Patent Description:
Mass spectrometry relies upon the measurement of physical values that can be related to the mass-to-charge ratio (m/z) to determine a mass of an ionic species or a compound within a sample. High mass accuracy requires calibration of the measured physical values against species of known m/z or mass. Calibration is generally accomplished with the use of a calibration mixture that produces multiple ionic species of known m/z. However, the choice of ions suitable for use in a calibration mixture may be limited.

From the foregoing it will be appreciated that a need exists for improved calibration methods for mass spectrometry.

<CIT> describes a method of calibrating a reflectron time-of-flight mass spectrometer using a spectrum generated by fragment ions wherein the mass of the fragment ion is assigned using the mono-isotopic peak only.

<NPL>examines the reliability of FTICR in obtaining exact mass-measurements for a series of compounds using several different calibration methods and data processing algorithms.

In a first aspect of the present invention, there is provided a method as set out in claim <NUM>.

In various embodiments of the first aspect, the mass analyzer can include a Fourier transform mass analyzer. In specific embodiments, the instrument parameter can be selected from the group consisting of frequency range for an image current, a digitizing rate, a filter bandwidth for an image current, and any combination thereof.

In various embodiments of the first aspect, the mass analyzer can include a quadrupole mass analyzer or a quadrupole ion trap mass analyzer. In specific embodiments, the instrument parameter can be selected from the group consisting of an RF voltage, a DC voltage, and any combination thereof.

In various embodiments of the first aspect, the mass analyzer can include a time-of-flight mass analyzer. In specific embodiments, the instrument parameter can be selected from the group consisting of an acquisition time window, a flight tube clearing pulse time, and any combination thereof.

In various embodiments of the first aspect, the sample can be provided by a gas chromatographic instrument.

In various embodiments of the first aspect, the sample can be provided by a liquid chromatographic instrument.

In various embodiments of the first aspect, the method can further include providing a second sample including a second sample ion species to the mass spectrometer; operating the mass spectrometer using the modified instrument parameter from the calibration; and measuring a third mass related physical value for the second sample ion species.

In specific embodiments, the method can further include shifting the calibration curve based on a measured mass-to-charge ratio of a third sample ion species within the second sample to account for scan specific drifts in the calibration curve, wherein the instrument parameter is not changed based on the shifting of the calibration curve. In specific embodiments, the second sample ion species can have a mass-to-charge ratio within the range of the mass-to-charge ratio of the first sample ion species and the mass-to-charge ratio of at least one of the calibrant ion species.

In various embodiments of the first aspect, the first sample ion species can have a mass-to-charge ratio above the range of the mass-to-charge ratios of the calibrant ion species.

In various embodiments of the first aspect, the first sample ion species can have a mass-to-charge ratio below the range of the mass-to-charge ratios of the calibrant ion species.

In a second aspect, there is provided a mass spectrometer as set out in claim <NUM>.

In various embodiments of the second aspect, the mass analyzer can include a Fourier transform mass analyzer. In specific embodiments, modifying the operation of the mass analyzer can include modifying a frequency range for an image current, a digitizing rate, a filter bandwidth for an image current, and any combination thereof.

In various embodiments of the second aspect, the mass analyzer can include a quadrupole mass analyzer or a quadrupole ion trap mass analyzer. In specific embodiments, modifying the operation of the mass analyzer can include modifying an RF voltage, a DC voltage, or any combination thereof.

In various embodiments of the second aspect, the mass analyzer can include a time-of-flight mass analyzer. In specific embodiments, modifying the operation of the mass analyzer can include modifying an acquisition time window, a flight tube clearing pulse time, or any combination thereof.

In various embodiments of the second aspect, the mass spectrometer can further include a gas chromatograph for supplying a sample to the ion source.

In various embodiments of the second aspect, the mass spectrometer can further include a liquid chromatograph for supplying a sample to the ion source.

