Abstract: Nanobodies have become popular targeting molecules in the field of molecular imaging due to their excellent targeting ability, superior stability, good solubility, and enhanced tissue penetration ability. A variety of nanobody-based molecular imaging probes have shown great promise for noninvasive diagnosis of human malignancies in preclinical and clinical settings. This review mainly introduces the latest research progress on nanobody-based molecular imaging probes in tumors, and further discusses the challenges and strategies for future clinical translation.