All lanes : Anti-SLC27A4 / FATP4 antibody [EPR17319] - C-terminal (ab199719) at 1/1000 dilution

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : SLC27A4 knockout HEK293T cell lysate
Lane 3 : HepG2 cell lysate
Lane 4 : HEK-293 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: 72 kDa
Observed band size: 72 kDa

Lanes 1-4: Merged signal (red and green). Green - ab199719 observed at 72 kDa. Red - loading control ab8245 observed at 36 kDa.

 ab199719 Anti-SLC27A4 / FATP4 antibody [EPR17319] - C-terminal  was shown to specifically react with SLC27A4 / FATP4 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266114 (knockout cell lysate ab257677) was used. Wild-type and SLC27A4 / FATP4 knockout samples were subjected to SDS-PAGE. ab199719 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HepG2 (Human liver hepatocellular carcinoma) cells labeling SLC27A4 / FATP4 with ab199719 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 dilution (green). Cytoplasm staining on HepG2 cell line is observed. The nuclear counterstain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution (red).
The negative controls are as follows:-
-ve control 1: ab199719 at 1/100 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 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 1/500 dilution.

 

Immunohistochemical analysis of paraffin-embedded Human kidney tissue labeling SLC27A4 / FATP4 with ab199719 at 1/100 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on Human kidney tissue is observed. Counter stained with Hematoxylin.
Negative control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051).

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

SLC27A4 / FATP4 was immunoprecipitated from 1mg of HEK293 (Human embryonic kidney) whole cell lysate with ab199719 at 1/50 dilution. Western blot was performed from the immunoprecipitate using ab199719 at 1/5000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution.

Lane 1: HEK293  whole cell lysate 10µg (Input). Lane 2: HEK293 whole cell lysate following IP. Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab199719 in HEK293 whole cell lysate.

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure is 3 minutes.

Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: SLC27A4/FATP4 knockout HAP1 cell lysate (20 µg)
Lane 3: HEK293 cell lysate (20 µg)
Lane 4: HepG2 cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab199719 observed at 70 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab199719 was shown to specifically react with SLC27A4/FATP4 when SLC27A4/FATP4 knockout samples were used. Wild-type and SLC27A4/FATP4 knockout samples were subjected to SDS-PAGE. ab199719 and ab8245 (loading control to GAPDH) were diluted 1/2000 and 1/10000 respectively and incubated overnight at 4°C. 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 1/10000 dilution for 1 h at room temperature before imaging.

All lanes : Anti-SLC27A4 / FATP4 antibody [EPR17319] - C-terminal (ab199719) at 1/10000 dilution

Lane 1 : HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate
Lane 2 : HepG2 (Human liver hepatocellular carcinoma) whole cell lysate
Lane 3 : HEK293 (Human embryonic kidney) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 72 kDa
Observed band size: 72 kDa


Exposure time: 1 minute

Blocking/dilution buffer: 5% NFDM/TBST.

All lanes : Anti-SLC27A4 / FATP4 antibody [EPR17319] - C-terminal (ab199719) at 1/2000 dilution

Lane 1 : Human fetal brain
Lane 2 : Human fetal kidney

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 72 kDa
Observed band size: 72 kDa


Exposure time: 1 minute

Blocking/dilution buffer: 5% NFDM/TBST.