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The accurate determination of specific biomolecules and their monitoring in complex biological systems will help us understand their role in the body. In comparison with cumulative fluorescent probes, responsive fluorescent probes exhibit a higher level of sensitivity for the detection of biomolecules. Recently, responsive fluorescent probes based on 1,8-naphthalimide have been used widely in the biomedical field because of their excellent luminescence performance. This review focuses on the progress in research on responsive fluorescent probes obtained by modifying the C4-position of 1,8-naphthalimide to achieve biological molecule detection over the last five years. Based on the differences in species, the corresponding recognition groups and responsive probes were classified systematically and thoroughly, and the significance of the probes used specifically for biological molecule detection was discussed. Finally, we discussed the prospect of developing this type of probe, to make it a powerful diagnostic tool in clinical research. [Display omitted] • The research progress of responsive fluorescent probes obtained by modifying the C4-position of 1.8-naphthalimide. • The corresponding recognition groups and responsive probes were classified systematically and in detail. • We also introduced the significance of responsive fluorescent probes in specific biological detection in recent five years. • We discussed the prospect of developing this type of probe, to make it a powerful diagnostic tool in clinical research. [ABSTRACT FROM AUTHOR]