Anti-Peroxiredoxin 1 antibody (ab15571)

  Immunocytochemistry - Peroxiredoxin 1 antibody (ab15571)

ab15571 staining Peroxiredoxin 1 in human TSU-Pr1 cells by Immunocytochemistry.

  Immunocytochemistry/ Immunofluorescence-Peroxiredoxin 1 antibody(ab15571)

ICC/IF image of ab15571 stained HeLa cells. The cells were 100% methanol fixed (5 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 (ab15571, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 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.

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Peroxiredoxin 1 antibody (ab15571)

IHC image of ab15571 staining in human liver carcinoma formalin fixed paraffin embedded tissue section, performed on a Leica Bond^TM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab15571, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  Western blot - Anti-Peroxiredoxin 1 antibody (ab15571)

All lanes : Anti-Peroxiredoxin 1 antibody (ab15571) at 1/1000 dilution

Lane 1 : Cell line indicated at 25 µg
Lane 2 : As above
Lane 3 : As above
Lane 4 : As above
Lane 5 : As above
Lane 6 : As above
Lane 7 : As above
Lane 8 : As above
Lane 9 : As above
Lane 10 : As above
Lane 11 : As above

Secondary
HRP-conjugated Goat anti-rabbit at 1/20000 dilution

Predicted band size : 22 kDa
Observed band size : 20 kDa (why is the actual band size different from the predicted?)
Additional bands at : 25 kDa. We are unsure as to the identity of these extra bands.

Western blot analysis of Peroxiredoxin was performed by loading 25ug of various whole cell lysates onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% Milk/TBST for at least 1 hour. Membranes were probed with ab15571 overnight at 4°C on a rocking platform. Membranes were washed in TBS-0.1%Tween 20 and probed with a goat anti-rabbit-HRP secondary antibody for at least one hour. Membranes were washed and chemiluminescent detection performed.