Fusion fx7 scanner

Cells were collected four to seven days post-transfection and lysed in sodium
dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) sample buffer
[13 ]. Cell lysates were separated
by SDS-PAGE and transferred onto nitrocellulose membranes. The membranes were
blocked with 5% fat-free milk powder in 150 mM NaCl, 50 mM Tris-HCL pH 8.0, 0.1%
Tween-20 (TBST) and incubated overnight at 4°C with the following antibodies: TERT
(Y182), PI3K p110α (04-399), ΔN-p63 (p40 [5 (link)]-[17 ] PC373),
GATA3 (09-076) (Millipore); BMI1 (D20B7), keratin 18 (DC10), cyclin D1 (DCS6), myc
(D84C12), AKT (9272), phospho-AKT-T308 (4056/244 F9), ERBB3 (4754) (Cell Signaling
Technology); FOXA1 (Ab55-178) (Abcam); AGR2 (1C3), tubulin (B-5-1-2) (Sigma);
keratin 14 (LL002, gift from Birgit Lane); ERα (Ab-16, RB-1493)
(ThermoScientific); p53 (D01, gift from David Lane). After three washes in TBST,
bound primary antibodies were detected by incubation with HRP-conjugated
anti-rabbit, anti-mouse or anti-goat IgG (GE Healthcare) at room temperature for
1 hour, washed again in TBST, and visualized using ECL reagents (GE Healthcare).
Images were captured on a Fusion FX7 scanner (Vilber Lourmat) or Hyperfilm (GE
Healthcare).

Mosquito hemolymph from 10 mosquitoes was collected by proboscis clipping directly into 20 μl of Laemmli buffer. Samples were heated at 90°C for 2–5 minutes and 2 to 8 μl (depending on the experiment) were loaded on polyacrylamide gels. Rabbit polyclonal antibodies against TEP1, APL1C, LRIM1, PPO2 and Vg were used. Secondary HRP conjugated antibodies were used at 1:15000 dilution. Images were collected on Fusion FX7 scanner (Vilber Lourmat).