti-SPT5 antibody [EPR5145(2)] (AB126592)

SPT5 western blot using anti-SPT5 antibody [EPR5145(2)] ab126592. Publication image and figure legend from Narain, A., Bhandare, P., et al., 2021, Mol Cell, PubMed 34233157.

ab126592 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab126592 please see the product overview.

Auxin-mediated depletion of SPT6 is rapid and specific(A) Schematic of the knockin strategy for AID-tagged SPT6. Shown are components of the knockin cassette. Positions of primers for genomic PCR are marked by arrows.(B) Agarose gel of PCR from U2OSSPT6-AID knockin clones. wt, wild-type; het, heterozygous; C1 and C2, homozygous clones.(C) Immunoblot of SPT6 in U2OSSPT6-AID-C2 cells treated with auxin for the indicated times. Vinculin, loading control.(D) Full-membrane immunoblots. U2OSSPT6-AID-C1 and U2OSSPT6-AID-C2 cells were treated with auxin, and the migration of SPT6 was compared with that in wild-type U2OS cells using antibodies for SPT6 and V5.(E) Immunoblot of SPT6 in U2OSSPT6-AID cells treated with auxin and then incubated in fresh medium for the indicated times. Vinculin, loading control.(F) Browser tracks of NPM1 gene showing SPT6 ChIP-Rx reads from U2OSSPT6-AID-C1.(G) Metagene plots from SPT6 ChIP-Rx and total-RNAPII. Input-normalized reads averaged over all expressed genes are shown. p values (two-sided Wilcoxon test) for the difference (auxin/control), calculated from the density values of individual genes at each genomic location, are shown in a heatmap.(H) Heatmaps showing Z scores calculated from input-normalized SPT6 ChIP-Rx reads in the presence (right) or absence (left) of auxin for 7,562 expressed genes (>5 kb), sorted by length.(I) Immunoblot of SPT6-interacting proteins. U2OSSPT6-AID-C1 cells were treated with auxin, and SPT6-associated proteins were analyzed on separate membranes. Tubulin, loading control.See also Figure S1.

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