15 μg of U-937 lysate was run on 6-18% SDS-PAGE under reducing conditions and blotted onto nitrocellulose membrane. antibody at 1 μg/mL was used as the primary antibody and peroxidase conjugated goat anti-mouse IgG was used as the secondary antibody. LAT2 band was visualized using ECL Western Blotting Substrate.

Result: antibody can detect LAT2 by Western blotting.

IHC-P analysis of tonsil tissue by LAT2 antibody.
IHC-P was performed using sections of the formalin-fixed paraffin-embedded tonsil tissue. Antigen was retrieved through addition of boiling Tris/EDTA buffer pH 9 in a pressure cooker for 3 min. Endogenous peroxidase activity was quenched by incubating the sections with 3% H₂O₂ for 30 min at room temperature. The sections were then incubated with LAT2 antibody at 7.5 μg/mL at room temperature for 1 h. Poly-peroxidase conjugated goat anti-mouse IgG was used as the secondary antibody. Diaminobenzidine was used as the chromogen. The section was counterstained with hematoxylin. A tissue section incubated with phosphate-buffered saline followed by incubation with the secondary antibody was used as the background control.

Result: Germinal center cells and non-germinal center cells are positively stained at the cytoplasm and membrane.

Immunoprecipitation was performed by incubation of 2.5 μg of antibody with U-937 lysate containing 200 μg of total protein. After absorption with Protein G beads, the mixture was run on 6-18% SDS-PAGE and blotted onto nitrocellulose membrane. LAT2 (antibody) at 1 μg/mL was used as the primary antibody and peroxidase conjugated goat anti-mouse IgG (Fc specific) was used as the secondary antibody.

Lane 1: U-937 lysate

Lane 2: LAT2 immunoprecipitated from U-937 lysate by antibody

Lane 3: The same as Lane 2 but KT82 was used as IgG isotype control antibody

Result: antibody can immunoprecipitate LAT2.