AUTOMATED SYSTEM FOR DELIVERING AT LEAST ONE SAMPLE FOR ELECTROSPRAY IONIZATION IN A MASS SPECTROMETER SYSTEM - Patent application

Abstract

An automation system (112) for providing at least one sample (136) for electrospray ionization in a mass spectrometer system (110) is disclosed. The automation system (112) includes at least one electrospray emitter (122) having at least one emitter end (120) with at least one emitter tip (124) and at least one fluid inlet end (126), at least one autosampler (132) having at least one autosampler outlet (134), the autosampler (132) configured to provide at least one sample (136) having at least one analyte, and at least one pipe (138) having at least one pipe fluid outlet end (142) and at least one pipe fluid inlet end (144), the pipe fluid outlet end (142) being configured to inject a sample into the at least one sample (136). The inlet end (144) comprises at least one pipe (138) fluidly connected to the autosampler outlet (134), and at least one liquid junction (146), the liquid junction (146) comprising at least one connecting element (148) formed of at least one electrically conductive material, the connecting element (148) receiving the fluid inlet end (126) of the electrospray emitter (122) and the pipe fluid outlet end (142) of the pipe (138) such that a fluid connection is established between the electrospray emitter (122) and the pipe (138), the connecting element (148) comprising at least one liquid junction (146) electrically connectable to at least one voltage source, and the emitter tip (124) comprises an opening having a diameter of 1 μm to 10 μm.

Description

[Technical field]

[0001] The present invention relates to an automated system for providing at least one sample for electrospray ionization in a mass spectrometer system, a mass spectrometer system, a method for providing at least one sample in a mass spectrometer system and ionizing by electrospray ionization, a method for optimizing at least one mass spectrometry (MS) parameter, a method for detecting at least one analyte in a sample by mass spectrometry (MS) and for use of the automated system and / or for use of the mass spectrometer system. The device and the method can have a wide range of applications in the research fields of proteomics, peptidomics, and metabolomics. Furthermore, the device and the method can be applied for the analysis of small molecules in metabolomics, drug discovery, and can also be useful for the analysis of therapeutic antibodies. However, other applications can also be implemented. [Background technology]

[0002] Direct detection of tumor-derived neoepitopes generally poses analytical challenges since neoepitopes may be of very low abundance in the context of personalized immunotherapy. Orbitrap instruments coupled to ion mobility-based interfaces such as nano-liquid chromatography (nanoLC-MS) and / or field asymmetric ion mobility spectrometry (FAIMS) could in principle increase the sensitivity required for detection of rare peptides. However, it requires fine-tuning of essential instrument parameters for each individual peptide.

[0003] In WO 2013 / 102670, an electrostatic spray ionization method is described. In the electrostatic spray ionization method for spraying a liquid layer from an insulating plate, the plate is placed between two electrodes. A constant high voltage power supply is provided, and an electric circuit is used to locally charge and discharge the surface of the liquid layer on the insulating plate by applying a power supply between the electrodes.

[0004] US Patent Application Publication No. 2019 / 0259597 describes a liquid junction for electrospray ionization in a mass spectrometer, the liquid junction comprising an electrospray emitter, a capillary conduit assembly for guiding a liquid to be sprayed with electrospray, and a union comprising a conductive material, the liquid junction being housed in a bottom sealed butt joint featuring a small dead volume while holding at least one of them pluggable and retractable in and out of the union comprising a conductive material, the liquid junction facilitating energizing the delivered liquid to a predetermined voltage level at the liquid junction upstream of the actual emitter tip where electrospray occurs, while holding at least one of the electrospray emitter and capillary conduit assembly pluggable in and retractable out of the union comprising a conductive material.

[0005] Giovanni D'Orazio et al., Journal of Chromatography A, 1317 (2013) 67-76, describe the study of a new nano-liquid junction interface for coupling both capillary electrochromatography (CEC) or nano-liquid chromatography (nano-LC) with mass spectrometry (MS). The interface is a small T-piece of polymeric material, in which the capillary column and the tip capillary are positioned at 180°, while the third outlet (at 90°) is occupied by a capillary conveying liquid-assisted spray ionization for CEC experiments, or by an electrode for high-voltage spray for nano-LC. Experiments were carried out to analyze a mixture of several organophosphorus drugs (OPPs) or a mixture of anti-inflammatory related acid drugs, with MS detection in positive or negative ion mode, respectively.

