Anti-GABARAP + GABARAPL1 + GABARAPL3 antibody [EPR18862] (ab191888) at 1/1000 dilution + Human GABARAPL1 full length recombinant protein at 0.02 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 14 kDa


Exposure time: 5 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

Human GABARAPL1 full length recombinant protein contains aa1-117 with a GST-Tag.

All lanes : Anti-GABARAP + GABARAPL1 + GABARAPL3 antibody [EPR18862] (ab191888) at 1/5000 dilution

Lane 1 : Recombinant human GABARAPL2 full length protein
Lane 2 : Recombinant human GABARAPL3 full length protein
Lane 3 : Recombinant human GABARAP full length protein

Lysates/proteins at 0.01 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 14 kDa

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure times: Lane 1 and 2: 5 seconds; Lane 3: 3 minutes.

All three recombinant human full length proteins contain aa1-117 with an N-terminal His-Tag^®. This product showed higher reactivity with GABARAPL1 and GABARAPL3, and lower reactivity with GABARAP.

Anti-GABARAP + GABARAPL1 + GABARAPL3 antibody [EPR18862] (ab191888) at 1/1000 dilution + Human brain lysate at 10 µg

Secondary
Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 14 kDa
Observed band size: 17 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.

The observed MW is a little higher than predicted one (14KD), which is similar with the published literatures (PMID: 16650615 & 21862879).

All lanes : Anti-GABARAP + GABARAPL1 + GABARAPL3 antibody [EPR18862] (ab191888) at 1/1000 dilution

Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution

Predicted band size: 14 kDa
Observed band size: 17 kDa why is the actual band size different from the predicted?


Exposure time: 30 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time:30 seconds

The observed MW is a little higher than predicted one (14KD), which is similar with the published literatures (PMID: 16650615 & 21862879).

All lanes : Anti-GABARAP + GABARAPL1 + GABARAPL3 antibody [EPR18862] (ab191888) at 1/1000 dilution

Lane 1 : Mouse brain lysate
Lane 2 : Mouse kidney lysate
Lane 3 : Mouse spleen lysate
Lane 4 : Rat brain lysate
Lane 5 : Rat kidney lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 14 kDa
Observed band size: 17 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.

The observed MW is a little higher than predicted one (14KD), which is similar with the published literatures (PMID: 16650615 & 21862879).

GABARAP + GABARAPL1 + GABARAPL3 was immunoprecipitated from 1 mg of Mouse brain lysate with ab191888 at 1/150 dilution. Western blot was performed from the immunoprecipitate using ab191888 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

Lane 1: Mouse brain lysate, 10µg (Input).

Lane 2: ab191888 IP in Mouse brain lysate.

Lane 3: Rabbit IgG,monoclonal [EPR25A]- Isotype Control (ab172730) instead of ab191888 in Mouse brain lysate.

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

Exposure time: 3 minutes.