SRM/MRM Assay for FASN Protein Quantitation
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Solution Overview
Problem
Current methods for analyzing the Fatty Acid Synthase (FASN) protein in formalin-fixed cancer tissue samples are inefficient due to the difficulty in detecting suitable peptides for mass spectrometry-based SRM/MRM assays, leading to inaccurate results and the inability to predict suitable peptides for use in these assays.
Innovation Solution
Development of a mass spectrometry-based SRM/MRM assay using specific peptides derived from the FASN protein, which are identified through experimental determination of their ionization properties and ability to survive sample preparation conditions, allowing for relative and absolute quantitation of the protein levels in formalin-fixed tissue samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional methods are used to analyze FASN protein in formalin-fixed tissue, then the analysis can be performed on readily available samples, but the detection accuracy and quantitation precision are insufficient
Solution Approach 1:
The patent applies preliminary action by experimentally determining the ionization properties and sample preparation survival of candidate peptides before developing the SRM/MRM assay. This pre-screening process identifies peptides that will actually be detectable in formalin-fixed tissue, ensuring the assay targets suitable analytes from the outset rather than attempting to detect unsuitable peptides during the actual analysis.
Solution Approach 2:
The patent employs parameter changes by optimizing multiple assay parameters including selecting specific peptide sequences with appropriate ionization characteristics, determining optimal fragmentation transitions, establishing suitable mass spectrometry acquisition parameters, and optimizing sample preparation conditions to preserve peptide integrity while maximizing detectability.
2Reliability
If SRM/MRM assay is developed without experimental validation of peptide properties, then the assay development would be faster, but the results would be inaccurate and unreliable
Solution Approach 1:
The patent performs preliminary experimental validation of peptide ionization properties and sample preparation stability before finalizing the SRM/MRM assay. This upfront investment in experimental characterization ensures that only peptides with proven detectability and stability are included in the final assay, guaranteeing reliable and accurate results without requiring later troubleshooting or rework.
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
The assay enables accurate and precise measurement of FASN protein levels in cancer tissue samples, aiding in cancer diagnosis and determining appropriate therapeutic agents, thereby facilitating personalized medicine approaches.
Implementation Method 1
mass spectrometry-based Selected Reaction Monitoring (SRM) assay
Implementation Method 2
measuring the amount of the FASN protein in a given protein preparation obtained from a biological sample by mass spectrometry
Data Source
AI summary
Specific peptides, and derived ionization characteristics of the peptides, from the Fatty acid synthase (FASN) protein are provided that are particularly advantageous for quantifying the FASN protein directly in biological samples that have been fixed in formalin by the method of Selected Reaction Monitoring (SRM) mass spectrometry, or what can also be termed as Multiple Reaction Monitoring (MRM) mass spectrometry. Such biological samples are chemically preserved and fixed and are selected from tissues and cells treated with formaldehyde containing agents/fixatives including formalin-fixed tissue/cells, formalin-fixed/paraffin embedded (FFPE) tissue/cells, FFPE tissue blocks and cells from those blocks, and tissue culture cells that have been formalin fixed and or paraffin embedded. A protein sample is prepared from said biological sample using the Liquid Tissue™ reagents and protocol and the FASN protein is quantitated in the Liquid Tissue™ sample by the method of SRM/MRM mass spectrometry by quantitating in the protein sample at least one or more of the peptides described. These peptides can be quantitated if they reside in a modified or an unmodified form. An example of a modified form of an FASN peptide is phosphorylation of a tyrosine, threonine, serine, and/or other amino acid residues within the peptide sequence.