Alexa fluor 568 goat anti rabbit igg h l antibody

Cells were transfected with plasmids or siRNAs and re-plated on Matrigel-coated coverslips after 24–48 h of transfection. The cells were allowed to adhere and spread for 24 h, and then fixed with 4% paraformaldehyde for 10 min at room temperature, followed by permeabilization using 0.2% Triton X-100 in phosphate-buffered saline (PBS) for 5 min. The coverslips were washed twice with PBS and incubated with the primary antibodies for 2 h at room temperature, and then treated with secondary antibodies for 1 h at room temperature. Alexa Fluor 568 Goat Anti-Rabbit IgG (H+L) Antibody and Alexa Fluor 647 Goat Anti-Mouse IgG (H+L) Antibody (both from Life Technologies) were used as secondary antibodies. The coverslips were mounted with PermaFluor Aqueous Mounting Medium (Thermo Scientific, Waltham, MA, USA) and observed using a FluoView 1000-D confocal microscope (Olympus, Tokyo, Japan).