Anti-PNPLA3 antibody [EPR29816-94]

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-PNPLA3 antibody [EPR29816-94] (AB326041)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HepG2 (human hepatocellular carcinoma epithelial cell) cells labelling PNPLA3 with ab326041 at 1/50 (10.66 µg/ml) dilution , followed by ab150088 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed (ab150088) antibody at 1/1000 dilution (Green).

Confocal image showing cytoplasmic staining in HepG2 cell line (shown in magenta). The counterstain was observed in green. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Lipid Droplets Green Fluorescence Assay Kit with BODIPY493/503 was used to counterstain tubulin at 1/200 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150088 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed (ab150088) at 1/1000 dilution.

• IP

Lab

Immunoprecipitation - Anti-PNPLA3 antibody [EPR29816-94] (AB326041)

PNPLA3 was immunoprecipitated from 0.35 mg HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate with ab326041 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab326041 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 2 : ab326041 IP in HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab326041 in HepG2 whole cell lysate

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

Exposure time : 10 seconds.

All lanes:

Immunoprecipitation - Anti-PNPLA3 antibody [EPR29816-94] (ab326041) at 1/1000 dilution

Lane 1:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 10 µg

Lane 2:

ab326041 at 1/30 IP in HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab326041 in HepG2 whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 70 kDa

false

Exposure time: 10s

• WB

Lab

Western blot - Anti-PNPLA3 antibody [EPR29816-94] (AB326041)

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

Low expression : Daudi, Raji.

Samples are non-boiled as boiling may cause protein aggregation.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa).

Exposure time : Lanes 1-3 : 102 seconds, Lanes 4 : 180 seconds

All lanes:

Western blot - Anti-PNPLA3 antibody [EPR29816-94] (ab326041) at 1/1000 dilution

Lane 1:

PAZ6 (human Adipose epithelial) whole cell lysate at 50 µg

Lane 2:

Daudi (human Burkitt's lymphoma lymphoblast) whole cell lysate at 50 µg

Lane 3:

Raji (human Burkitt's lymphoma B lymphocyte) whole cell lysate at 50 µg

Lane 4:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 50 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/1000 dilution

Observed band size: 70 kDa,36 kDa

false

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-PNPLA3 antibody [EPR29816-94] (AB326041)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Daudi (human Burkitt's lymphoma lymphoblast) cells labelling PNPLA3 with ab326041 at 1/50 (10.66 µg/ml) dilution , followed by ab150088 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed (ab150088) antibody at 1/1000 dilution (Green).

Confocal image showing no staining in Daudi cell line (shown in magenta). The counterstain was observed in green. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/1000 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150088 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed (ab150088) at 1/1000 dilution.