In various embodiments of the second aspect, the controller can be further configured to operate the ion source to provide a second sample including a second sample ion species to the mass spectrometer; operate the mass spectrometer using the modified instrument parameter from the calibration; and obtain a third set of mass related physical values for the second sample ion species. In specific embodiments, the controller can be further configured to shift the calibration curve based on a measured mass-to-charge ratio of a third sample ion species within the second sample to account for scan specific drift in the calibration curve, wherein the operation of the mass analyzer is not changed based on the shifting of the calibration curve.

In various embodiments of the second aspect, first sample ion species can have a mass-to-charge ratio above the range of the mass-to-charge ratios of the calibrant ion species.

In various embodiments of the second aspect, first sample ion species can have a mass-to-charge ratio below the range of the mass-to-charge ratios of the calibrant ion species.

Embodiments of systems and methods for mass calibration are described herein.

In this detailed description of the various embodiments, for purposes of explanation, numerous specific details are set forth to provide a thorough understanding of the embodiments disclosed. One skilled in the art will appreciate, however, that these various embodiments may be practiced with or without these specific details, while remaining within the scope of the invention, which is defined by the claims. In other instances, structures and devices are shown in block diagram form.

Unless described otherwise, all technical and scientific terms used herein have a meaning as is commonly understood by one of ordinary skill in the art to which the various embodiments described herein belongs.

The mass spectrometry platform <NUM> includes components as displayed in the block diagram of <FIG>. In various embodiments, elements of <FIG> can be incorporated into mass spectrometry platform <NUM>. According to various embodiments, mass spectrometer <NUM> includes an ion source <NUM>, a mass analyzer <NUM>, an ion detector <NUM>, and a controller <NUM>.

In various embodiments, the ion source <NUM> generates a plurality of ions from a sample. The ion source can include, but is not limited to, a matrix assisted laser desorption/ionization (MALDI) source, electrospray ionization (ESI) source, atmospheric pressure chemical ionization (APCI) source, atmospheric pressure photoionization source (APPI), inductively coupled plasma (ICP) source, electron ionization source, chemical ionization source, photoionization source, glow discharge ionization source, thermospray ionization source, and the like.

In various embodiments, the mass analyzer <NUM> separates ions based on a mass-to-charge ratio of the ions. For example, the mass analyzer <NUM> can include a quadrupole mass filter analyzer, a quadrupole ion trap analyzer, a time-of-flight (TOF) analyzer, an electrostatic trap (e.g., Orbitrap) mass analyzer, Fourier transform ion cyclotron resonance (FT-ICR) mass analyzer, and the like. In various embodiments, the mass analyzer <NUM> can also be configured to fragment the ions using collision induced dissociation (CID) electron transfer dissociation (ETD), electron capture dissociation (ECD), photo induced dissociation (PID), surface induced dissociation (SID), and the like, and further separate the fragmented ions based on the mass-to-charge ratio.

In various embodiments, the ion detector <NUM> detects ions. For example, the ion detector <NUM> can include an electron multiplier, a Faraday cup, and the like. Ions leaving the mass analyzer can be detected by the ion detector. In various embodiments, the ion detector can be quantitative, such that an accurate count of the ions can be determined.

In various embodiments, the controller <NUM> can communicate with the ion source <NUM>, the mass analyzer <NUM>, and the ion detector <NUM>. For example, the controller <NUM> can configure the ion source or enable/disable the ion source. Additionally, the controller <NUM> can configure the mass analyzer <NUM> to select a particular mass range to detect. Further, the controller <NUM> can adjust the sensitivity of the ion detector <NUM>, such as by adjusting the gain. Additionally, the controller <NUM> can adjust the polarity of the ion detector <NUM> based on the polarity of the ions being detected. For example, the ion detector <NUM> can be configured to detect positive ions or be configured to detected negative ions.

