Mouse anti v5 primary antibody

The following antibodies were used for western blot analysis at the indicated dilutions in either blotto (1x Phosphate Buffered Saline containing 10 mM Na2HPO4 and 1.8 mM KH2PO4 at pH 7.4, 137 mM NaCl, 2.7 mM KCl, 0.1% tween-20 and 5% w/v nonfat powdered milk) or TBST (1x Tris-HCl buffered saline containing 50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 0.1% tween-20, and 5% w/v bovine serum albumin), as specified by the antibody′s manufacturer.
Mouse anti-Tubulin (Cell Signaling Technologies, Cat# 3873) at 1:1000 in blotto.
Rabbit anti-Myc (Cell Signaling Technologies, Cat# 2278S) at 1:5000 in TBST.
Mouse anti-V5 primary antibody (Invitrogen; catalog no.: R960–25) at 1:5000 dilution in blotto.
Goat anti-rabbit-HRP secondary antibody (Cell Signaling Technologies, Cat# 7074P2 or Sigma, Cat# AP187P) at 1:5000 in blotto.
Goat anti-mouse-HRP (Thermo Fisher, Cat# 31430) at 1:5000 in blotto.

Immunofluorescence (IF) was described previously (47 (link)). After transfection, cells grown on coverslips were fixed in 2% formaldehyde for 15 min, washed 2 times with phosphate buffered saline (PBS) and permeabilized with PBS-Triton X-100 (0.1%). Cells were washed with PBS and with PBS plus (PBS with 0.5% bovine serum albumin (BSA), 0.15% glycine), followed by primary antibody incubation overnight at 4°C. Cells on coverslips were washed with PBS plus for 4 times and were incubated with secondary antibody. After washing with PBS plus and PBS DNA was stained with DAPI or Hoechst (Thermo Fisher Scientific) for 5 min followed by washing with PBS. Cover slips were mounted in Vectashield (Agar Scientific). Mouse anti-v5 primary antibody (Invitrogen) at a 1:10 000 dilution, Rabbit anti-DDX6 primary antibody (Novus) at 1:1000, Alexa 594 anti-mouse (Invitrogen) secondary antibody was diluted as 1:1500, Alexa 633 anti-rabbit (Invitrogen) secondary antibody was diluted as 1:1000. Images were taken by a Leica sp8 confocal microscope and edited by Fiji.