A simple and accurate high throughput evaluation method for larval zebrafish xenografts

The advantages of short assay duration, low cost, and high-throughput make larval zebrafish xenografts a popular in vivo model in cancer research, especially in large-scale screening of anti-tumor drug and target identification. Recently, the great potential of larval zebrafish xenografts in precision cancer therapy has also been demonstrated. Establishing an accurate high-throughput analytical method is now the main challenge of this model. In this study, we, for the first time, compared the current mainstream analytical methods. Calculating the volume of xenografts based on confocal imaging was accurate but labor-intensive and time-consuming. Although measuring tumor area is an effective method to achieve high-throughput analysis, its accuracy was low. Based on our experience, we found that analyzing the maximum fluorescence intensity (mFI) avoided manual error, reduced the intra-group variability, and enable high-throughput analysis. Moreover, mFI accurately reflected the change of xenograft size and significantly reduced the sample size required for one test. This study provided a simple and accurate high-throughput analytical method for assessing larval zebrafish xenografts, and the development of an automatic high-throughput analysis platform based on this method will further promote anti-tumor drug development and precision cancer therapy.