o-Phthalaldehyde (OPA), a primary amino compound and a thiol react to produce a ternary isoindole-type fluorophore. The effect of the structure of the constituent amino compound on the chemical stability of isoindole fluorophores was investigated. The amino compounds with steric hindrance around the -NH2group such as L-tryptophan and L-Dopa exhibited a marked stabilization effect on the isoindole fluorophores. By using OPA and L-typtophan as reagents, a procedure was established for the fluorometric determination of thiols which permitted the determination of 2 nmol of L-cysteine (CySH). The possible mechanism for the instability of isoindole accelerated by increasing concentration of OPA was also discussed.