Anti-KAT1 / HAT1 antibody [RP23040271] - ChIP Grade

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-KAT1 / HAT1 antibody [RP23040271] - ChIP Grade (AB308104)

For immunofluorescence analysis, HeLa cells were fixed and permeabilized for detection of endogenous HAT1 using ab308104 at 1/100 dilution and labeled with Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor 488 conjugate at 1/2000 dilution. Panel a) shows representative cells that were stained for detection and localization of HAT1 protein (green) Panel b) is stained for nuclei (blue) using DAPI Panel c) represents cytoskeletal F-actin staining using Rhodamine Phalloidin at 1/300 dilution Panel d) is a composite image of Panels a, b and c clearly demonstrating nuclear localization of HAT1 Panel e) represents control cells with no primary antibody to assess background The images were captured at 60X magnification

• ChIP

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ChIP - Anti-KAT1 / HAT1 antibody [RP23040271] - ChIP Grade (AB308104)

Chromatin Immunoprecipitation (ChIP) assay of endogenous HAT1 protein using ab308104 : ChIP was performed using 5 µg ab308104 on sheared chromatin from 2 million HeLa cells. Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by qPCR using primers binding to transcriptional start site and gene body (+2Kb) of GAPDH, RPL30 transcriptional start site, CDKN1A intron 1 and SAT2 satellite repeats. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

• WB

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Western blot - Anti-KAT1 / HAT1 antibody [RP23040271] - ChIP Grade (AB308104)

Western blot was performed using ab308104 and a ~49.5 kDa band corresponding to HAT1 was observed across cell lines tested. Nuclear enriched extracts (30 µg lysate) of MCF7 (Lane 1), HCT 116 (Lane 2), LNCaP (Lane 3), U-2 OS (Lane 4), HeLa (Lane 5) and Jurkat (Lane 6) were electrophoresed using a 10% Bis-Tris gel, electrophoresis system and pre-stained protein standard. Resolved proteins were then transferred onto a nitrocellulose membrane. The blot was probed with ab308104 at 1/1000 dilution and detected by chemiluminescence using a Goat anti-Rabbit IgG (H+L) Secondary Antibody, HRP conjugate at 1/4000 dilution. Chemiluminescent detection was performed (ECL).

All lanes:

Western blot - Anti-KAT1 / HAT1 antibody [RP23040271] - ChIP Grade (ab308104) at 1/1000 dilution

Lane 1:

MCF7 cell lysate at 30 µg

Lane 2:

HCT 116 cell lysate at 30 µg

Lane 3:

LNCaP cell lysate at 30 µg

Lane 4:

U-2 OS cell lysate at 30 µg

Lane 5:

HeLa cell lysate at 30 µg

Lane 6:

Jurkat cell lysate at 30 µg

Secondary

All lanes:

HRP conjugated Goat anti-Rabbit IgG (H+L) Secondary Antibody at 1/4000 dilution

Observed band size: 49.5 kDa

true

• WB

Supplier Data

Western blot - Anti-KAT1 / HAT1 antibody [RP23040271] - ChIP Grade (AB308104)

Knockdown of HAT1 was achieved by transfecting HepG2 cells with HAT1 specific siRNA. Western blot analysis was performed using whole cell extracts from HAT1 knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with ab308104 and a Goat anti-Rabbit IgG (H+L) Secondary Antibody, HRP conjugate at 1/4000 dilution. Loss of signal upon siRNA mediated knockdown confirms that antibody is specific to HAT1.

All lanes:

Western blot - Anti-KAT1 / HAT1 antibody [RP23040271] - ChIP Grade (ab308104) at 1/1000 dilution

Lane 1:

untransfected HepG2 cell lysate

Lane 2:

non-specific siRNA transfected HepG2 cell lysate

Lane 3:

HAT1 siRNA transfected HepG2 cell lysate

Secondary

All lanes:

HRP conjugated Goat anti-Rabbit IgG (H+L) Secondary Antibody at 1/4000 dilution

false