µSPIM toolset: a software platform for selective plane illumination microscopy

Background: Selective Plane Illumination Microscopy (SPIM) is a fluorescence imaging technique that allows volumetric imaging at high spatio-temporal resolution to monitor neural activity in live organisms such as larval zebrafish. A major challenge in the construction of a custom SPIM microscope using a scanned laser beam is the control and synchronization of the various hardware components. New Method: We present an open-source software, µSPIM Toolset, built around the widely adopted MicroManager platform, that provides control and acquisition functionality for a SPIM. A key advantage of µSPIM Toolset is a series of calibration procedures that optimize acquisition for a given set-up, making it relatively independent of the optical design of the microscope or the hardware used to build it. Results: µSPIM Toolset allows imaging of calcium activity throughout the brain of larval zebrafish at rates of 100 planes per second with single cell resolution. Comparison with Existing Methods: Several designs of SPIM have been published but are focused on imaging of developmental processes using a slower setup with a moving stage and therefore have limited use for functional imaging. In comparison, µSPIM Toolset uses a scanned beam to allow imaging at higher acquisition frequencies while minimizing disturbance of the sample. Conclusions: The µSPIM Toolset provides a flexible solution for the control of SPIM microscopes and demonstrated its utility for brain-wide imaging of neural activity in larval zebrafish.