<FIG> is a graph illustrating a calibration curve when using only low mass ions from a calibrant mixture and when using the low mass ions in a calibration mixture in combination with a high mass ion from a sample. <FIG> is an enlargement of the circled region around the point corresponding to the high mass ion. The solid line is a fit of the standard calibrant ions to equation <NUM>. The standard calibrant ions are <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM>. At m/z <NUM>, the fit is significantly off. The dashed line is a fit of the standard calibrant ions plus the higher mass ion to equation <NUM>. This fit has improved accuracy over the range from low mass to high mass.

<FIG> is a flow diagram for calibrating a mass analyzer, such as mass analyzer <NUM> of <FIG>. At <NUM>, ions are generated from a calibrant mixture. In various embodiments, the calibrant mixture includes a multiple species of known mass which covers a range of masses. The ions of the calibrant mixture are analyzed by the mass analyzer at <NUM> and a set of physical values related to the m/z is measured. Depending on the type of mass analyzer, the mass related physical value can be a frequency, a time, a voltage, or the like.

In various embodiments, when using a Fourier transform mass analyzer, such as an Orbitrap or a Fourier Transform Ion Cyclotron Resonance (FTICR) mass analyzer, the m/z ratio can be related to a measured frequencyf of an induced current caused by the oscillation of the ion in the mass analyzer. The relationship can be described using Equation <NUM>, <NUM>, or <NUM> for an Orbitrap or Equation <NUM> for a FTICR mass analyzer. <MAT> <MAT> <MAT> <MAT>.

In various embodiments, when using a quadrupole mass analyzer, such as a quadrupole mass filter or a quadrupole ion trap, the m/z ratio can be related to a voltage V applied to the quadrupole. Additionally, there can be a time delay between when a voltage is set and when the voltage is applied to the quadrupole and the ions respond to the change in the field. When scanning across a mass range, it may be necessary to correct for the response delay. In various embodiments, this can be accomplished by introducing a voltage offset B which may be a function of scan rate. The relationship can be described using Equation <NUM> and the correction for the response delay can be described using Equation <NUM>. <MAT> <MAT>.

In various embodiments, when using a time-of-flight mass analyzer the m/z ratio can be related to a time t it takes for the ion to travel the flight path. The relationship can be described using Equations <NUM> and <NUM>. <MAT> <MAT>.

In various embodiments, a calibration for the physical value is determined from the measured values and the known masses, as indicated at <NUM>.

At <NUM>, ions are generated from a sample containing at least one species of known mass. The species of known mass can have a mass outside of the range covered by the calibrant, either lower or higher or even in between. In various embodiments, the sample can be a calibration sample used to calibrate a chromatograph, such as a gas chromatograph or a liquid chromatograph. In this way, calibration of the mass analyzer can occur concurrently with calibration of the chromatograph. In other embodiments, calibration of the chromatograph may occur less frequently than calibration of the mass analyzer, and a species known to be in a sample or spiked into a sample can be used.

At <NUM>, the mass related physical value is measured for the ions within the sample, including the species of known mass. At <NUM>, a calibration curve is calculated for the mass related physical value. The calibration uses the set of mass related physical values measured for the calibrant ion species and the mass related physical value measured for the species of known mass in the sample. In various embodiments, the calibration curve can be used to determine masses of unknown ions from the measured mass related physical value with an error of not greater than about <NUM> ppm, such as not greater than about <NUM> ppm, such as not greater than about <NUM> ppm, or even not greater than about <NUM> ppm.

At <NUM>, the operation of the mass analyzer is modified based on the calibration curve. In embodiments, when using a Fourier transform mass analyzer, a frequency range for an image current, a digitizing rate, a filter bandwidth for an image current, or any combination thereof can be modified based on the calibration curve. In other embodiments, when using a quadrupole mass analyzer or a quadrupole ion trap mass analyzer, an RF voltage, a DC voltage, or any combination thereof can be modified based on the calibration curve. In further embodiments, when using a time-of-flight mass analyzer, an acquisition time window, a flight tube clearing pulse time, or any combination thereof can be modified based on the calibration curve.

