Molecular　imaging　is　a　non-invasive　method　to　image　and　analyze　the　concentration　and　activity　of　functional　biomolecules　in　cells　or　in　vivo　at　molecular　level,　and　plays　an　increasing　role　in　deep　understanding　of　biological　processes,　early　and　accurate　diagnosis　of　diseases,　and　evaluation　of　treatment.　Nowadays,　numerous　novel　molecular　imaging　probes　have　been　developed,　involving　every　biomedical　imaging　modality,　such　as　optical　imaging,　photoacoustic　imaging,　magnetic　resonance　imaging,　single-photon-emission　computed　tomography,　and　positron　emission　tomography.　In　this　review,　we　summarize　the　development　of　current　state-of-the-art　molecular　imaging　probes.　We　introduce　the　design　strategies　of　molecular　probes　and　detailed　imaging　modalities,　and　highlight　the　properties　of　probes　and　biomedical　imaging　applications　in　cells　and　in　vivo,　including　disease　diagnosis,　drug　tracking,　and　imaging-guided　surgery.　Then　we　discuss　the　perspectives　and　challenges　in　this　emerging　field.　We　expect　this　review　could　inspire　more　effective　molecular　imaging　probes　to　be　developed,　achieving　the　goal　towards　clinical　practices.　[Figure　not　available:　see　fulltext.]　©　2023,　Science　China　Press.