A new cationic indolium based styryl dye (Ci) as a fluorescent probe was synthesized and its anions selectivity/sensitivity properties/molecular interactions with protease enzymes (pepsin/trypsin) and ctDNA has been studied by spectroscopic and computational methods. The fluorescence measurements at different temperatures indicated that quenching mechanism of enzymes by Ci was static. Delta H and Delta S data pointed out electrostatic/hydrophobic interactions with pepsin, and also hydrogen bonds/van der Waals forces with trypsin of Ci. According to Forster's non-radiative energy transfer, binding distances (r) were calculated as 3.53/3.27 nm for pepsin/trypsin. It was also investigated that groove binding is effective in interaction with ctDNA. The results were supported with molecular docking analyzes which have same tendency. Ci has been demonstrated hypsochromic effect with a decrease in polarity of solvents and it showed highly selective colorimetric and fluorometric sensing behavior for cyanide in organic solvent and in aqueous solution. H-1 NMR titration was performed to examine the interaction mechanism between Ci and cyanide. The LOD values of cyanide ion were reported as 4.87 x 10(-9) M and 9.70 x 10(-7) M in DMSO and DMSO/H2O binary mixture, respectively. In addition, sensitivity of Ci as a chemosensor to cyanide was investigated in bitter almond samples.