At <NUM>, ions can be generated from a second sample, and at <NUM>, the mass related physical value can be measured using the modified operation of the mass analyzer.

At <NUM>, the mass of the ions in the second sample can be determined based on the calibration curve. In various embodiments, a lock mass can be used to modify the calibration curve to further improve mass accuracy. The lock mass can be derived from a species of known mass within the second sample and can be used to shift or rotate the curve to account for run specific changes in the operation of the mass analyzer. Various factors can contribute to run specific alterations to the measured mass related physical value, such as a temperature of various components of the mass analyzer, a total number of ions within the mass analyzer, and other factors. The use of the lock mass can correct for these run specific affects without altering the operation of the mass analyzer, and can be applied while the data is collected for the sample and before saving the data, or can be applied after the data is collected for the sample.

<FIG> is a block diagram that illustrates a computer system <NUM>, upon which embodiments of the present teachings may be implemented which may incorporate or communicate with a system controller, for example controller <NUM> shown in <FIG>, such that the operation of components of the associated mass spectrometer may be adjusted in accordance with calculations or determinations made by computer system <NUM>. Computer system <NUM> can include a bus <NUM> or other communication mechanism for communicating information, and a processor <NUM> coupled with bus <NUM> for processing information. Computer system <NUM> can also include a memory <NUM>, which can be a random access memory (RAM) or other dynamic storage device, coupled to bus <NUM> for determining base calls, and instructions to be executed by processor <NUM>. Memory <NUM> also can be used for storing temporary variables or other intermediate information during execution of instructions to be executed by processor <NUM>. Computer system <NUM> can further include a read only memory (ROM) <NUM> or other static storage device coupled to bus <NUM> for storing static information and instructions for processor <NUM>. A storage device <NUM>, such as a magnetic disk or optical disk, can be provided and coupled to bus <NUM> for storing information and instructions.

Processor <NUM> can include a plurality of logic gates. The logic gates can include AND gates, OR gates, NOT gates, NAND gates, NOR gates, EXOR gates, EXNOR gates, or any combination thereof. An AND gate can produce a high output only if all the inputs are high. An OR gate can produce a high output if one or more of the inputs are high. A NOT gate can produce an inverted version of the input as an output, such as outputting a high value when the input is low. A NAND (NOT-AND) gate can produce an inverted AND output, such that the output will be high if any of the inputs are low. A NOR (NOT-OR) gate can produce an inverted OR output, such that the NOR gate output is low if any of the inputs are high. An EXOR (Exclusive-OR) gate can produce a high output if either, but not both, inputs are high. An EXNOR (Exclusive-NOR) gate can produce an inverted EXOR output, such that the output is low if either, but not both, inputs are high.

One of skill in the art would appreciate that the logic gates can be used in various combinations to perform comparisons, arithmetic operations, and the like. Further, one of skill in the art would appreciate how to sequence the use of various combinations of logic gates to perform complex processes, such as the processes described herein.

In an example, a <NUM>-bit binary comparison can be performed using a XNOR gate since the result is high only when the two inputs are the same. A comparison of two multi-bit values can be performed by using multiple XNOR gates to compare each pair of bits, and the combining the output of the XNOR gates using and AND gates, such that the result can be true only when each pair of bits have the same value. If any pair of bits does not have the same value, the result of the corresponding XNOR gate can be low, and the output of the AND gate receiving the low input can be low.

In another example, a <NUM>-bit adder can be implemented using a combination of AND gates and XOR gates. Specifically, the <NUM>-bit adder can receive three inputs, the two bits to be added (A and B) and a carry bit (Cin), and two outputs, the sum (S) and a carry out bit (Cout). The Cin bit can be set to <NUM> for addition of two one bit values, or can be used to couple multiple <NUM>-bit adders together to add two multi-bit values by receiving the Cout from a lower order adder. In an exemplary embodiment, S can be implemented by applying the A and B inputs to a XOR gate, and then applying the result and Cin to another XOR gate. Cout can be implemented by applying the A and B inputs to an AND gate, the result of the A-B XOR from the SUM and the Cin to another AND, and applying the input of the AND gates to a XOR gate.

