SPATIALLY ENCODED BIOLOGICAL ASSAYS
The present invention provides assays and assay systems for use in spatially encoded biological assays. The invention provides an assay system comprising an assay capable of high levels of multiplexing where reagents are provided to a biological sample in defined spatial patterns; instrumentation capable of controlled delivery of reagents according to the spatial patterns; and a decoding scheme providing a readout that is digital in nature.
1 . (canceled)
2 . A system for analyzing a target biological molecule of a tissue sample, the system comprising:
a laser that is positionable relative to a location of the tissue sample and that is configured to release a mass tag or a portion of a mass tag from a region of interest in the tissue sample, wherein the tissue sample comprises a plurality of probes, and wherein a probe of the plurality of probes comprises a capture agent conjugated to the mass tag;
an imager configured to obtain an image of the tissue sample;
a mass analyzer configured to perform mass spectrometry of the released mass tag or the released portion of the mass tag; and
a processing circuit arrangement connected to the laser and the imager and further comprising software;
wherein the software comprises instructions configured to cause the processing circuit arrangement to:
display an image of the tissue sample obtained by the imager on a visual display to obtain information about the region of interest in the tissue sample based on the image of the tissue sample, wherein the region of interest is less than the entire tissue sample; and
control the laser to the region of interest in the tissue sample.
3 . The system of claim 2 , wherein the target biological molecule comprises a protein.
4 . The system of claim 2 , wherein the capture agent comprises an antibody.
5 . The system of claim 4 , wherein the antibody is conjugated to the mass tag by a linker.
6 . The system of claim 2 , wherein the mass tag comprises a chelator or a plurality of chelators.
7 . The system of claim 2 , wherein the mass tag comprises a metal or a plurality of metals.
8 . The system of claim 7 , wherein the probe further comprises a linker disposed between the capture agent and the metal or between the capture agent and each of the plurality of metals.
9 . The system of claim 2 , wherein the probe is configured to be exposed to radiation.
10 . The system of claim 9 , wherein the radiation comprises ultraviolet light.
11 . The system of claim 10 , wherein the laser is configured to provide the ultraviolet light.
12 . The system of claim 2 , wherein each probe of the plurality of probes comprises a capture agent conjugated to a mass tag.
13 . The system of claim 2 , wherein the plurality of probes includes different probes.
14 . The system of claim 2 , wherein the plurality of probes is configured to be transformed via cleavage.
15 . The system of claim 2 , wherein the probe comprises:
a primary antibody that specifically binds the target biological molecule of the tissue sample; and
a secondary antibody conjugated to the mass tag.
16 . The system of claim 2 , wherein the tissue sample is affixed to a support.
17 . The system of claim 2 , wherein the tissue sample comprises a plurality of cells.
18 . The system of claim 2 , wherein the tissue sample comprises a tissue section or a fixed tissue section.
19 . The system of claim 18 , wherein the laser is positionable atop the tissue section to access discrete portions of the tissue section.
20 . The system of claim 2 , wherein the laser is configured for laser desorption and/or laser ionization.
21 . The system of claim 2 , wherein the imager comprises a CCD array or a CMOS array.
22 . The system of claim 2 , wherein the mass analyzer is configured to perform time-of-flight mass spectrometry.
23 . The system of claim 2 , wherein the region of interest is configured for a topology of the tissue sample, configured within a grid, or configured for a group of cells, a cell layer, or a cell type in the tissue sample.
24 . The system of claim 2 , wherein the software comprises instructions configured to cause the processing circuit arrangement to obtain the information about the region of interest in the tissue sample from the image of the tissue sample.
25 . The system of claim 24 , wherein the software comprises instructions configured to cause the processing circuit arrangement to obtain the information about the region of interest in the tissue sample based on differentiated immunohistochemistry, histology, or differentiated staining of the tissue sample identified in the image of the tissue sample.
26 . The system of claim 2 , wherein the software comprises instructions configured to cause the processing circuit arrangement to receive the information about the region of interest in the tissue sample from a user of the system.
