APRIL Antibody [Aprily-8]

APRIL antibody can be used to recognize human APRIL by Flow Cytometry, immunocytochemistry and Western Blot. Method: 3 days after transfection of cells with the indicated constructs, cells were fixed with 4% formaldehyde 5 min at RT. After a wash in PBS, samples were dehydrated by washes in 60%, 80%, 90%, 100% EtOH and xylol. Cells were then dried and embedded in paraffin. Sections were cut, mounted on slides and dried overnight at 50˚ C. Slides were then successively washed 2x 10min in xylol, 2x 10min in 100% EtOH, and then treated 10 min in 100% MeOH/ 0.6% H202 to inhibit endogenous peroxidase. Samples were rehydrated by washes in 90%, 80%, 60% EtOH and PBS. After micro-wave treatment, slides were washed 3x in PBS, blocked with IgG, and incubated for 1 hour with 5 μ g/mL Aprily-8 or control mouse IgG (isotype control) in 1%BSA / 1x PBS for 1 hour. After PBS washes, samples were incubated with the secondary Ab for 1 hour, washed in PBS and revealed with StreptABComplex/HRP (Vector) and AEC. Method: HEK 293 cells were mock transfected or transfected with an expression plasmid coding for human APRIL. Cells (5x105) were incubated on ice for 30 min in 50 μ l FACS buffer (PBS, 5% fetal calf serum, 0.02% azide) containing 10 μ g/mL of Aprily-8 antibody. After washing in FACS buffer, FITC-conjugated antibody to mouse IgG was added. Cells were incubated on ice for 30 min, washed and analyzed by flow cytometry. Method: 10 μ g of protein was applied to the gel, and revealed with Aprily-8 (1 μ g/mL) and HRP-coupled anti-mouse secondary antibody (1:4,000) .Note: The hAPRIL construct used in this experiment is an uncleavable fusion protein between hBAFF (aa 1-132) and hAPRIL (aa 93-233) . This product is for research use only.