Computer system <NUM> can be coupled via bus <NUM> to a display <NUM>, such as a cathode ray tube (CRT) or liquid crystal display (LCD), for displaying information to a computer user. An input device <NUM>, including alphanumeric and other keys, can be coupled to bus <NUM> for communicating information and command selections to processor <NUM>. Another type of user input device is a cursor control <NUM>, such as a mouse, a trackball or cursor direction keys for communicating direction information and command selections to processor <NUM> and for controlling cursor movement on display <NUM>. This input device typically has two degrees of freedom in two axes, a first axis (i.e., x) and a second axis (i.e., y), that allows the device to specify positions in a plane.

A computer system <NUM> can perform the present teachings. Consistent with certain implementations of the present teachings, results can be provided by computer system <NUM> in response to processor <NUM> executing one or more sequences of one or more instructions contained in memory <NUM>. Such instructions can be read into memory <NUM> from another computer-readable medium, such as storage device <NUM>. Execution of the sequences of instructions contained in memory <NUM> can cause processor <NUM> to perform the processes described herein. Instructions in the memory can sequence the use of various combinations of logic gates available within the processor to perform the processes describe herein. Alternatively hard-wired circuitry can be used in place of or in combination with software instructions to implement the present teachings. The hard-wired circuitry can include the necessary logic gates, operated in the necessary sequence to perform the processes described herein. Thus implementations of the present teachings are not limited to any specific combination of hardware circuitry and software.

The term "computer-readable medium" as used herein refers to any media that participates in providing instructions to processor <NUM> for execution. Such a medium can take many forms, including but not limited to, non-volatile media, volatile media, and transmission media. Examples of non-volatile media can include, but are not limited to, optical or magnetic disks, such as storage device <NUM>. Examples of volatile media can include, but are not limited to, dynamic memory, such as memory <NUM>. Examples of transmission media can include, but are not limited to, coaxial cables, copper wire, and fiber optics, including the wires that comprise bus <NUM>.

Common forms of non-transitory computer-readable media include, for example, a floppy disk, a flexible disk, hard disk, magnetic tape, or any other magnetic medium, a CD-ROM, any other optical medium, punch cards, paper tape, any other physical medium with patterns of holes, a RAM, PROM, and EPROM, a FLASH-EPROM, any other memory chip or cartridge, or any other tangible medium from which a computer can read.

Instructions configured to be executed by a processor to perform a method are stored on a computer-readable medium.

The methods of the present teachings may be implemented in a software program and applications written in conventional programming languages such as C, C++, C#, etc..

While the present teachings are described in conjunction with various embodiments, it is not intended that the present teachings be limited to such embodiments. On the contrary, the present teachings encompass various alternatives, modifications, and equivalents, as will be appreciated by those of skill in the art.

The embodiments described herein, can be practiced with other computer system configurations including hand-held devices, microprocessor systems, microprocessor-based or programmable consumer electronics, minicomputers, mainframe computers and the like. The embodiments can also be practiced in distributing computing environments where tasks are performed by remote processing devices that are linked through a network.

It should also be understood that the embodiments described herein can employ various computer-implemented operations involving data stored in computer systems. These operations are those requiring physical manipulation of physical quantities. Usually, though not necessarily, these quantities take the form of electrical or magnetic signals capable of being stored, transferred, combined, compared, and otherwise manipulated. Further, the manipulations performed are often referred to in terms, such as producing, identifying, determining, or comparing.

