What is the difference between ProLong and SlowFade antifade reagents?
Our ProLong antifade reagents dispense as a liquid that will solidify upon the evaporation of water. SlowFade antifade reagents remain liquid. If you are going to image right away and then dispose of your sample, you do not need a mountant that hardens, such as the SlowFade reagents. If you wish to archive your slide for more than a day, you will want a mounting medium that hardens (or cures). This hardening will limit the off-rates of various dye-conjugated antibodies and provides a better refractive index. Also, there will be a lower diffusion rate of free radicals, limiting photobleaching.
Some antifade mounting media stay as liquid whereas others harden. What is the benefit of having one that hardens?
If you are going to image right away and then dispose of your sample, you probably want a mountant that does not harden. If you wish to archive your slide for more than a day, you want a mountant that hardens (or "cures"). This hardening will slow or prevent off-rate of your dye or conjugate and often produces a better refractive index. Secondary sealing is usually not necessary. Also there will be lower diffusion of free radicals, thus limiting photobleaching.
Does Slowfade Antifade Mountant require using a nail polish for sealing?
No. Samples mounted with SlowFade mountants need not be sealed; they are intended for immediate viewing. The coverslip may be anchored (to prevent movement while viewing) by applying molten paraffin to three or four spots around the edge of the coverslip.
DAPI and Hoechst dyes are quite similar to each other. Why would I choose one over the other?
DAPI is a very common blue-fluorescent dye for nuclear counterstaining and gives very bright labeling on nuclei in fixed and permeabilized cells and tissues. However, it is considered to be a semi-permeant to impermeant stain and provides inconsistent staining of live cells. Hoechst 33342 dye is cell-permeant and stains with the same binding mechanism and fluorescent color; it is preferred for live-cell imaging and is just as good as DAPI for fixed cell labeling.