Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

This data was developed using the same antibody clone in a different buffer formulation (ab109506).

Peroxiredoxin 1/PAG (PRDX1) staining observed in wild-type HEK293T cells and PRDX1 knockout HEK293T cells (ab266842). The cells were fixed with 100% methanol then permeabilized with 0.1% Triton X-100. The cells were then incubated with ab109506 at 1/50 dilution and followed by secondary antibody ab150077 AlexaFluor®488 Goat anti-Rabbit secondary at 1/1000 dilution (shown in green). ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) was used to counterstain at 1/200 dilution (shown in red). Nuclear DNA was labelled in blue with DAPI.

Confocal image showing cytoplasmic staining in wild-type HEK-293Tcell line, and no staining in PRDX1 knockout HEK-293T cell line.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

This data was developed using the same antibody in a different buffer formulation (ab109506).

Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling Peroxiredoxin 1/PAG with unpurified ab109506 at 1/20 dilution (10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor^®488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

This data was developed using the same antibody in a different buffer formulation (ab109506).

ICC/IF image of ab109506 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab109506, 1/100 dilution) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43μM.

• ICC/IF

Collaborator

Immunocytochemistry/ Immunofluorescence - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

This data was developed using the same antibody clone in a different buffer formulation (ab109506). ab109506 was shown to react with PRDX1 in wild-type U-2 OS cells in Immunocytochemistry with loss of signal observed in a PRDX1 knockout cell line. Wild-type and knockout cells were mixed and pelleted at a 1 : 1 ratio on coverslips. The cells were fixed with then permeabilized with and then blocked with 1/10000. The cells were then incubated with ab109506 at 1/250. 1/5000 would be better dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a secondary antibody to (Alexa Fluor^® 555) at 0.5 μg/ml. Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880). These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

• IP

Supplier Data

Immunoprecipitation - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

This data was developed using the same antibody in a different buffer formulation (ab109506).

Immunoprecipitation of PRDX1 in U-2 OS cells. Lysates were prepared and immunoprecipitation was performed using 1.0 μg of ab109506 pre-coupled to prot.A-Sepharose beads. Samples were washed and processed for western blot with Peroxiredoxin 1 Antibody at 1/10000. SM=10% starting material; UB=10% unbound fraction; IP=immunoprecipitate. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Immunoprecipitation - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (ab211538)

Predicted band size: 22 kDa

false

• WB

Supplier Data

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

This data was developed using the same antibody in a different buffer formulation (ab109506).

Lane 1 : Wild-type HAP1 cell lysate (20 μg)
Lane 2 : Peroxiredoxin 1/PAG knockout HAP1 cell lysate (20 μg)
Lane 3 : A431 cell lysate (20 μg)
Lane 4 : Jurkat cell lysate (20 μg)
Lanes 1 - 4 : Merged signal (red and green). Green - ab109506 observed at 23 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab109506 was shown to specifically react with Peroxiredoxin 1/PAG when Peroxiredoxin 1/PAG knockout samples were used. Wild-type and Peroxiredoxin 1/PAG knockout samples were subjected to SDS-PAGE. ab109506 and ab8245 (loading control to GAPDH) were both diluted 1/10 000 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-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (ab211538)

Predicted band size: 22 kDa

false

• WB

Collaborator

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

This data was developed using the same antibody in a different buffer formulation (ab109506).

ab109506 was shown to react with PRDX1 in wild-type U-2 OS cells in Western blot with loss of signal observed in a PRDX1 knockout cell line. Wild-type U-2 OS and PRDX1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab109506 overnight at 4 °C at a 1/5000 dilution. Blots were incubated with goat anti-rabbit HRP secondary antibodies at 0.2ug/mL before imaging. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] (<a href='/en-us/products/primary-antibodies/peroxiredoxin-1-pag-antibody-epr5434-ab109506'>ab109506</a>) at 1/5000 dilution

Lane 1:

Wild-type U-2 OS cell lysate at 20 µg

Lane 2:

PRDX1 knockout U-2 OS cell lysate at 20 µg

Predicted band size: 22 kDa

false

• WB

Unknown

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

This data was developed using the same antibody clone in a different buffer formulation (ab109506).

Lanes 1-3 : Merged signal (red and green). Green - ab109506 observed at 26 kDa. Red - loading control ab8245 observed at 36 kDa.

ab109506 Anti-Peroxiredoxin 1/PAG antibody [EPR5434] was shown to specifically react with Peroxiredoxin in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266842 (knockout cell lysate ab257040) was used. Wild-type and Peroxiredoxin knockout samples were subjected to SDS-PAGE. ab109506 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-Peroxiredoxin 1/PAG antibody [EPR5434] (<a href='/en-us/products/primary-antibodies/peroxiredoxin-1-pag-antibody-epr5434-ab109506'>ab109506</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

PRDX1 knockout HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human PRDX1 (Peroxiredoxin 1/PAG) knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-prdx1-peroxiredoxin-1-pag-knockout-hek-293t-cell-line-ab266842'>ab266842</a>)

Lane 3:

Jurkat cell lysate at 20 µg

Secondary

All lanes:

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

Predicted band size: 22 kDa

Observed band size: 26 kDa

false

• WB

Supplier Data

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (AB211538)

All lanes:

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5434] - BSA and Azide free (ab211538) at 1/10000 dilution

Lane 1:

293T cell lysate at 10 µg

Lane 2:

K562 cell lysate at 10 µg

Lane 3:

U87-MG cell lysate at 10 µg

Secondary

All lanes:

Standard HRP labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 22 kDa

false