Anti-NDUFS3 antibody [EPR12782] - C-terminal

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

Immunohistochemcal analysis of paraffin-embedded Human kidney tissue labeling NDUFS3 with ab177471 at 1/50 dilution.

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

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

Immunofluorescence analysis of 293 cells labeling NDUFS3 with ab177471 at 1/50 dilution (green). DAPI nuclear staining (blue).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

Immunohistochemcal analysis of paraffin-embedded Human heart tissue labeling NDUFS3 with ab177471 at 1/50 dilution.

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

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

Intracellular Flow Cytometry analysis of HEK293 (Human embryonic kidney epithelial cell) cells labeling NDUFS3 (red) with purified ab177471 at a 1/200 dilution (10ug/mL). Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A goat anti rabbit IgG (Alexa Fluor^®488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (Black) (ab172730). Blue (unlabeled control) - Cell without incubation with primary antibody and secondary antibody (Blue).

• WB

Lab

Western blot - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

Lanes 1- 4 : Merged signal (red and green). Green - ab177471 observed at 30 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.

ab177471 was shown to react with NDUFS3 in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266419 (knockout cell lysate ab257556) was used. Wild-type HEK-293T and NDUFS3 knockout HEK-293T cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab177471 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) overnight at 4°C at a 1 in 1000 dilution and a 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-NDUFS3 antibody [EPR12782] - C-terminal (ab177471) at 1/1000 dilution

Lane 1:

Wild-type HEK-293T cell lysate at 20 µg

Lane 2:

NDUFS3 knockout HEK-293T cell lysate at 20 µg

Lane 2:

Western blot - Human NDUFS3 knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-ndufs3-knockout-hek-293t-cell-line-ab266419'>ab266419</a>)

Lane 3:

HepG2 cell lysate at 20 µg

Lane 4:

HL60 cell lysate at 20 µg

Predicted band size: 30 kDa

Observed band size: 30 kDa

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• WB

AbReview51458****

Western blot - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

All lanes:

Western blot - Anti-NDUFS3 antibody [EPR12782] - C-terminal (ab177471) at 1/1000 dilution

Lanes 1 - 2:

Whole cell lysates from NDUFS3 KO cells. at 50 µg

Lanes 3 - 4:

Whole cell lysates from NDUFS3 WT cells. at 50 µg

Lanes 5 - 6:

Isolated mitochondria from NDUFS3 KO Cells. at 50 µg

Lanes 7 - 8:

Isolated mitochondria from NDUFS3 WT Cells. at 50 µg

Secondary

All lanes:

Goat anti-Rabbit polyclonal HRP conjugate. at 1/5000 dilution

Predicted band size: 30 kDa

true

Exposure time: 1min

Image is courtesy of an anonymous Abreview.

• IP

Supplier Data

Immunoprecipitation - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

Western blot analysis on immunoprecipitation pellet from (1) human fetal heart lysate or (2) 1XPBS (negative control) using ab177471at 1/10 dilution, and HRP-conjugated anti-rabbit IgG preferentially detecting the non-reduced form of rabbit IgG.

All lanes:

Immunoprecipitation - Anti-NDUFS3 antibody [EPR12782] - C-terminal (ab177471)

Predicted band size: 30 kDa

true

• WB

Lab

Western blot - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

Lanes 1 - 2 : Merged signal (red and green). Green - ab177471 observed at 27 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55kDa.

ab177471 was shown to react with NDUFS3 in HEK-293T wild-type cells in western blot with loss of signal observed in NDUFS3 knockout cell line ab266419 (NDUFS3 knockout cell lysate ab257556). Wild-type and NDUFS3 knockout HEK-293T cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab177471 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-NDUFS3 antibody [EPR12782] - C-terminal (ab177471) at 1/1000 dilution

Lane 1:

Wild-type HEK-293T cell lysate at 20 µg

Lane 2:

NDUFS3 knockout HEK-293T cell lysate at 20 µg

Lane 2:

Western blot - Human NDUFS3 knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-ndufs3-knockout-hek-293t-cell-line-ab266419'>ab266419</a>)

Predicted band size: 30 kDa

Observed band size: 27 kDa

false

• WB

Supplier Data

Western blot - Anti-NDUFS3 antibody [EPR12782] - C-terminal (AB177471)

All lanes:

Western blot - Anti-NDUFS3 antibody [EPR12782] - C-terminal (ab177471) at 1/1000 dilution

Lane 1:

HepG2 lysate at 10 µg

Lane 2:

293T lysate at 10 µg

Lane 3:

HL60 lysate at 10 µg

Lane 4:

Fetal heart lysate at 10 µg

Secondary

All lanes:

Goat-anti-rabbit HRP at 1/2000 dilution

Predicted band size: 30 kDa

true