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ABSTRACT In order to develop a model for fluorescence-guided surgery (FGS), 143B human osteosarcoma cells expressing red fluorescent protein (RFP) were injected into the intramedullary cavity of the tibia in nude mice. The fluorescent areas of residual tumors after bright-light surgery (BLS) and FGS were 10.2 ± 2.4 mm2 and 0.1 ± 0.1 mm2, respectively ( p < 0.001). The BLS-treated mice and BLS + cisplatinum (CDDP)-treated mice had significant recurrence. In contrast, the FGS mice and FGS + CDDP mice had very little recurring tumor growth. Disease-free survival (DFS) in the BLS-, BLS + CDDP-, FGS-, and FGS + CDDP-treated mice was 12.5%, 37.5%, 75.0%, and 87.5%, respectively. The FGS-treated mice had a significantly higher DFS rate than the BLS-treated mice ( p = 0.021). The FGS + CDDP-treated mice had significantly higher DFS rate than the BLS + CDDP-treated mice ( p = 0.043). Although chemotherapy significantly reduced multiple metastases ( p = 0.033), there was no significant correlation between FGS and lung metastasis. FGS significantly reduced the recurrence of the primary tumor but did not reduce lung metastasis. The combination of FGS and adjuvant CDDP reduced tumor recurrence and prevented multiple metastases. FGS and adjuvant chemotherapy should be performed as early as possible in the disease to prevent both recurrence and metastatic development. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:1596-1601, 2014. [ABSTRACT FROM AUTHOR]