LAG-3 Recombinant Rabbit Monoclonal Antibody [PS01-33]

Immunohistochemical analysis of paraffin-embedded human tonsils tissue with Rabbit anti-LAG-3 antibody (HA721358) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA721358) at 1/200 dilution for 1 hour 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.

☑ Cell treatment (CT)

Western blot analysis of LAG-3 on different lysates with Rabbit anti-LAG-3 antibody (HA721358) at 1/1,000 dilution.

Lane 1: NIH/3T3 transfected with LAG3 expression vector, whole cell lysate
Lane 2: NIH/3T3 cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 57.4 kDa
Observed band size: 57/70 kDa

Exposure time: 2 minutes;

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 (HA721358) at 1/1,000 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.

☑ Cell treatment (CT)

Immunocytochemistry analysis of NIH/3T3 cells transfected with or without LAG-3 labeling LAG-3 with Rabbit anti-LAG-3 antibody (HA721358) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 10 minutes at 37 ℃, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes, and then blocked with 1% BSA for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-LAG-3 antibody (HA721358) 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 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

mIHC analysis of human tonsils tissue (Formalin/PFA-fixed paraffin-embedded sections) with Rabbit anti-LAG-3 antibody (HA721358) at 1/400 dilution. The immunostaining was performed with the IRISKitCmTSA Kit (900808). 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.