Light-sheet microscope for 3D high-resolution imaging of large biological samples

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Background:

The NIDA IRP Histology and Imaging Core needs a light-sheet microscope system for 3D high-resolution imaging of large biological samples. Of many applications the microscope will be used for towards the NIDA IRP mission, it will be used to determine neural mechanisms in the brain that mediate behavior in different animal models used in drug addiction research. More specifically, this microscope will be used to analyze fluorescently labeled elements (e.g. neural activity markers such as Fos and cell-type markers) in whole rat and mouse brains that were altered by drug experience. Our main use of a light-sheet microscope is to visualize fluorescently labeled elements, such as activity markers and cell type markers, in the whole brain in one image scan. This allows for rapid acquisition of labeling in all brain areas in an unbiased manner, which is important for: (a) identification of brain regions and (b) identification (through correlational analysis) of combinations of brain regions or circuits, that were not previously considered in mediating a behavior of interest.

Requirement:

The necessary requirements are:

1. For this to be effective, the microscope Requirement (1) must be able to image whole rat and mouse brains in one scan. Requirement (2) is ease of use by non-expert users, such as minimal number of adjustments and manual steps to use for imaging, and a lower probability of damage to machine. Requirement (3) is the ability to run these brains in batch mode so the microscope can be run at all times, which will greatly increase throughput of the microscope that will be in demand for many experiments by different researchers. Requirement (4) is responsiveness of tech support, which has been a severe problem in the past. Requirement (5) is to obtain optically flat images with minimal shadowing to minimize post-processing times. Requirement (6) is for high sensitivity camera acquisition to (a) minimize scan times but (b) more importantly to use lower excitation light intensities to minimize photobleaching of sample brains. Requirement (7) is for automated sample stage and imaging objectives to scan the same samples at multiple resolutions in a single imaging session. The appropriate microscope that fulfills all of these Requirements will allow us to answer critical questions about how drug experience alters the brain to produce the different behavioral components of addiction, such as drug craving, dependence, and learned associations between cues and drugs of abuse, that were not previously possible for our program to answer.

Salient Characteristics:

The characteristics in the following table are minimal criteria for a system that meets all of our requirements.

General Requirements

(1) Must be able to image whole rat and whole mouse brains in one scan.

(2) Ease of use by non-expert users, such as minimal number of adjustments and manual steps to use for imaging, and a lower probability of damage to machine.

(3) Ability to run these brains in batch mode so the microscope can be run at all times, which will greatly increase throughput of the microscope that will be in demand for many experiments by different researchers.

(4) Promise of quick responsiveness of tech support in quote, including site visit when necessary.

(5) Obtain optically flat images with minimal shadowing to minimize post-processing times.

(6) High sensitivity camera acquisition to (a) minimize scan times but (b) more importantly to use lower excitation light intensities to minimize photobleaching of sample brains.

(7) The instrument should be accompanied by a 1-year original warranty period.