Galectin 1 Recombinant Rabbit Monoclonal Antibody [JM13-37]

☑ Relative expression (RE)

Western blot analysis of Galectin 1 on different lysates with Rabbit anti-Galectin 1 antibody (ET1705-83) at 1/2,000 dilution.

Lane 1: HL-60 cell lysate
Lane 2: LNCaP cell lysate (negative)
Lane 3: A549 cell lysate
Lane 4: HCT 116 cell lysate

Lysates/proteins at 15 µg/Lane.

Predicted band size: 15 kDa
Observed band size: 14 kDa

Exposure time: 1 minute 17 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 (ET1705-83) 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 Galectin 1 on different lysates with Rabbit anti-Galectin 1 antibody (ET1705-83) at 1/2,000 dilution.

Lane 1: NIH/3T3 cell lysate (20 µg/Lane)
Lane 2: L6 cell lysate (20 µg/Lane)
Lane 3: Mouse kidney tissue lysate (40 µg/Lane)
Lane 4: Mouse colon tissue lysate (40 µg/Lane)

Predicted band size: 15 kDa
Observed band size: 14 kDa

Exposure time: 20 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 (ET1705-83) 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 NIH/3T3 cells labeling Galectin 1 with Rabbit anti-Galectin 1 antibody (ET1705-83) at 1/100 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-Galectin 1 antibody (ET1705-83) 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 (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.

Immunocytochemistry analysis of L6 cells labeling Galectin 1 with Rabbit anti-Galectin 1 antibody (ET1705-83) at 1/100 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-Galectin 1 antibody (ET1705-83) 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 (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.

Immunohistochemical analysis of paraffin-embedded rat stomach tissue using anti-Galectin 1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1705-83, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

Immunohistochemical analysis of paraffin-embedded human placenta tissue using anti-Galectin 1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1705-83, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

Flow cytometric analysis of NIH/3T3 cells labeling Galectin 1.

Cells were fixed and permeabilized. Then stained with the primary antibody (ET1705-83, 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).

Flow cytometric analysis of L6 cells labeling Galectin 1.

Cells were fixed and permeabilized. Then stained with the primary antibody (ET1705-83, 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).

Immunoprecipitation (IP)

Galectin 1 was immunoprecipitated in 0.2 mg HL-60 (Human acute promyelocytic leukemia cell) cell lysate with ET1705-83 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using ET1705-83 at 1/2,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: HL-60 (Human acute promyelocytic leukemia cell) cell lysate (input)
Lane 2: ET1705-83 IP in HL-60 (Human acute promyelocytic leukemia cell) cell lysate
Lane 3: Rabbit IgG instead of ET1705-83 in HL-60 (Human acute promyelocytic leukemia cell) cell lysate

Exposure time: 2 seconds
Blocking: 5% NFDM/TBST, 1 hour at room temperature

Primary dilution: ET1705-83, 1/2,000 in primary antibody dilution buffer (K1803), 2 hours at room temperature

Predicted band size: 15 kDa
Observed band size: 15 kDa