Immunodepletion of Abundant Peptides for Mass Spectrometry
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Solution Overview
Problem
In mass spectrometry, the detection of all analytes in complex samples is hindered by ionization suppression, differences in ionization potential, and the limited dynamic range of detectors, particularly in proteomics where high abundance proteins interfere with the detection of lower abundance proteins, leading to incomplete sequence coverage and incorrect protein identification.
Innovation Solution
A method involving the physical removal of highly abundant and well-ionizing peptides from samples before mass spectral analysis using immunodepletion with antibodies specific to these peptides, which are identified by cleaving proteins into peptides and ranking them by ionization potential, allowing for the selective removal of problematic peptides.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Object-affected harmful factors
If high abundance proteins are removed by affinity depletion prior to digestion, then ionization suppression is reduced, but non-specific removal of low abundance proteins occurs
Solution Approach 1:
The invention segments the protein removal process by digesting proteins into peptides first, then selectively removing specific peptides using antibodies. This allows targeted depletion of high abundance protein peptides without affecting intact low abundance proteins, resolving the non-specific removal issue while maintaining ionization suppression reduction benefits
Solution Approach 2:
The invention performs preliminary enzymatic digestion of proteins into peptides before the depletion step. This preliminary action enables subsequent antibody-based selective removal of specific peptide sequences from high abundance proteins, preventing the non-specific removal problem that occurs with intact protein affinity depletion
2Device complexity
If peptide masses from contaminant proteins are removed from peak lists, then database searching is simplified, but ionization suppression and detector saturation remain unresolved
Solution Approach 1:
The invention performs preliminary removal of contaminant and high abundance protein peptides from the sample using antibody-based immunodepletion before mass spectrometry analysis. This physical removal prevents ionization suppression and detector saturation during data acquisition, addressing the fundamental detection problems that data processing approaches cannot resolve
Solution Approach 2:
The invention extracts and removes problematic peptide components (contaminant and high abundance protein peptides) from the complex peptide mixture using specific antibodies. This extraction eliminates the source of ionization suppression and detector saturation, allowing better detection of low abundance protein peptides
3Quantity of substance
If intact high abundance proteins are removed by affinity depletion, then dynamic range issues are reduced, but low abundance proteins are non-specifically depleted
Solution Approach 1:
The invention segments proteins into peptides through enzymatic digestion, then applies antibody-based depletion to specific peptide sequences. This segmentation allows selective removal of high abundance protein peptides while preserving low abundance proteins, as antibodies can be designed to target unique peptide sequences without cross-reacting with peptides from low abundance proteins
Solution Approach 2:
The invention applies local quality by using antibodies with high specificity to target only particular peptide sequences from high abundance proteins. This ensures that depletion occurs at the specific peptide level rather than affecting all proteins globally, maintaining reliability by preventing non-specific removal of low abundance proteins
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 enhances the detection of low abundance proteins by reducing ionization suppression and improving spectral quality, enabling more accurate protein identification and detection at higher sensitivity levels, such as the femtomolar or attomolar level.
Implementation Method 1
the peptides are removed by immunodepletion
Implementation Method 2
one or more proteins in the sample are cleaved to peptides by an enzymatic or chemical method
Data Source
AI summary
The present invention relates, e.g., to a method for pre-processing a sample for mass spectral analysis, comprising cleaving proteins in the sample to peptides and immunodepleting highly abundant and/or well-ionizing and/or proteotypic peptides from the sample. Also described are methods for identifying well-ionizing peptides for use in this and other methods; analytic (diagnostic) methods using antibodies against highly ionizable peptides from a protein target of interest; and compositions, kits and devices comprising antibodies of the invention.