EGFR Recombinant Rabbit Monoclonal Antibody [SZ40-19]

☑ Relative expression (RE)

Western blot analysis of EGFR on different lysates with Rabbit anti-EGFR antibody (ET1603-37) at 1/2,000 dilution and competitor's antibody at 1/2,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: A431 cell lysate
Lane 3: MDA-MB-468 cell lysate
Lane 4: MCF7 cell lysate (low expression)

Lysates/proteins at 15 µg/Lane.

Predicted band size: 134 kDa
Observed band size: 150 kDa

Exposure time: 21 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 (ET1603-37) at 1/2,000 dilution and competitor's antibody at 1/2,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

☑ Knockdown (KD)

Western blot analysis of EGFR on different lysates with Rabbit anti-EGFR antibody (ET1603-37) at 1/2,000 dilution.

Lane 1: A431-si NT cell lysate
Lane 2: A431-si EGFR cell lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 134 kDa
Observed band size: 150 kDa

Exposure time: 2 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 (ET1603-37) at 1/2,000 dilution was used in 5% NFDM/TBST 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 A431 cells labeling EGFR with Rabbit anti-EGFR antibody (ET1603-37) at 1/500 dilution and competitor's antibody at 1/200 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-EGFR antibody (ET1603-37) at 1/500 dilution and competitor's antibody at 1/200 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 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 A431 cells labeling EGFR.

Cells were fixed and permeabilized. Then stained with the primary antibody (ET1603-37, red) at 1/1,000 dilution and competitor's antibody (red) at 1/200 dilution, 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).

ICC staining of EGFR in Hela cells (red). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 10% negative goat serum for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1603-37, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®594 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).

ICC staining of EGFR in HepG2 cells (red). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 10% negative goat serum for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1603-37, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®594 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).

mIHC analysis of human lung cancer tissue (Formalin/PFA-fixed paraffin-embedded sections) with Rabbit anti-EGFR antibody (ET1603-37) at 1/1,000 dilution. The immunostaining was performed with the IRISKit® HyperView mTSA Kit (MH900206). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.

mIHC analysis of human kidney cortex tissue (Formalin/PFA-fixed paraffin-embedded sections) with Rabbit anti-EGFR antibody (ET1603-37) at 1/1,000 dilution. The immunostaining was performed with the IRISKit® HyperView mTSA Kit (MH900206). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.

mIHC analysis of human kidney medulla tissue (Formalin/PFA-fixed paraffin-embedded sections) with Rabbit anti-EGFR antibody (ET1603-37) at 1/1,000 dilution. The immunostaining was performed with the IRISKit® HyperView mTSA Kit (MH900206). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.