The invention provides an acridone type stable isotope mercapto compound labeling reagent as well as a synthesis method and application thereof, and belongs to the field of organic small molecule stable isotope labelling technology application. The stable isotope labeling reagent is N-(4-(acridone-10-group)phenyl)-N-[d0]/[d2]maleimide. A synthesis method is characterized in that acridone and 4-acetbromanilide asepsin are subjected to substitution reaction to obtain an intermediate of I10-((4-acetamido)-phenyl)-acridone; then, the intermediate I is heated and hydrolyzed under the alkaline condition to obtain an intermediate II10-((4-amidogen)-phenyl)-acridone; the intermediate II and [d0]/[d2]maleic anhydride are subjected to acylation condensation reaction to generate a target product of N-(4-(acridone-10-group)phenyl)-N-[d0]/[d2]maleimide. The synthesis method is simple and convenient; the realization is easy; the stable isotope deuterated sulfhydryl compound labeling reagent is obtained; the isotope labeling position is stable; various kinds of sulfhydryl compounds can be selectively labeled; the advantages of high labeling product yield, high labeling reaction rate and high sulfhydryl compound selectivity are realized.