[0006] US Patent No. 10,983,098 describes a liquid chromatograph, which includes a column, a liquid delivery unit configured to deliver a mobile phase to the column at a pressure higher than atmospheric pressure, a first pipe having one end connected to an outlet of the column, and a second pipe having one end connected to an end face at the other end of the first pipe, the second pipe having the other end located in an ionization chamber having a pressure lower than or equal to atmospheric pressure, with a connection gap sandwiched between said one end and said end face of the second pipe.

[0007] WO 98 / 35226 describes the design of a simple, robust, sheathless interface for capillary electrophoresis / electrospray ionization-mass spectrometry (CE / ESI-MS) using common laboratory tools and chemicals. The interface uses a small platinum (Pb) wire that is inserted into the CE capillary through a small hole near the end. The position of the wire inside the CE capillary and in the buffer solution is similar to standard CE separation operation, where the end of the CE capillary is placed inside a buffer reservoir with a grounded platinum electrode. By combining the use of an in-capillary electrode interface with a sharpened fused silica tip at the outlet of the CE capillary, a stable electrospray current was maintained for extended periods of time.

[0008] US Patent No. 5,993,633 refers to an interface between an end of a capillary electrophoresis separation capillary and an end of an electrospray ionization mass spectrometry emitter capillary for transporting an analyte sample from the capillary electrophoresis separation capillary to an electrospray ionization mass spectrometry emitter capillary, the interface having (a) a charge transfer fitting surrounding both the end of the capillary electrophoresis capillary and the end of the electrospray ionization mass spectrometry emitter capillary, (b) a reservoir containing an electrolyte surrounding the charge transfer fitting, and (c) an electrode immersed in the electrolyte that closes the capillary electrophoresis circuit and provides charge transfer across the charge transfer fitting while avoiding substantial bulk liquid transport across the charge transfer fitting.

[0009] In Kegi Tang et al., J Am Soc Mass Spectrom 2004, 15, 1416-1423, experimental investigations and theoretical analysis of charge competition in electrospray ionization (ESI) and its effect on the linear dynamic range of ESI mass spectrometry (MS) measurements are reported. Experiments confirmed that MS sensitivity increases as expected with decreasing ESI flow rate. However, even at the same concentration and same electrospray operating conditions, different compounds show somewhat different mass spectral peak intensities even at the lowest flow rate. The MS response for each compound solution shows good linearity at low concentrations and levels off at higher concentrations, consistent with "saturation" of the analyte in the ESI process. The degree of charge competition leading to saturation in the ESI process is consistent with the relative magnitude of the excess charge in the electrospray compared to the total number of analyte molecules in solution.

[0010] WO 03 / 042684 describes an apparatus for connecting a capillary for capillary electrophoresis (CE) to an ionization source in an apparatus for mass spectrometry (MS), comprising a chamber for electrolyte with a first inlet for the capillary, a second inlet for electrolyte from a reservoir and an outlet for the capillary, a tubular electrode through which the capillary is passed is arranged connected to said outlet, electrolyte can flow around the capillary through said electrode, a flow chamber is arranged upstream with respect to said electrode and in the flow chamber a conductive surface is arranged which is electrically connected to said electrode.

[0011] Additionally, Advion offers an in-chip based electrospray ionization technology called TriVersa NanoMate LESA® (link: https: / / www.advion.com / products / triversa-nanomate / ). Additionally, emitters are described on the PepSep website (link: https: / / pepsep.com / products.html).

[0012] Despite the advantages achieved by the above mentioned devices and methods, some technical challenges remain. Most of the above mentioned devices are linked to chromatographic and electrophoretic separation techniques, which allow the complexity of the mixture of analytes to be reduced. However, such devices may be limited to a certain range of physicochemical properties of the analytes. A typical static spray workflow can allow the direct injection of microliter amounts of samples for their unbiased analysis, usually for a period of several hours, during which a large number of parameters can be changed and the corresponding data can be recorded. However, it generally suffers from poor reproducibility due to the manufacture of coated glass capillaries, which can have a metal coating, for example by sputter deposition of a thin metal layer. The metal coating can easily erode and result in unevenness at the tip of the capillary, which can lead to electrospray instability. As a result, the lack of reproducibility, as well as sample loss and the tediousness of manual handling, have led to the decline of the use of static spray in mass spectrometry-based proteomics and peptidomics workflows. Summary of the Invention [Problem to be solved by the invention]

