Zebrafish (Danio rerio) larvae are widely used to study otic functions because they possess all five typical vertebrate senses including hearing and balance. Powerful genetic tools and the transparent body of the embryo and larva also make zebrafish a unique vertebrate model to study otic development. Due to its small larval size and moisture requirement during experiments, accurately acquiring the vestibulo-ocular reflex (VOR) of zebrafish larva is challenging. In this report, a new VOR testing device has been developed for quantifying linear VOR (LVOR) in zebrafish larva, evoked by the head motion about the earth horizontal axis. The system has a newly designed larva-shaped chamber, by which live fish can be steadily held without anesthesia, and the system is more compact and easier to use than its predecessors. To demonstrate the efficacy of the system, the LVORs in wild-type (WT), dlx3b and dlx4b morphant zebrafish larvae were measured and the results showed that LVOR amplitudes were consistent with the morphological changes of otoliths induced by morpholino oligonucleotides (MO). Our study represents an important advance to obtain VOR and predict the vestibular conditions in zebrafish.