Anti-ADAR1 抗体 [EPR7033]

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-ADAR1 antibody [EPR7033] (AB126745)

Intracellular flow cytometric analysis of permeabilized Ramos cells, staining ADAR1 (red) with ab126745. 1x10⁶ cells were collected and washed with blocking buffer. Cells were fixed with 2% paraformaldehyde, permeabilized with 1X FACS permeabilizing solution and blocked with blocking buffer for 30 minutes at room temperature. Cells were incubated with primary antibody (1/10) for 30 minutes at room temperature before a Fluorescently-conjugated secondary antibody or 30 min at room temperature. A rabbit IgG was used as a negative control (green).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ADAR1 antibody [EPR7033] (AB126745)

ab126745, at 1/50 dilution, staining ADAR1 in paraffin-embedded Human brain tissue by Immunohistochemistry.

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

• IP

Supplier Data

Immunoprecipitation - Anti-ADAR1 antibody [EPR7033] (AB126745)

ADAR1 was immunoprecipitated from 0.35 mg HeLa whole cell lysate with ab307585 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307585 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate Lane 2 : ab307585 IP in HeLa whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab307585 in HeLa whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : Lanes 1-3 : 5 seconds (left), Lanes 1-3 : 3 seconds (right). The IP experiment was performed by ab307585 using HeLa cells. On the left the IP blot was probed with ab307585 and on the right the blot was probed by another anti-ADAR1 antibody (ab126745)(1 : 1000 dilution).

Lane 2:

Immunoprecipitation - Anti-ADAR1 antibody [EPR25431-60] (<a href='/products/primary-antibodies/adar1-antibody-epr25431-60-ab307585'>ab307585</a>) at 1/30 dilution

Lane 2:

Immunoprecipitation - Anti-ADAR1 antibody [EPR7033] (ab126745) at 1/1000 dilution

All lanes:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

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Exposure time: 3s

• WB

Lab

Western blot - Anti-ADAR1 antibody [EPR7033] (AB126745)

Lane 1 : Wild-type HAP1 cell lysate (20 μg)
Lane 2 : ADAR1 knockout HAP1 cell lysate (20 μg)
Lane 3 : HepG2 cell lysate (20 μg)
Lane 4 : HeLa cell lysate (20 μg)
Lanes 1 - 4 : Merged signal (red and green). Green - ab126745 observed at 150 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab126745 was shown to recognize ADAR1 when ADAR1 knockout samples were used, along with additional cross-reactive bands. Wild-type and ADAR1 knockout samples were subjected to SDS-PAGE. ab126745 and ab8245 (loading control to GAPDH) were diluted at 1/1000 and 1/10 000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-ADAR1 antibody [EPR7033] (ab126745)

Predicted band size: 136 kDa

Observed band size: 150 kDa

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• WB

Unknown

Western blot - Anti-ADAR1 antibody [EPR7033] (AB126745)

All lanes:

Western blot - Anti-ADAR1 antibody [EPR7033] (ab126745) at 1/1000 dilution

Lane 1:

HeLa (treated with IFN-alpha) cell lysate at 10 µg

Lane 2:

HeLa cell lysate at 10 µg

Lane 3:

Ramos cell lysate at 10 µg

Lane 4:

SH-SY5Y cell lysate at 10 µg

Secondary

All lanes:

HRP labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 136 kDa

Observed band size: 150 kDa

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• WB

Lab

Western blot - Anti-ADAR1 antibody [EPR7033] (AB126745)

Lanes 1-3 : Merged signal (red and green). Green - ab126745 observed at 130 kDa. Red - loading control ab8245 observed at 36 kDa.

ab126745 Anti-ADAR1 antibody [EPR7033] was shown to specifically react with ADAR1 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266846 (knockout cell lysate ab257131) was used. Wild-type and ADAR1 knockout samples were subjected to SDS-PAGE. ab126745 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-ADAR1 antibody [EPR7033] (ab126745) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

ADAR knockout HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human ADAR (ADAR1) knockout HEK-293T cell line (<a href='/products/cell-lines/human-adar-adar1-knockout-hek-293t-cell-line-ab266846'>ab266846</a>)

Lane 3:

HeLa cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 136 kDa,19 kDa,29 kDa,35 kDa,36 kDa

Observed band size: 130 kDa,20 kDa,29 kDa,35 kDa,36 kDa

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