Method and System for Imaging and Analysis of a Biological Specimen
AssigneeThe Board of Trustees of the Leland Stanford Junior University
InventorsKarl A. Deisseroth, William E. Allen, Brian Hsueh, Li Ye
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Abstract
The present disclosure provides methods of preparing a biological specimen for imaging analysis, comprising fixing and clearing the biological specimen and subsequently analyzing the cleared biological specimen using microscopy. Also included are methods of quantifying cells, for example, active populations of cells in response to a stimulant. The present disclosure also provides devices for practicing the described methods. A flow-assisted clearing device provides rapid clearing of hydrogel-embedded biological specimens without the need of specialized equipment such as electrophoresis or perfusion devices.
Claims37
A method of preparing a biological specimen for imaging analysis, the method comprising:
- i) fixing the biological specimen with a plurality of hydrogel monomers to produce a hydrogel-fixed specimen;
- ii) clearing the hydrogel-fixed specimen using a flow-assisted clearing device to produce a cleared specimen, wherein the clearing is performed by:
- a) immersing the hydrogel-fixed specimen in a buffer;
- b) flowing the buffer through the hydrogel-fixed specimen, wherein the buffer flow is unidirectional;
- and
- iii) contacting the cleared specimen with a refractive index matching solution prior to imaging analysis.
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A flow-assisted clearing device, the device comprising:
- a sample chamber comprising an inlet and an outlet, wherein the inlet delivers the buffer into the sample chamber and the outlet draws the buffer out of the sample chamber;
- a sample holder, wherein the sample holder can be removably placed inside the sample chamber, and wherein the sample holder allows the buffer to flow through;
- and
- a buffer circulator.
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Dependencies are inferred from the claim text.