The invention discloses a long-wavelength fluorescence probe, and a preparation method and application thereof. The preparation method comprises the following steps: reacting 1,3,5,7-tetramethyl-8-phenyl-(2'-nitro)-boron difluoride-dimethyl pyrrole and benzaldehyde to generate a conjugated system with a larger 3,5-distyryl group, reducing -NO2 into -NH2 with zinc powder, carrying out addition reaction with maleic anhydride, and reacting with acetic anhydride to obtain the long-wavelength fluorescence probe 1,7-dimethyl-3,5-distyryl-8-phenyl-(2'-maleimido)-boron difluoride dipyrromethene. The preparation method is simple; the prepared long wavelength fluorescence probe has the advantages of small influence by environmental pH value, favorable selectivity for sulfhydryl compounds, and high sensitivity; the autofluorescence intensity of the fluorescence probe is very weak, but can be enhanced after reaction with sulfhydryl group, and thus, the fluorescence probe can be used for detecting sulfhydryl compounds; and the fluorescence probe can generate small damage to organisms, and thus, has wide application prospects in the fields of biological tissues, cell fluorescence imaging and the like.