[0013] It is therefore desirable to provide an automated system for providing at least one sample for electrospray ionization in a mass spectrometer system, a mass spectrometer system, a method for providing at least one sample in a mass spectrometer system and ionizing by electrospray ionization, a method for optimizing at least one mass spectrometry (MS) parameter, a method for detecting at least one analyte in a sample by mass spectrometry (MS), and the use of the automated system and / or the mass spectrometer system that at least partially addresses the above-mentioned technical challenges. In particular, measurement reproducibility and throughput will be increased. [Means for solving the problem]

[0014] This problem is addressed by an automation system for providing at least one sample for electrospray ionization in a mass spectrometer system, a mass spectrometer system, a method for providing at least one sample in a mass spectrometer system and ionizing by electrospray ionization, a method for optimizing at least one mass spectrometry (MS) parameter, a method for detecting at least one analyte in a sample by mass spectrometry (MS), and the use of the automation system and / or the mass spectrometer system with the features of the independent claims. Advantageous embodiments, which may be realized in isolation or in any combination, are listed in the dependent claims as well as in the entire specification.

[0015] As used below, the terms "have", "comprise", "include" or any grammatical variants thereof are used in a non-exclusive manner. Thus, these terms can refer both to the situation where there are no further features in the entity described in this context other than the features introduced by these terms, and to the situation where one or more further features are present. As an example, the expressions "A has B", "A comprises B" and "A includes B" can refer both to the situation where there are no other elements in A other than B (i.e., A consists solely and exclusively of B), and to the situation where there are one or more further elements in entity A other than B, such as element C, elements C and D, or even further elements.

[0016] Furthermore, it should be noted that the terms "at least one," "one or more," or similar expressions indicating that a feature or element may be present once or more than once are generally used only once when introducing each feature or element. In the following, in most cases, when referring to each feature or element, the expressions "at least one" or "one or more" will not be repeated, despite the fact that each feature or element may be present once or more than once.

[0017] Furthermore, as used below, the terms "preferably", "more preferably", "particularly", "more particularly", "particularly", "more particularly" or similar terms are used with optional features without limiting the alternative possibilities. Features introduced by these terms are therefore optional features and are not intended to limit the scope of the claims in any way. The invention can be practiced by using alternative features, as understood by those skilled in the art. Similarly, features introduced by the phrase "in an embodiment of the invention" or similar phrases are intended to be optional features, without any limitations on alternative embodiments of the invention, without any limitations on the scope of the invention, and without any limitations on the possibility of combining the feature so introduced with other optional or non-optional features of the invention.

[0018] In a first aspect of the present invention, an automated system for providing at least one sample for electrospray ionization in a mass spectrometer system, in particular for transporting said at least one sample, is disclosed. The automated system comprises at least one electrospray emitter comprising at least one emitter end with at least one emitter tip and at least one fluid inlet end. Furthermore, the automated system comprises at least one autosampler. The autosampler comprises at least one autosampler outlet. The autosampler is configured to provide at least one sample having at least one analyte. Furthermore, the automated system comprises at least one pipe, in particular at least one fused silica capillary, having at least one pipe fluid outlet end and at least one pipe fluid inlet end. The pipe fluid inlet end is fluidly connected to the autosampler outlet. Furthermore, the automated system comprises at least one liquid junction. The liquid junction comprises at least one connection element formed of at least one electrically conductive material. The connecting element receives the fluid inlet end of the electrospray emitter and the pipe fluid outlet end of the pipe such that a fluid connection is established between the electrospray emitter and the pipe. The connecting element is electrically connectable to at least one voltage source. The emitter tip includes an opening having a diameter of 1 μm to 10 μm.

[0019] The term "system" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to any particular or customized meaning. The term may specifically, but not be limited to, refer to a group of at least two elements that can interact with each other to perform at least one common function. The at least two components may be treated independently or may be linked, connectable, or integrated to form a common component.

