Fitc conjugated anti ha antibody

Cells were adhered to glass coverslips, treated with PEME buffer for 5 min at room temperature, and then fixed with 4% paraformaldehyde for 10 min at room temperature. Cells were washed once with PBS, incubated with the blocking buffer (3% BSA in PBS) for 1 h at room temperature, and then incubated with the primary antibody for 1 h at room temperature. The following primary antibodies were used: anti-CIF1 polyclonal antibody (1:1,000 dilution) (Zhou et al., 2018a (link)), anti-CIF2 polyclonal antibody (1: 1,000 dilution) (Zhang et al., 2019 (link)), anti-Protein A (anti-ProtA) polyclonal antibody (1:400 dilution) (Sigma-Aldrich), and FITC-conjugated anti-HA antibody (1:400 dilution) (Sigma-Aldrich). Cells were washed three times with PBS containing 0.1% Triton X-100. Except for the FITC-conjugated anti-HA antibody, cells were subsequently incubated with Cy3-conjugated anti-rabbit IgG at room temperature for 1 h. The slides were mounted in VectaShield mounting medium (Vector Labs) containing DAPI and examined using an inverted microscope (Model IX71, Olympus) equipped with a cooled CCD camera (model Orca-ER, Hamamatsu) and a PlanApo N 60 × 1.42-NA DIC objective. Images were acquired and processed using the Slidebook5 software (Intelligent Imaging Innovations, Inc).