Il md led inverted fluorescence microscope

Immunofluorescence (IF) and immunocytochemistry (ICC) were performed as follows. Cells were fixed with 4% paraformaldehyde (PFA) for 10 min at room temperature and permeabilized with 0.2% Triton X-100 before immunostaining. The cells were washed in PBS and blocked for 1 h in blocking buffer (10% goat serum in PBS). Samples were incubated with Anti-Fibrillarin antibody (G-8) (sc-374022_Santa Cruz Biotechnology), Anti-Ribosomal ProteinL28 (A-16) (sc-14151_Santa Cruz Biotechnology), Anti-Histone H1 (AE-4) (sc-8030_Santa Cruz Biotechnology), and our Anti RpL22/H5 for 1 h at r.t., washed three times in PBS and incubated with Alexa Fluor 488 goat anti-rabbit secondary antibody (Life Technologies Carlsbad, CA, USA, 1:200 dilution) and Alexa Fluor 488 goat anti-mouse secondary antibody (Life Technologies, Carlsbad, CA, USA 1:200 dilution) for 1 h at r.t. for detection. Counterstaining was done with DAPI. Images were acquired using a Leica IL MD LED inverted fluorescence microscope. To ensure the validity and specificity of the anti RpL22/H5 antibody, we conducted IF and ICC experiments using the pre-immune serum under the same experimental conditions described above without obtaining any signal on the tested cells or tissues.