[0020] The term "mass spectrometer system" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to any special or customized meaning. The term can specifically refer to, but is not limited to, any group of at least two elements, one of which is a mass spectrometry analyzer. The other of the at least two elements may specifically be a device configured to provide and / or prepare at least one sample for performing at least one mass spectrometry analysis. Further details are given in more detail below.

[0021] The term "mass spectrometry analyzer" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to any special or customized meaning. The term may specifically refer to, but is not limited to, any analytical technique configured to measure the mass-to-charge ratio of ions. The mass spectrometry analyzer may comprise at least one mass analyzer and at least one detector. The mass analyzer and detector may be disposed within an ionization chamber. The ionization chamber may also be referred to as an "ion source." The mass analyzer may be configured to classify and differentiate a plurality of ions according to their mass-to-charge ratio. The mass spectrometry analyzer may comprise several ion optics, such as skimmers, focusing lenses, electromagnetic elements such as multipoles. The ion optics may be configured to convey the ions to a region inside the ionization chamber where the mass analyzer is disposed. The mass spectrometry analyzer may be selected from the group consisting of a time-of-flight analyzer, a triple quadrupole analyzer, an ion trap, an ion cyclotron resonance cell, and an orbitrap. However, other embodiments may be implemented.

[0022] The mass spectrometer system may further comprise at least one ion mobility spectrometry device, preferably a high field asymmetric waveform ion mobility spectrometer. The ion mobility spectrometry device may be configured to separate a plurality of ions generated in the ion source. The term "ion mobility spectrometry device" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically refer to, but is not limited to, any analytical technique configured to separate and identify a plurality of ionized molecules in the gas phase based on their mobility in a carrier buffer gas. The ion mobility spectrometry device may be configured to actively correct the drift of some ions from the ion path towards the entrance of the mass spectrometer by a so-called compensation voltage (CV) so that these ions can enter the mass spectrometer. These ions may have some specificity, such as a predefined charge state or a predefined range of charge states. Thus, by applying the above-mentioned CV, a cloud of ions of a particular charge state may be separated from further ions.

[0023] The term "electrospray ionization (ESI)" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to any special or customized meaning. The term can specifically refer to, but is not limited to, any process in which ions are generated using electrospray, whereby a high voltage can be applied to a liquid. Mass spectrometry using ESI is commonly referred to as electrospray ionization mass spectrometry (ESI-MS) or, less commonly, electrospray mass spectrometry (ES-MS). In electrospray ionization, a sample containing an analyte solution can be passed through a capillary and a voltage can be applied to the sample.

[0024] The automated system may be specifically configured to provide at least one sample for static spray ionization. The term "static spray ionization" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically refer to, but is not limited to, any electrospray ionization process in which the electrospray does not change systematically over time. Thus, as an example, at the 1 minute time point, the same ions are expected to be detected as at the 10 minute time point. Specifically, static spray ionization refers to an offline analysis of a sample. The term "offline analysis" may refer to an analytical technique in which the injection of a sample may be performed without any specific separation being applied to the sample prior to its ionization. In contrast, online analysis may include the use of separation chromatography or the use of electrophoretic devices or elements.

[0025] The term "automation system" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term can specifically refer to, but is not limited to, any group of at least two elements that can interact with each other to perform at least one common function, where at least one of these two elements can be handled at least partially automatically, e.g., where at least one of these two elements is operable independently of continuous or constant human interference. In particular, an autosampler, more specifically one or more of the valves, fluid loading device, and pumps of the autosampler, and / or liquid junctions can be configured to be handled automatically, e.g., without user interaction. For this purpose, in particular, at least one processing device can be used. The term "processing device" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically, but not limited to, refer to any logic circuitry configured to perform basic operations of a computer or system, and / or may generally refer to a device configured to perform calculations or logical operations. A processing device may be configured to process basic instructions that run a computer or system. By way of example, a processing device may include at least one arithmetic logic unit (ALU), at least one floating point unit (FPU), such as a math coprocessor and a numeric coprocessor, a number of registers, specifically a number of registers configured to provide operands to the ALU and store results of operations, and memory, such as L1 and L2 cache memories. A processing device may be a multi-core processor. A processing device may be or include a central processing unit (CPU).Additionally or alternatively, the processing device may be or comprise a microprocessor, and thus, in particular, the elements of the processor may be included in a single integrated circuit (IC) chip. Additionally or alternatively, the processing device may be or comprise one or more application specific integrated circuits (ASICs), and / or the processing device may be or comprise one or more field programmable gate arrays (FPGAs), and / or the processing device may be or comprise one or more tensor processing units (TPUs), and / or the processing device may be or comprise one or more chips, such as a dedicated machine learning optimization chip. The processing device may be configured to perform one or more evaluation operations, such as by software programming. By way of example, the processing device may comprise software code stored thereon and comprising a number of computer instructions. A processing device may provide one or more hardware elements for performing one or more operations and / or may provide one or more processors having software operating thereon.

