A composition including a N-[4-[[4-(diethyl amino) phenyl](2,5-disulfophenyl)
methylene]-2,5-cyclohexadien-1-ylidene]-N-ethylethanaminium
sodium salt, where the N-4-4-(diethylamino)phenyl (2,5-disulfophenyl)
methylene-2,5-cyclohexadien-1- ylidene-N-ethylethanaminium
sodium salt is at least 90% of the weight of such composition. A method of making the composition by a process including sulphonating 2-chlorobenzaldehyde, treatment with
sodium sulphite and subsequent basification, a condensation step, an oxidation step and a purification step comprising utilizing HPLC to monitor the sulphonating step, the treatment step, the condensation step, and / or the oxidation step, to determine when one or more reactions of such sulphonating step, treatment step, condensation step, and / or oxidation step are substantially complete and ending such one or more reactions on the basis of the HPLC monitoring. The purification step utilizes flash
chromatography to obtain a high purity N-[4-[[4-(diethyl amino) phenyl](2,5-disulfophenyl)
methylene]-2,5-cyclohexadien-1-ylidene]-N-ethylethanaminium
sodium salt.