Anti-Optineurin antibody [RP23040250]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Optineurin Antibody [RP23040250] (AB313418)

Immunofluorescent analysis of NIH 3T3 cells fixed with 4% formaldehyde reconstituted in 1X PBS for 10 min at room temperature and permeabilized with 0.1 % Triton X-100 in PBS for 15 min at room temperature labeling endogenous Optineurin with ab313418 at 5 µg/mL dilution followed by Goat anti-Rabbit IgG (H+L) Secondary Antibody, Alexa Fluor® 488 conjugate at 1/2000 dilution. Panel a) shows representative cells that were stained for detection and localization of Optineurin protein (green), Panel b) is stained for nuclei (blue) using DAPI. Panel c) represents cytoskeletal F-actin staining using Rhodamine Phalloidin at 1/300 dilution. Panel d) is a composite image of Panels a, b and c clearly demonstrating cytoplasmic localization of Optineurin. Panel e) represents control cells without primary antibody to assess background. The images were captured at 60X magnification.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-Optineurin Antibody [RP23040250] (AB313418)

Immunofluorescent analysis of frozen mouse retina tissue fixed with 4% PFA for 20 min, permeabilized using 0.1% Triton X 100 for 10 mins and blocked for 1 hour with 2% BSA labeling Optineurin with ab313418 at 1/250 dilution followd by Goat anti-Rabbit IgG (H+L) Secondary Antibody, Alexa Fluor® 488 conjugate at 1/2000 dilution. Nuclei (blue) were stained using DAPI, Panel a) represents staining with the matched isotype control. Panel b) shows a representative mouse retinal section stained for Optineurin (green). The images were captured at 20X magnification.

• WB

Lab

Western blot - Anti-Optineurin antibody [RP23040250] (AB313418)

Western blot : Anti-Optineurin Antibody [RP23040250] ab313418 staining at 1 µg/mL, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 73 kDa in Wild-type MCF7 cell lysates with no signal observed at this size in OPTN knockout MCF7 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc Milk in TBS-0.1 % Tween^™ 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Optineurin antibody [RP23040250] (ab313418) at 1 µg/mL

Lane 1:

Wild-type MCF7 whole cell lysate at 20 µg

Lane 2:

OPTN knockout MCF7 whole cell lysate at 20 µg

Lane 3:

U-2 OS whole cell lysate at 20 µg

Lane 4:

Raji whole cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 66 kDa

Observed band size: 73 kDa

false

• WB

Supplier Data

Western blot - Anti-Optineurin Antibody [RP23040250] (AB313418)

All lanes:

Western blot - Anti-Optineurin Antibody [RP23040250] (ab313418) at 2.5 µg/mL

Lane 1:

NIH/3T3 cell lysate at 30 µg

Lane 2:

NCCIT cell lysate at 30 µg

Lane 3:

HeLa cell lysate at 30 µg

Lane 4:

Mouse brain tissue lysate at 30 µg

Lane 5:

Rat brain tissue lysate at 30 µg

Lane 6:

Mouse liver tissue lysate at 30 µg

Lane 7:

Mouse eye tissue lysate at 30 µg

Secondary

All lanes:

Goat anti-Rabbit IgG (H+L) Secondary Antibody, HRP conjugate at 1/4000 dilution

Observed band size: 66 kDa

true