ab16085 at 1ug/ml detecting  membrane-bound human APRIL/TNFSF13 on HEK 293T cells by FACS analysis.

HEK 293T cells (5x105) were mock transfected (thin line) or transfected with an expression plasmid enabling surface expression of a non-cleavable mouse A proliferation-inducing ligand (thick line). Cells were incubated on ice for 30 min. in 50ul FACS buffer (PBS, 5% fetal calf serum, 0.02% azide) containing 1ug/ml of MAb to APRIL (Sacha-1). After washing in FACS buffer, PE-conjugated antibody to rat IgG was added. Cells were incubated on ice for 30 min., washed and then analyzed by flow cytometry.

Immunostaining of HEK-293T cells transfected with a murine APRIL/TNFSF13 expression plasmid (left panels), or mock transfected (right panel).

Three days after transfection of cells with the indicated constructs, cells were fixed with acetone. Slides were blocked with IgG, and incubated for 1 hour with 5ug/ml ab16085 or control murine IgG (isotype control) in 1%BSA / 1x PBS for 1 hour. After washes wish PBS, samples were incubated with the secondary Ab for 1 hour, washed in PBS and revealed with StreptABComplex/HRP (Vector) and AEC.