SynaptoGreen™ and SynaptoRed™ dyes vary in the length of the lipophilic tail and the number of double bonds linking the two aromatic rings in the dye (Fig. 2). In general, dyes with longer tails and more double bonds have a higher affinity for membrane and thus a higher on-rate and lower off-rate. AM and HM dyes possess an additional aldehyde-fixable amine or hydrazide group, which makes them more water-soluble with a higher off-rate and lower on-rate than the corresponding SynaptoGreen™/SynaptoRed™ or FM® dye