Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)

ab192890 staining LC3B in HeLa cells +/- Chloroquine (50μM, 24 hours). The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab192890 at 1μg/ml and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 594), at 1/250 dilution (shown in pseudocolor red) followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab192890).

Immunocytochemistry/ Immunofluorescence analysis of HeLa cells labeling LC3B with ab192890 at 1/500 dilution. Cells were fixed in Methanol. Staining with ab192890 at 1/500 was carried out for 1 hour at 22°C in PBS buffer. ab150081, a Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed secondary antibody was used at 1/200 dilution. DAPI was used to counterstain.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab192890).

LC3B was immunoprecipitated from 1mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab192890 at 1/50 dilution. Western blot was performed from the immunoprecipitate using ab192890 at 1/1000 dilution. Veriblot for IP (HRP) (ab131366), was used as secondary antibody at 1/10000 dilution.
Lane 1: HeLa whole cell lysate 10µg (Input).
Lane 2: ab192890 IP in Jurkat whole cell lysate.
Lane 3: Rabbit IgG,monoclonal [EPR25A] -Isotype Control (ab172730) instead of ab192890 in HeLa whole cell cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 30 seconds.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab192890).

This ICC data was generated using the same anti-LC3B antibody clone, EPR18709, in a different buffer formulation (cat# ab192890).

ab192890 staining LC3B in HAP1 cells (wildtype and MAP1LC3B knockout) +/- Chloroquine (50μM, 24 hours). The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab192890 at 1μg/ml and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 594), at 1/250 dilution (shown in pseudocolor red) followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).