Isoform 1 is on the cell surface. Western blots of fractions of surface biotinylated HEK cells transfected with HA tagged GPR110 isoform 1 constructs (HA466 and HA1393), or with no tag (Iso1). (A) Developed with antibody to GAPDH (first blot), to integrin (second blot), or HA.11 antibody (third blot). L, lysate; U, unbound, and B, bound cell surface fraction. GAPDH is a cytosolic protein (arrow on the first blot); and integrin β1 is a surface protein (arrow on the second blot). Arrow on the third blot, 100 kD band of isoform 1. (B) Treatment of tagged GPR110 isoform 1 surface fractions with (+) or without (-) PNGaseF; developed with HA.11 antibody. Arrows point to the 60 and 75 kD bands, a result of the PNGaseF treatment; and to a 25 kD band, of the putative SEA fragment.