[0026] As outlined above, the automated system is configured to provide at least one sample for electrospray ionization in a mass spectrometer system. Thus, the automated system may also be referred to as a sample transport system. The term "sample transport system" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically refer to any system configured to transport a sample in order to use the sample for a special purpose, such as, but not limited to, performing analytical measurements. During the transport of the sample, at least one characteristic of the sample may change. Further details about the components of the automated system will be given in more detail below.

[0027] The term "sample" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term can specifically refer to any sample, such as, but not limited to, a biological sample or a synthetic sample. The sample can contain one or more analytes of interest. The sample can specifically be a liquid sample, in particular a liquid sample containing at least one biological material. The sample can be used directly as obtained from the respective source or can be subject to a pre-treatment and / or sample preparation workflow. Furthermore, the sample can be subjected to one or more processing steps. Thus, at least one characteristic of the sample can be changed.

[0028] The sample may in particular be a biological sample, such as a protein-containing sample, a chemical sample, a synthetic sample, and / or an environmental sample. Furthermore, the sample may be a sample of a subject, preferably a sample of a subject suffering from or suspected of suffering from a disease, preferably cancer. Furthermore, the sample may be a sample of a cell lysate, or a sample of a body fluid. Other embodiments may also be implemented.

[0029] As outlined above, a sample can contain at least one analyte. The term "analyte" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term can specifically refer to any chemical or biological substance or species, such as, but not limited to, a molecule or compound, to be detected and / or measured. In particular, the presence, absence, concentration, and / or amount of the analyte in the sample can be detected or measured.

[0030] The analyte may in particular be a biomolecule, preferably a protein. Furthermore, the biomolecule may be a polypeptide and / or a small molecule metabolite. In particular, the polypeptide may be a peptide presented by an antibody, a major histocompatibility antigen or a peptide in a body fluid, and / or a metabolite, and said small molecule metabolite may be a disease marker or an illegal drug and / or a metabolite thereof. Furthermore, the analyte may be a neo-epitope presented by a cancer cell. Other embodiments are also possible.

[0031] The terms "inlet" and "outlet" as used herein are broad terms and are to be given their ordinary and customary meaning to one of ordinary skill in the art and should not be limited to any special or customized meaning. The terms can specifically, but are not limited to, refer to the portions of an element through which a fluid medium enters or exits the element.

[0032] The term "fluidly connected" as used herein is a broad term and is to be given its ordinary and customary meaning to those of skill in the art and should not be limited to any special or customized meaning. The term can specifically, but is not limited to, refer to the property that two or more elements are connected in a manner that provides for the transfer of any fluid medium from one of the two elements to the other of the two elements and vice versa.

[0033] As outlined above, the automated system comprises an electrospray emitter. The term "electrospray emitter" is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term can specifically refer to, but is not limited to, any device configured to emit an electrospray. The electrospray can include charged droplets of a sample, specifically fine highly unipolar charged droplets of a sample with a narrow size distribution. The electrospray emitter can specifically comprise at least one fused silica capillary. The term "capillary" generally refers to any small, elongated void volume portion, such as a small tube. Generally, the capillary can have dimensions in the sub-millimeter range. Generally, the fluid medium can move through the capillary by capillary action, where the fluid medium can flow through the narrow space of the capillary without the aid of an external force such as gravity, due to intermolecular forces between the fluid medium and the surface of the capillary facing the fluid medium.

