Anti-Peroxiredoxin 1/PAG antibody [EPR5433]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Peroxiredoxin 1/PAG antibody [EPR5433] (AB109498)

Immunohistochemical analysis of paraffin-embedded Human thyroid cancer tissue labeling Peroxiredoxin 1/PAG with ab109498 at 1/1000 (0.155 ug/ml) dilution. The section was incubated with ab109498 for 30 mins at room temperature and followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Counterstained with Hematoxylin.

Positive staining on the human thyroid cancer, performed on a Leica Biosystems BOND® RX instrument.

Secondary antibody only control : Rabbit specific IHC polymer detection kit HRP/DAB (ab209101)

Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Peroxiredoxin 1/PAG antibody [EPR5433] (AB109498)

Immunohistochemical analysis of paraffin-embedded Mouse kidney tissue labeling Peroxiredoxin 1/PAG with ab109498 at 1/4000 (0.038 ug/ml) dilution. The section was incubated with ab109498 for 30 mins at room temperature and followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Counterstained with Hematoxylin.

Positive staining on the mouse kidney, performed on a Leica Biosystems BOND® RX instrument.

Secondary antibody only control : Rabbit specific IHC polymer detection kit HRP/DAB (ab209101)

Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins.

• WB

Supplier Data

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5433] (AB109498)

Blocking and diluting buffer and concentration : 5% NFDM/TBST

All lanes:

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5433] (ab109498) at 1/10000 dilution

Lane 1:

MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Lane 3:

NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 4:

Mouse brain lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 22 kDa

Observed band size: 22 kDa

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Exposure time: 10s

• WB

Lab

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5433] (AB109498)

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 - ab109498 observed at 23 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab109498 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. ab109498 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 [EPR5433] (ab109498)

Predicted band size: 22 kDa

false

• WB

Lab

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5433] (AB109498)

Lanes 1-3 : Merged signal (red and green). Green - ab109498 observed at 26 kDa. Red - loading control ab8245 observed at 36 kDa.

ab109498 Anti-Peroxiredoxin 1/PAG antibody [EPR5433] was shown to specifically react with Peroxiredoxin 1/PAG 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 1/PAG knockout samples were subjected to SDS-PAGE. ab109498 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 [EPR5433] (ab109498) 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,41 kDa,45 kDa,78 kDa

Observed band size: 25 kDa,26 kDa,47 kDa,49 kDa,78 kDa

false

• WB

CiteAb

Western blot - Anti-Peroxiredoxin 1/PAG antibody [EPR5433] (AB109498)

Peroxiredoxin 1/PAG western blot using anti-Peroxiredoxin 1/PAG antibody [EPR5433] ab109498. Publication image and figure legend from Ziegler, Y. S., Moresco, J. J., et al., 2016, PLoS One, PubMed 27355404.

ab109498 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 ab109498 please see the product overview.

Identification of proteins in BC and MCF-10A cell secretomes.A. A workflow was established to analyze the secretomes of BC and benign mammary epithelial cells. B. Benign (MCF-10A), ERα-positive BC (MCF-7), and TNBC (MDA-MB-231, DT22, and DT28) cells were established in 3D cultures using Matrigel extracellular matrix. Live (top row) and fixed (bottom row) cultures were imaged by phase contrast or immunofluorescence, respectively. Markers of proliferation (Ki67-red), basement membrane (laminin-green), and nuclei (DAPI-blue) were utilized. C. 3D cultures of MCF-10A cells were exposed to CM from MCF-10A, MCF-7, MDA-MB-231, DT22, or DT28 cells. Proliferation (red) and nuclei (blue) are indicated by Ki67 and DAPI staining, respectively. Scale bars indicate 25 μm. D. 3D cultures described in panel C were analyzed to determine the percent of proliferating MCF-10A cells for each treatment. An asterisk (*) indicates a p-value <0.05. E. CM from each cell line was subjected to MS and compiled results are shown. F. CM from 3D cultures of BC and MCF-10A cells were subjected to Western blot analysis using antibodies to cathepsin D (CTSD), extracellular matrix protein 1 (ECM1), peroxiredoxin 1 (PRDX1), or 14-3-3 sigma (SFN). The number of spectral counts (SCs) is indicated above each lane. To visualize SFN, the CM was concentrated before blot. MDA-MB-231 and MCF-10A cells are labeled as 231 and 10A, respectively. (Abbreviations : BC = breast cancer, TNBC = triple negative breast cancer, 3D = three dimensional, CM = conditioned medium, MS = mass spectroscopy)

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