Photocleavable Mass-Tag Probes for High-Multiplex Tissue MSI
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Solution Overview
Problem
Current methods for multiplexed detection and mapping of biomolecules in tissues, such as immunohistochemistry and in situ hybridization, are limited by the inability to simultaneously detect multiple biomarkers due to spectral overlap in fluorescence microscopy and poor sensitivity in mass spectrometric imaging (MSI).
Innovation Solution
The use of photocleavable mass-tag reagents attached to probes like antibodies and nucleic acids, which allow for multiplex immunohistochemistry and in situ hybridization, coupled with mass spectrometric imaging (MSI) as the detection method, enabling the simultaneous imaging of multiple biomarkers on a single slide.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If fluorescence microscopy is used for multiplexed detection of biomarkers, then visualization of spatial distribution is achieved, but the number of simultaneously detectable biomarkers is limited to 3-5 due to spectral overlap
Solution Approach 1:
The patent replaces the optical detection system (fluorescence microscopy) with a mass spectrometric detection system. Instead of using fluorophores that emit light at different wavelengths, the invention uses mass tags with distinct mass-to-charge ratios that are detected by mass spectrometry. This substitution eliminates spectral overlap issues and enables simultaneous detection of 5 or more biomarkers, as mass spectrometry can resolve many more distinct mass values than optical wavelengths can be distinguished.
Solution Approach 2:
The invention changes the detection parameter from optical wavelength (fluorescence emission spectrum) to mass-to-charge ratio. By using mass tags with carefully selected distinct masses rather than fluorophores with distinct emission wavelengths, the system overcomes the fundamental limitation of spectral overlap. The mass tags are designed to have masses that can be clearly distinguished by the mass spectrometer, allowing for higher multiplexing capacity.
2Measurement precision
If mass spectrometric imaging is used for detection, then sensitivity is improved, but the ability to detect multiple biomarkers simultaneously is reduced
Solution Approach 1:
The patent segments the detection system into two independent components: (1) highly sensitive mass tag labels that provide strong mass spectrometric signals for each biomarker, and (2) a mass spectrometric imaging platform that can detect multiple distinct mass values simultaneously. The mass tags are designed with high ionization efficiency and distinct mass-to-charge ratios, allowing each biomarker to be detected with high sensitivity while maintaining the ability to distinguish multiple different mass tags in the same sample.
Solution Approach 2:
The mass spectrometric imaging platform serves multiple functions simultaneously: it detects multiple different mass tags with high sensitivity, provides spatial mapping information, and maintains the ability to distinguish between different biomarkers based on their unique mass-to-charge ratios. The system is universally applicable to detect any number of biomarkers as long as distinct mass tags are used, making it more versatile than fluorescence microscopy for multiplexed detection.
3Quantity of substance
If photocleavable mass-tag reagents are used, then multiplexing capability is improved to 5 or more biomarkers, but the complexity of probe labeling is increased
Solution Approach 1:
The patent uses photocleavable linkers as intermediary components that connect the mass tags to the biomolecule targets. These linkers serve as temporary carriers that allow the mass tags to be attached to various probes (antibodies, nucleic acids, etc.) through standard conjugation chemistry. The photocleavable nature of the linkers enables controlled release of the mass tags during mass spectrometric analysis, simplifying the detection process while maintaining the ability to multiplex multiple biomarkers simultaneously.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach allows for the co-detection of 5 or more different types of biomarkers in a tissue sample, overcoming the multiplexing limits of traditional fluorescence microscopy and achieving high sensitivity in MSI, thereby facilitating comprehensive biomarker analysis in tissues.
Implementation Method 1
photocleavable mass-tag reagents which are attached to probes such as antibodies and nucleic acids
Data Source
AI summary
The field of this invention relates to immunohistochemistry (IHC) and in situ hybridization (ISH) for the targeted detection and mapping of biomolecules (e.g., proteins and miRNAs) in tissues or cells for example, for research use and for clinical use such by pathologists (e.g., biomarker analyses of a resected tumor or tumor biopsy). In particular, the use of mass spectrometric imaging (MSI) as a mode to detect and map the biomolecules in tissues or cells for example. More specifically, the field of this invention relates to photocleavable mass-tag reagents which are attached to probes such as antibodies and nucleic acids and used to achieve multiplex immunohistochemistry and in situ hybridization, with MSI as the mode of detection/readout. Probe types other than antibodies and nucleic acids are also covered in the field of invention, including but not limited to carbohydrate-binding proteins (e.g., lectins), receptors and ligands. Finally, the field of the invention also encompasses multi-omic MSI procedures, where MSI of photocleavable mass-tag probes is combined with other modes of MSI, such as direct label-free MSI of endogenous biomolecules from the biospecimen (e.g., tissue), whereby said biomolecules can be intact or digested (e.g., chemically digested or by enzyme).