Compositions and processes for improved mass spectrometry analysis
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Publication Number
US-9310378-B2
Publication Date
2016-04-12
Expiration Date
Abstract
The invention provides a novel additive for improved analysis by mass spectrometry. More specifically, ascorbic acid has been found to reduce or eliminate the presence of adducts commonly present in mass spectra. The improved processes and compositions of the invention allow for increased accuracy, sensitivity and throughput for samples analyzed by mass spectrometry.
Core Innovation
The described invention relates to a Matrix-Assisted Laser Desorption/Ionization mass spectrometry substrate for nucleic acids that uses a MALDI matrix and an additive to reduce adduct formation under MALDI conditions. The MALDI matrix is selected from 3-hydroxypicolinic acid, di-ammoniumcitrate, or a mixture of 3-hydroxypicolinic acid and di-ammoniumcitrate. The substrate includes a support with an array of spots, some or all of which comprise the nucleic-acid MALDI matrix and a specified additive.
The additive comprises ascorbic acid, or a salt, tautomer or analog thereof, wherein the amount of the additive reduces the amount of an adduct formed under MALDI mass spectrometry conditions. The analog is defined according to a structural formula with substituent definitions for R1 and R2, and for R3 and R4, including options such as OH, halogen, azido, cyano, alkyl, and optionally substituted aryl or heterocyclic rings. This combination is applied to spots on a substrate to suppress undesired adducts associated with MALDI measurement of nucleic acids.
The invention also includes a mass spectrometry system that performs Matrix-Assisted Laser Desorption/Ionization Time-of-Flight mass spectrometry and uses the same MALDI substrate structure. The system includes a mass spectrometer for MALDI-TOF, and the substrate comprises the nucleic-acid MALDI matrix and the ascorbic-acid additive in an amount that reduces adduct formation under MALDI conditions. The system and substrate are defined for use with an array of spots, with certain embodiments specifying support formats and spot characteristics.
The problem being solved is alkali and ammonia adduct formation in MALDI/electrospray-compatible mass spectrometry, which can reduce spectral accuracy and adversely affect nucleic-acid measurements. The disclosed approach uses an ascorbic-acid additive with nucleic-acid MALDI matrix choices to reduce adduct formation, including suppression of ammonia-related adduct peaks and reduction of false positive genotype calls. The document also reports matrix stability over time while preserving signal characteristics.
Claims Coverage
The partial content provided includes two independent claims: a MALDI mass spectrometry substrate for nucleic acids and a mass spectrometry system that incorporates such a substrate. Each independent claim centers on an array-of-spots MALDI substrate using a nucleic-acid matrix (3-hydroxypicolinic acid and/or di-ammoniumcitrate) together with an ascorbic-acid-based additive that reduces adduct formation under MALDI conditions, where the ascorbic-acid analog is defined by a specific formula.
Nucleic-acid MALDI array with adduct-reducing ascorbic acid additive
A Matrix-Assisted Laser Desorption/Ionization mass spectrometry substrate with an array of spots on a support, where some or all spots comprise a nucleic-acid MALDI matrix selected from 3-hydroxypicolinic acid, di-ammoniumcitrate, or a mixture of 3-hydroxypicolinic acid and di-ammoniumcitrate, and an additive that comprises ascorbic acid or a salt, tautomer or analog thereof in an amount that reduces the amount of an adduct formed under Matrix-Assisted Laser Desorption/Ionization mass spectrometry conditions, wherein the analog of ascorbic acid is according to a specified formula with defined substituent groups for R1, R2, R3, and R4.
MALDI-TOF mass spectrometry system using the nucleic-acid adduct-reducing substrate
A mass spectrometry system comprising a mass spectrometer for performing Matrix-Assisted Laser Desorption/Ionization Time-of-Flight mass spectrometry and a Matrix-Assisted Laser Desorption/Ionization mass spectrometry substrate, where the substrate includes an array of spots and some or all spots comprise a nucleic-acid MALDI matrix selected from 3-hydroxypicolinic acid, di-ammoniumcitrate, or a mixture of 3-hydroxypicolinic acid and di-ammoniumcitrate, and an additive that comprises ascorbic acid or a salt, tautomer or analog thereof in an amount that reduces the amount of an adduct formed under Matrix-Assisted Laser Desorption/Ionization mass spectrometry conditions, wherein the analog is defined according to the specified formula with defined substituent groups for R1, R2, R3, and R4.
Across both independent claims, the central coverage is an array-of-spots MALDI substrate for nucleic acids that combines a defined nucleic-acid MALDI matrix with an ascorbic-acid-based additive in an amount that reduces adduct formation under MALDI conditions; the second independent claim further ties this substrate to a MALDI-TOF mass spectrometry system.
Stated Advantages
Improves spectral accuracy by reducing adduct formation in MALDI mass spectrometry conditions.
Improves sensitivity, including signal-to-noise ratio (SNR) and adduct score performance.
Improves throughput for nucleic-acid measurements by reducing unwanted adducts and associated measurement issues.
Reduces false positive genotype calls associated with adduct peaks.
Documented Applications
Mass spectrometry of nucleic acids, including nucleic-acid genotyping in contexts such as Sequenom iPLEX, MassARRAY, and SpectroChip (platform ions/ion analyzers are mentioned).
Use with nucleic acids such as DNA, RNA, and oligonucleotides, including primer extension and genotyping contexts.
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