Anti-Myoferlin antibody [EPR18887]

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Myoferlin antibody [EPR18887] (AB178386)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling Myoferlin with ab178386 at 1/400 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor^® 488) at 1/2000 dilution was used as the secondary antibody.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Myoferlin antibody [EPR18887] (AB178386)

Immunofluorescent analysis of 100% methanol-fixed HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling Myoferlin with ab178386 at 1/100 dilution followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on HeLa cells. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (Alexa Fluor^® 594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

The negative controls are as follows :
-ve control 1 : ab178386 at 1/100 dilution followed by ab150120 (Alexa Fluor^® 594 Goat anti-Mouse secondary) at 1/1000 dilution.

-ve control 2 : ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor^® 488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Myoferlin antibody [EPR18887] (AB178386)

Immunofluorescent analysis of 100% methanol-fixed C2C12 (mouse myoblast cell line) cells labeling Myoferlin with ab178386 at 1/100 dilution followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on C2C12 cells. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (Alexa Fluor^® 594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

The negative controls are as follows :
-ve control 1 : ab178386 at 1/100 dilution followed by ab150120 (Alexa Fluor^® 594 Goat anti-Mouse secondary) at 1/1000 dilution.

-ve control 2 : ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor^® 488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

• WB

Supplier Data

Western blot - Anti-Myoferlin antibody [EPR18887] (AB178386)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure times : Lane 1/5 : 10 seconds; Lane 2 : 15 seconds; Lane 3/4 : 3 minutes.

The molecular weight observed is consistent with what has been described in the literature (PMID : 22135466, PMID : 23499551).

Lanes 1 - 2:

Western blot - Anti-Myoferlin antibody [EPR18887] (ab178386) at 1/20000 dilution

Lanes 3 - 5:

Western blot - Anti-Myoferlin antibody [EPR18887] (ab178386) at 1/2000 dilution

Lane 1:

MCF7 (human breast adenocarcinoma cell line) whole cell lysate at 10 µg

Lane 2:

MDA-MB-231 (human breast adenocarcinoma cell line) whole cell lysate at 10 µg

Lane 3:

Human fetal kidney lysate at 10 µg

Lane 4:

Human fetal heart lysate at 10 µg

Lane 5:

HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 10 µg

Secondary

Lanes 1 - 3:

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 1/100000 dilution

Lanes 4 - 5:

Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/100000 dilution

Predicted band size: 235 kDa

Observed band size: 180 kDa,250 kDa

false

• WB

Lab

Western blot - Anti-Myoferlin antibody [EPR18887] (AB178386)

Lanes 1-4 : Merged signal (red and green). Green - ab178386 observed at 250,180 kDa. Red - loading control ab7291 observed at 50 kDa.

ab178386 Anti-Myoferlin antibody [EPR18887] was shown to specifically react with Myoferlin in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265782 (knockout cell lysate ab257547) was used. Wild-type and Myoferlin knockout samples were subjected to SDS-PAGE. ab178386 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) 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-Myoferlin antibody [EPR18887] (ab178386) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

MYOF knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human MYOF (Myoferlin) knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-myof-myoferlin-knockout-hela-cell-line-ab265782'>ab265782</a>)

Lane 3:

MDA-MB-231 cell lysate at 20 µg

Lane 4:

SK-BR-3 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: 235 kDa

Observed band size: 180 kDa,250 kDa

false

• WB

Supplier Data

Western blot - Anti-Myoferlin antibody [EPR18887] (AB178386)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 3 minutes; Lane 2/4 : 10 seconds; Lane 3 : 3 seconds.

All lanes:

Western blot - Anti-Myoferlin antibody [EPR18887] (ab178386) at 1/2000 dilution

Lane 1:

Rat kidney lysate at 10 µg

Lane 2:

C6 (rat glial tumor cell line) whole cell lysate at 10 µg

Lane 3:

NIH/3T3 (mouse embryonic fibroblast cell line) whole cell lysate at 10 µg

Lane 4:

C2C12 (mouse myoblast cell line) whole cell lysate at 10 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 1/100000 dilution

Predicted band size: 235 kDa

Observed band size: 180 kDa,250 kDa

false