[0034] An electrospray emitter may comprise two opposite ends, specifically a fluid inlet end, which may also be referred to as an inlet, and an emitter end, which may also be referred to as an outlet. As outlined above, the emitter end comprises an emitter tip. The term "tip" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to any special or customized meaning. The term may specifically refer to, but is not limited to, the sharp end of any element, specifically the sharp end of an elongated element. Specifically, the emitter tip may be conically tapered at the emitter end. The emitter tip comprises an opening having a diameter of 1 μm to 10 μm. Specifically, the opening of the emitter tip has a diameter of 2 μm to 6 μm, preferably a diameter of 2.5 μm to 5.5 μm, and most preferably a diameter of 3 μm to 5 μm. The opening of the emitter tip may be referred to as an orifice.

[0035] As outlined above, the automated system comprises an autosampler. The term "autosampler" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term can specifically refer to any device that automatically delivers samples to laboratory equipment, such as, but not limited to, a mass spectrometry analyzer. The autosampler can specifically be configured to provide at least one sample. Thus, specifically, the autosampler can comprise at least one storage element, such as a flask or vessel, configured to provide a liquid and, specifically, to store a liquid. Specifically, the autosampler can comprise multiple storage elements. Further, additionally or alternatively, the autosampler can comprise at least one connection line, such as at least one tube. The connection line can be configured to transport the liquid to a desired location or to transport the liquid to another element. Thus, the autosampler can also be referred to as a liquid supply.

[0036] The autosampler may specifically include at least one sample tray configured to deliver at least one sample. The term "sample tray" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically refer to, but is not limited to, any device configured to receive and hold at least one sample, specifically at least one liquid sample. Specifically, the sample tray may be or include a holder for a plurality of flasks or vessels configured to deliver, specifically to store, samples. Furthermore, the sample tray may include a plurality of flasks or vessels. Furthermore, the autosampler may include at least one needle. The needle may be connected or connectable to a tube, which may be connected or connectable to a sample port of the first valve, which will be further described in more detail below. The needle may be configured to be selectively received in one of the flasks or vessels of the sample tray and at least one needle receiving seat of the autosampler.

[0037] The needle seat may be configured to receive the needle. In particular, the autosampler may be configured to wash the needle when the needle is received in the needle seat, as described in more detail further below.

[0038] Moreover, the autosampler may specifically comprise at least one first valve having at least one sample loop configured to receive a sample. The terms "first valve", "second valve", and "third valve" may be considered merely as names, without numbering or ranking the named elements, without specifying an order, and without excluding the possibility that there may be more than one type of first valve, second valve, or third valve. Furthermore, there may be additional blocks, ports, or positions, such as one or more fourth valves.

[0039] The term "valve" as used herein is a broad term and is to be given its ordinary and customary meaning to one of ordinary skill in the art and should not be limited to a special or customized meaning. The term can specifically refer to, but is not limited to, any device configured to block or control the flow of a fluid. Specifically, the valve can include at least one rotatable element, such as a disk with one or more channels configured to receive a fluid. Additionally, the valve can include at least one stationary element having one or more ports and can optionally include additional channels configured to flow and / or direct the fluid. The channels of the rotatable element can be selectively connectable to the ports and optionally connectable to additional channels of the stationary element by rotation of the rotatable element.

[0040] The term "sample loop" as used herein is a broad term and is to be given its ordinary and customary meaning to one of skill in the art and should not be limited to any special or customized meaning. The term can specifically refer to, but is not limited to, any element that can have an elongated shape, can provide a free volume or lumen, and allows for the flow of a sample therethrough. Thus, a sample loop can be configured to receive a sample and / or provide for the movement of the sample from one end of the sample loop to the other end of the sample loop.

[0041] Further, the autosampler may specifically comprise at least one fluid loading device, specifically at least one syringe, configured to load samples provided by a sample tray into the sample loop of the first valve. The term "fluid loading device" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically refer to, but is not limited to, any device configured to apply a fluid, specifically a defined or desired amount of fluid, to another object. The fluid loading device may specifically be a syringe. The term "syringe" may generally refer to a reciprocating pump that comprises a plunger or piston that fits tightly inside a cylindrical tube or barrel. The plunger or piston can be pulled or pushed linearly along the inside of said tube or barrel, allowing the syringe to take in and expel fluid through a discharge orifice at the open end of said tube. The open end of the syringe can be attached to tubing to direct flow into and out of the barrel.

