Tyrosinase Recombinant Rabbit Monoclonal Antibody [JA52-11]

Western blot analysis of Tyrosinase on SK-MEL-28 cell lysates with Rabbit anti-Tyrosinase antibody (ET1704-18) at 1/2,000 dilution.

Lysates/proteins at 20 µg/Lane.

Predicted band size: 60 kDa
Observed band size: 60 kDa

Exposure time: 46 seconds; ECL: K1802;

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 (ET1704-18) 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.

Western blot analysis of Tyrosinase on B16F1 cell lysates. Proteins were transferred to a PVDF membrane and blocked with 5% BSA in PBS for 1 hour at room temperature. The primary antibody (ET1704-18, 1/500) was used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.

Immunocytochemistry analysis of SK-MEL-28 cells labeling Tyrosinase with Rabbit anti-Tyrosinase antibody (ET1704-18) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 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-Tyrosinase antibody (ET1704-18) at 1/100 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 (HA601187, 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.

Application: Immunohistochemistry (IHC-P)

Species: Human
Tissue: Melanoma
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95℃.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: ET1704-18, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.

Application: Immunohistochemistry (IHC-P)

Species: Human
Tissue: Skin
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95℃.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: ET1704-18, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.

☑ Relative expression (RE)

Application: Immunohistochemistry (IHC-P)

Species: Human
Tissue: Liver
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95℃.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: ET1704-18, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.

Negative expression of Tyrosinase protein in liver is consistent with the predicted expression pattern.

Tyrosinase was immunoprecipitated from 0.2 mg SK-MEL-28 cell lysate with ET1704-18 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using ET1704-18 at 1/2,000 dilution. HRP Conjugated Anti-Rabbit IgG for IP Nano-secondary antibody at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: SK-MEL-28 cell lysate (input)
Lane 2: ET1704-18 IP in SK-MEL-28 cell lysate
Lane 3: Rabbit IgG instead of ET1704-18 in SK-MEL-28 cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 3 minutes; ECL: K1801

Flow cytometric analysis of SK-MEL-28 cells labeling Tyrosinase.

Cells were washed twice with cold PBS and resuspend. Then stained with the primary antibody (ET1704-18, 1/1,000) (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).