RAB8A Recombinant Rabbit Monoclonal Antibody [JE64-79]

Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (HA721127) at 1/1,000 dilution.

Lane 1: HeLa (Human cervical adenocarcinoma cell)
Lane 2: Hep G2 (Human hepatocellular carcinoma cell)

Lysates/proteins at 15 µg/Lane.
Exposure time: 1 minutes; ECL: K1801


Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: HA721127, 1/1,000 in 5% NFDM/TBST, overnight at 4 ℃
Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature

Predicted band size: 24 kDa
Observed band size: 24 kDa

Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (HA721127) at 1/500 dilution.

Lane 1: NIH/3T3 cell lysate
Lane 2: RAW264.7 cell lysate
Lane 3: C2C12 cell lysate
Lane 4: L6 cell lysate

Lysates/proteins at 30 µg/Lane.

Predicted band size: 24 kDa
Observed band size: 24 kDa

Exposure time: 2 minutes 16 seconds;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721127) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:100,000 dilution was used for 1 hour at room temperature.

☑ Knockdown (KD)

Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (HA721127) at 1/1,000 dilution.

Lane 1: HAP1-parental cell lysate
Lane 2: HAP1-RAB8A KD cell lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 24 kDa
Observed band size: 24 kDa

Exposure time: 30 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721127) at 1/1,000 dilution was used in K1803 at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

Immunocytochemistry analysis of HeLa cells labeling RAB8A with Rabbit anti-RAB8A antibody (HA721127) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 10 minutes at 37 ℃, permeabilized with 0.05% Triton X-100 in PBS for 20 minutes, and then blocked with 2% negative goat serum for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-RAB8A antibody (HA721127) at 1/100 dilution in 2% negative goat serum overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/200 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

Flow cytometric analysis of HeLa cells labeling RAB8A.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA721127, 1ug/ml) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

RAB8A was immunoprecipitated from 0.2 mg HeLa cell lysate with HA721127 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA721127 at 1/1,000 dilution. Anti-Rabbit IgG for IP, AlpSdAbs® VHH(HRP) (025-100-005) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: HeLa cell lysate (input)
Lane 2: HA721127 IP in HeLa cell lysate
Lane 3: Rabbit IgG instead of HA721127 in HeLa cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 4 seconds; ECL: K1801