Method and system for imaging and analysis of a biological specimen
AssigneeThe Board of Trustees of the Leland Sanford Junior Univeristy
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.
Claims20
A method for quantifying an active neuronal population of a subject animal exposed to a stimulant, the method comprising:
- i) delivering a stimulant to the subject animal;
- ii) isolating the brain of the subject animal;
- iii) preparing the brain of the animal to produce a cleared brain by:
- a) fixing the brain with a plurality of hydrogel monomers to produce a hydrogel-fixed brain;
- b) clearing the hydrogel-fixed brain using a non-electrophoretic flow-assisted clearing device to produce a cleared brain, wherein the clearing is performed by immersing the hydrogel-fixed brain in a buffer in a sample chamber of the flow-assisted clearing device and flowing the buffer through the hydrogel-fixed brain by using a buffer circulator, wherein the buffer flow through the hydrogel-fixed brain is unidirectional;
- and
- c) contacting the cleared brain with a refractive index matching solution;
- iv) imaging the cleared brain;
- and
- v) identifying the active neuronal population.
Dependencies are inferred from the claim text.