Anti ev d68 vp1 antibody

Cell samples were harvested and lysed in RIPA buffer (1 M Tris pH 7.8, 1M NaCl, 1% NP-40, 0.5 M EDTA). The cell lysate was separated on 12% SDS-PAGE gels and then transferred to nitrocellulose membranes using a semidry apparatus (Bio-Rad). Anti-HA was purchased from Thermo Fisher Scientific (Waltham, MA, United States). The anti-GAPDH antibody and anti-α-Tubulin antibody were purchased from GenScript Biotech Corp (NJ, United States). Anti-EV-D68 VP1 antibody and anti-EV-A71 VP1 antibody were purchased from GeneTex (San Antonio, TX, United States).

The lung sections were blocked for two hours in 10% normal goat serum to reduce nonspecific antibody binding. The lung was then incubated with anti-EV-D68VP1 antibody 5 μg/mL (GeneTex, Inc., Irvine, CA, USA) and with biotinylated MAA (α2,3-linkage) and SNA (α2,6-linkage) 5 μg/mL (Vector Laboratories, Inc., CA, USA) at 4 °C overnight. The sections were rinsed extensively in Tris-buffered saline. The cell nuclei were strained by DAPI (Beyotime Biotech, Co., Ltd., Shanghai, China). The primary antibody was detected using 5 μg/mL of Fluorescein Avidin DCS (Vector Laboratories, Inc., Burlingame, CA, USA) and 2 μg/mL Texas-Red-conjugated goat anti-rabbit IgG antibodies (Molecular Probes, Carlsbad, CA, USA) [20 (link),21 (link)]. The stained slides were analyzed under a Leica TCS SP8Laser Confocal microscope (Leica Microsystems, Wetzlar, Germany).