27 . The system of claim 2 , wherein the software comprises instructions configured to cause the processing circuit arrangement to communicate the information about the region of interest to the laser.
28 . The system of claim 2 , wherein the software comprises instructions configured to cause the processing circuit arrangement to obtain information about multiple regions of interest in the tissue sample based on the image of the tissue sample, and wherein the multiple regions of interest comprise at least the region of interest.
29 . The system of claim 28 , wherein:
the multiple regions of interest comprise a first region of interest and a second region of interest;
the region of interest is the first region of interest; and
the software comprises further instructions configured to cause the processing circuit arrangement to control the laser to the first region of interest and then to the second region of interest.
30 . A method of analyzing a target biological molecule of a tissue sample, the method comprising:
binding a plurality of probes to the tissue sample disposed on a surface of a support, wherein a probe of the plurality of probes comprises a capture agent conjugated to a mass tag;
releasing the mass tag or a portion of the mass tag from a region of interest in the tissue sample using a laser that is positionable relative to a location of the tissue sample; and
performing mass analysis of the released mass tag or the released portion of the mass tag using mass spectrometry to determine information about the target biological molecule at the region of interest in the tissue sample.
31 . The method of claim 30 , wherein the target biological molecule comprises a protein.
32 . The method of claim 30 , wherein the capture agent comprises an antibody.
33 . The method of claim 32 , wherein the antibody is conjugated to the mass tag by a linker.
34 . The method of claim 30 , wherein the mass tag comprises a chelator or a plurality of chelators.
35 . The method of claim 30 , wherein the mass tag comprises a metal or a plurality of metals.
36 . The method of claim 35 , wherein the probe further comprises a linker disposed between the capture agent and the metal or between the capture agent and each of the plurality of metals.
37 . The method of claim 30 , wherein the probe is configured to be exposed to radiation.
38 . The method of claim 37 , wherein the radiation comprises ultraviolet light.
39 . The method of claim 30 , wherein each probe of the plurality of probes comprises a capture agent conjugated to a mass tag.
40 . The method of claim 30 , wherein the plurality of probes includes different probes.
41 . The method of claim 30 , wherein the plurality of probes is configured to be transformed via cleavage.
42 . The method of claim 30 , wherein the probe comprises:
a primary antibody that specifically binds the target biological molecule of the tissue sample; and
a secondary antibody conjugated to the mass tag.
43 . The method of claim 30 , wherein the tissue sample is affixed to a support.
44 . The method of claim 30 , wherein the tissue sample comprises a plurality of cells.
45 . The method of claim 30 , wherein the tissue sample comprises a tissue section or a fixed tissue section.
46 . The method of claim 45 , wherein the laser is positionable atop the tissue section to access discrete portions of the tissue section.
47 . The method of claim 30 , wherein the releasing comprises laser desorption and/or laser ionization.
48 . The method of claim 30 , wherein the performing comprises time-of-flight mass spectrometry.
49 . The method of claim 30 , wherein the region of interest is configured for a topology of the tissue sample, configured within a grid, or configured for a group of cells, a cell layer, or a cell type in the tissue sample.
50 . The method of claim 30 , further comprising:
obtaining an image of the tissue sample.
51 . The method of claim 50 , further comprising:
obtaining information about the region of interest in the tissue sample from the image of the tissue sample.
52 . The method of claim 51 , wherein the information about the region of interest in the tissue sample is based on differentiated immunohistochemistry, histology, or differentiated staining of the tissue sample identified in the image of the tissue sample.
53 . The method of claim 51 , wherein the information about the region of interest in the tissue sample is provided from a user.
54 . The method of claim 51 , wherein the information about the region of interest is provided to the laser.
55 . The method of claim 30 , further comprising:
releasing the mass tag or a portion of the mass tag from multiple regions of interest in the tissue sample using the laser, wherein the multiple regions of interest comprise at least the region of interest.
56 . The method of claim 55 , wherein:
the multiple regions of interest comprise a first region of interest and a second region of interest;
the region of interest is the first region of interest; and
the releasing comprises using the laser at the first region of interest and then the second region of interest.