All lanes : Anti-NDUFV2 antibody [EPR15351(B)] (ab183715) at 1/1000 dilution

Lane 1 : Wild-type HeLa cell lysate
Lane 2 : NDUFV2 knockout HeLa cell lysate
Lane 3 : A549 cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

Predicted band size: 27 kDa
Observed band size: 27 kDa

Lanes 1-3: Merged signal (red and green). Green - ab183715 observed at 27 kDa. Red - loading control ab7291 observed at 50 kDa.

ab183715 Anti-NDUFV2 antibody [EPR15351(B)] was shown to specifically react with NDUFV2 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265619 (knockout cell lysate ab258068) was used. Wild-type and NDUFV2 knockout samples were subjected to SDS-PAGE. ab183715 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.

 

Immunocytochemistry/Immunofluorescence analysis of Jurkat cells labelling NDUFV2 with ab183715 at 1/1000. Cells were fixed with 100% Methanol. ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody.

Control: PBS only.
Nuclear counter stain: DAPI.

Immunohistochemical analysis of paraffin-embedded, Human hepatocellular carcinoma tissue labeling NDUFV2 with ab183715 at a 1/500 dilution. Counter stained with hematoxylin.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Flow Cytometry analysis of Jurkat (Human T cell leukemia T lymphocyte) cells labeling NDUFV2 with purified ab183715 at 1/80 dilution (10ug/mL) (red). Cells were fixed with 80% methanol and permeabilised using 0.1% Tween-20. A Goat anti rabbit IgG (Alexa Fluor^®488) (ab150077) secondary antibody was used at 1/2000 dilution. Isotype control - Rabbit monoclonal IgG (Black) (ab172730). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

All lanes : Anti-NDUFV2 antibody [EPR15351(B)] (ab183715) at 1/1000 dilution

Lane 1 : A549 cell lysate
Lane 2 : Jurkat cell lysate
Lane 3 : HeLa cell lysate
Lane 4 : Ramos cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : goat anti-rabbit IgG, (H+L), peroxidase conjugated at 1/1000 dilution

Developed using the ECL technique.

Predicted band size: 27 kDa
Observed band size: 24 kDa why is the actual band size different from the predicted?

Anti-NDUFV2 antibody [EPR15351(B)] (ab183715) at 1/20000 dilution + Human fetal heart lysate at 20 µg

Secondary
goat anti-rabbit IgG, (H+L), peroxidase conjugated at 1/1000 dilution

Developed using the ECL technique.

Predicted band size: 27 kDa
Observed band size: 24 kDa why is the actual band size different from the predicted?

Western blot analysis on immunoprecipitation from 1) Human fetal heart lysate, or 2) TBS, labeling NDUFV2 using ab183715 at 1/50 dilution and HRP-conjugated anti-rabbit IgG preferentially detecting the non-reduced form of rabbit IgG at a 1/1500 dilution.