Western Blot analysis of cell lysates.

Lane 1: HeLa cell lysates
Lane 2: Jurkat cell lysates
Lane 3: Mouse brain
Lane 4: Rat brain

The band marked with NS is probably non-specific.

ICC/IF image of ab16731 stained Hela cells. The cells were 4% PFA 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 (ab16731, 5µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) 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.

Ab16731 staining human normal adrenal gland tissue. Staining is localised to intracellular compartment (peroxisomes).
Left panel: with primary antibody at 1 ug/ml. Right panel: isotype control.
Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffer EDTA pH 9.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification

All lanes : Anti-Catalase antibody - Peroxisome Marker (ab16731) at 1/2000 dilution

Lane 1 : 40ug supernatant of mouse liver homogenate
Lane 2 : 20ug supernatant of mouse liver homogenate
Lane 3 : 5ug supernatant of mouse liver homogenate

Secondary
All lanes : HRP conjugated donkey anti-rabbit antibody

Developed using the ECL technique.

Performed under reducing conditions.

Predicted band size: 60 kDa
Observed band size: 60 kDa


Exposure time: 1 minute


This image is courtesy of an Abreview submitted by Sandra Sobocanec on 16 March 2006.

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ab16731 at 1/200 dilution staining Catalase in human 293FT cells by Immunocytochemistry/ Immunofluorescence. Cells were fixed in formaldehyde and blocked in 5% BSA for 1 hour at 25°C. The primary antibody was used at 1/200 dilution in PBS and incubated with sample at 4°C for 12 hours. An Alexa Fluor® 488 conjugated Goat polyclonal to rabbit IgG was used undiluted as secondary.   

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ab16731 at a 1/200 dilution staining Catalase in mouse bone marrow cells by Immunocytochemistry/ Immunofluorescence, incubated for 9 hours at 4°C. Formalin fixed. Blocked with 2% BSA for 30 minutes at 20°C. Secondary used at 1/200 dilution polyclonal Goat anti-rabbit IgG conjugated to Alexa Fluor 488 (green). Nuclei stained with DAPI (blue).

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