After washing, secondary antibody was incubated for 30 minutes.
After washing, secondary antibody and BGT-AF647 were added and incubated over night at 4°C.
Primary antibodies were applied overnight at 4°C and, after washing, secondary antibodies were applied for 1 h at room temperature.
After washing, secondary antibodies, which were conjugated to either Alexa-488 or Alexa-568 (Invitrogen, Carlsbad, CA), were added at a 1∶200 dilution and incubated for 1 2 hrs.
After washing, secondary antibodies against the species were combined and diluted 1∶500 in 1% GS-PBS (Alexa 488 anti-mouse and Alexa 546 anti-rabbit from Invitrogen).
After PBS washes, secondary antibodies (30 µg/ml goat anti-mouse or -rabbit IgG conjugated with FITC, Jackson ImmunoResearch) were applied at room temperature for 1 h.
After washing, secondary horseradish peroxidase conjugated antibody was added, and the membranes were developed using the Enhanced Chemiluminescence Plus Western Blotting Detection System (Amersham Biosciences, Piscataway, NJ).
After three 10 minute 1XPBS-T washes secondary goat anti-rabbit antibody (Jackson Immunologicals) was incubated at 1∶500 for 2 hrs at room temperature.
After washing, secondary donkey-anti-goat antibody SC-20200 (Santa Cruz Biotech, Santa Cruz, CA) was applied at a 1∶100 ratio in PBST with a 1∶10000 dilution of DAPI stain (Invitrogen) for one hour.
After washing, secondary antibodies were added and sections incubated for 1.5 h at room temperature.