Any of the operations that form part of the embodiments described herein are useful machine operations. The embodiments, described herein, also relate to a device or an apparatus for performing these operations. The systems and methods described herein can be specially constructed for the required purposes or it may be a general purpose computer selectively activated or configured by a computer program stored in the computer. In particular, various general purpose machines may be used with computer programs written in accordance with the teachings herein, or it may be more convenient to construct a more specialized apparatus to perform the required operations.

The computer readable medium is any data storage device that can store data, which can thereafter be read by a computer system. Examples of the computer readable medium include hard drives, network attached storage (NAS), read-only memory, random-access memory, CD-ROMs, CD-Rs, CD-RWs, magnetic tapes, and other optical and non-optical data storage devices. The computer readable medium can also be distributed over a network coupled computer systems so that the computer readable code is stored and executed in a distributed fashion.

The mass analyzer is calibrated using MS Grade Perfluorotributylamine (PFTBA) from SynQuest Labs, Alachua, FL. The PFTBA calibration compound produces ions having theoretical masses of <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM>. A mixture of bromobiphenyl ethers from tri- to deca- (from Sigma-Aldrich, St Louis, MO) is analyzed.

<FIG> shows an experimental mass spectrum of the bromobiphenyl ethers using the calibration derived from the PFTBA calibration compound as compared to a theoretical mass spectrum of the bromobiphenyl ethers. The error in the experimentally determined mass at m/z <NUM> is <NUM> ppm. The data are tabulated in Table <NUM>. <FIG> is a graph of the mass errors as a function of m/z ratio. The mass errors are stable within the m/z range covered by the PFTBA calibration compound, but increase as the m/z ratio moves further from the calibration range.

<FIG> shows an experimental mass spectrum of the bromobiphenyl ethers using the calibration derived from the PFTBA calibration compound and decabromobiphenyl ether in the sample as compared to a theoretical mass spectrum of the bromobiphenyl ethers. The error in the experimentally determined mass at m/z <NUM> is <NUM> ppm. The data are tabulated in Table <NUM>. <FIG> is a graph of the mass errors as a function of m/z ratio. <FIG> compares the mass error curve from <FIG> (mass calibration with PFTBA only) to the mass error curve for multiple runs calibrated using the PFTBA calibration compound and the decabromobphenyl ether within the sample. The mass errors when using both PFTBA and decabromobphenyl ether are relatively consistent across the measured m/z range. Although, the errors tend to be negative and the masses tend to be systematically underestimated for these samples.

<FIG> shows an experimental mass spectrum of the bromobiphenyl ethers using the calibration derived from the PFTBA calibration compound and decabromobiphenyl ether and using decabromobiphenyl ether as a lock mass as compared to a theoretical mass spectrum of the bromobiphenyl ethers. The error in the experimentally determined mass at m/z <NUM> is <NUM> ppm. The data are tabulated in Table <NUM>. <FIG> is a graph of the mass errors as a function of m/z ratio. <FIG> compares the mass error curve from <FIG> (mass calibration with PFTBA only) to the mass error curve for multiple runs when the PFTBA calibration compound and the decabromobphenyl ether are used for the calibration curve and decabromobphenyl ether (alone or in combination with Heptabromobiphenyl ether) is used as a lock mass. The mass errors when using both PFTBA and decabromobphenyl ether along with a lock mass are relatively consistent across the measured m/z range.

Claim 1:
A method comprising:
producing ions from one or more calibrant species and delivering the ions to a mass analyzer (<NUM>);
measuring a first set of mass related physical values for the ions from the one or more calibrant species;
producing ions from a sample and delivering the ions to a mass analyzer (<NUM>), the sample including ion species of unknown mass and a first sample ion species of known mass, the first sample ion species having a mass-to-charge ratio outside of the range of the mass-to-charge ratios of the calibrant ion species;
measuring a second mass related physical value for the first sample ion species;
calculating a calibration curve based on the first set of mass related physical values for the plurality calibrant ion species and second mass related physical value for the first sample ion species; and
modifying at least one instrument parameter based on the calibration curve.