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.

中文翻译：

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μSPIMToolset：用于选择性平面照明显微镜的软件平台

背景

选择性平面照明显微镜（SPIM）是一种荧光成像技术，可以在高时空分辨率下进行体积成像，以监测诸如斑马鱼幼体等活生物体的神经活动。使用扫描的激光束构造定制SPIM显微镜的主要挑战是各种硬件组件的控制和同步。

新方法

我们提供了一个基于广泛采用的MicroManager平台构建的开源软件μSPIMToolset，该软件为SPIM提供控制和获取功能。μSPIMToolset的一个关键优势是一系列校准程序，可以针对给定的设置优化采集，使其相对独立于显微镜的光学设计或用于构建它的硬件。

结果

μSPIMToolset可以以每秒100个平面的速度以单个细胞分辨率对整个幼虫斑马鱼大脑中的钙活动进行成像。

与现有方法的比较

SPIM的几种设计已发布，但专注于使用较慢的设置和移动平台来对发育过程进行成像，因此对功能成像的使用受到限制。相比之下，μSPIMToolset使用扫描光束可以在更高的采集频率下成像，同时最大程度地减少了样品干扰。

结论

μSPIMToolset为SPIM显微镜的控制提供了灵活的解决方案，并证明了其可用于全脑成像幼虫斑马鱼的神经活动。