[0042] Furthermore, the autosampler can include at least one third valve. Furthermore, the autosampler can include at least one washing liquid container configured to supply at least one washing liquid. The third valve can be configured to selectively connect the fluid loading device to the washing liquid container or to the fluid loading device port of the first valve. The third valve can also be called a fluid loading device valve or a syringe valve. For the design and structure of the third valve, reference can be made to the description of the first valve and the second valve. Specifically, the third valve can include four ports. However, other embodiments of the third valve can also be implemented.

[0043] Furthermore, the autosampler may specifically comprise at least one pump. The pump may be configured to transport the sample from the sample loop of the first valve to the autosampler outlet and to the electrospray emitter. The pump may specifically be a nanopump. The term "moving" or "transporting" may generally refer to the active transport of any material from one location to another or vice versa. In that regard, the term "active transport" generally means that the transport is assisted by an external force and / or an actuation means such as a pump or a valve used for the directed transport of the material. Thus, the term "active transport" may also refer to a defined manipulation of the material.

[0044] The first valve may further comprise at least one sample port, at least one fluid loading device port, at least one pump port, and at least one outlet port. The sample tray may be fluidly connectable to the sample loop of the first valve through the sample port. The fluid loading device may be fluidly connectable to the sample loop of the first valve through the fluid loading device port. The pump may be fluidly connectable to the sample loop of the first valve through the pump port. The autosampler outlet may be fluidly connectable to the sample loop of the first valve through the outlet port and the inlet port of the second valve. The automation system may be configured to load the sample into the sample loop of the first valve by establishing a fluid connection between the fluid loading device port and the sample port through the sample loop. Furthermore, the automation system may be configured to inject the sample into the pipe by establishing a fluid connection between the pump port and the outlet port through the sample loop.

[0045] Furthermore, the autosampler may comprise at least one second valve. The second valve may be configured to selectively fluidly connect the outlet port of the first valve to the autosampler outlet or to at least one waste container. The waste container may be any container configured to receive and / or store fluids that are no longer required for sample preparation and / or measurement purposes. The second valve may be configured to return the pump outlet to the waste container as soon as the sample is transferred from the sample loop of the first valve to the pipe fluid inlet end of the pipe. With regard to the design of the second valve, reference may be made to the above description of the first valve.

[0046] As outlined above, the automated system comprises a pipe. The term "pipe" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically, but not limited to, refer to any connection element that may be configured to supply a sample from an autosampler to an electrospray emitter via a liquid junction. Thus, the pipe may also be referred to as a supply line. The pipe may specifically be or comprise a fused silica capillary. However, other embodiments may also be implemented. Specifically, the pipe fluid inlet end of the pipe may be directly fluidly connected to the autosampler outlet. Thus, a separation element may not be disposed between the liquid junction and the autosampler. Specifically, a liquid column may not be disposed between the liquid junction and the autosampler.

[0047] As outlined above, the automated system comprises a liquid junction. The term "liquid junction" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term can specifically refer to, but is not limited to, any device configured to couple at least two elements for the purpose of conveying a liquid from one of the elements to another of the elements. Thus, specifically, a fluid outlet of one of the elements may be fluidly connected or fluidly connectable to a fluid inlet of another of the elements. Furthermore, the liquid junction may be configured to charge at least one component of the sample by applying a voltage. Specifically, the liquid junction may be configured to achieve stable and robust electrospray ionization.

[0048] As outlined above, the liquid junction comprises at least one connection element made of at least one electrically conductive material. The electrically conductive material may specifically be steel, such as stainless steel. However, other materials are also feasible. The term "connection element" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically refer to, but is not limited to, any device configured to establish a fluid connection between at least two elements. The fluid connection may specifically be established between a fluid outlet of one of the elements and a fluid inlet of another of the elements. As outlined above, the connection element receives a fluid inlet end of the electrospray emitter and a pipe fluid outlet end of the pipe such that a fluid connection between the electrospray emitter and the pipe is established. The connection element may specifically comprise at least one hollow cylinder having at least one first end and at least one opposite second end. The fluid inlet end of the electrospray emitter may be received at the first end and the pipe fluid outlet end may be received at the second end. The fluid inlet end of the electrospray emitter and the pipe fluid outlet end of the pipe may be received at the connecting element, in particular such that a nanogap is formed between the fluid inlet end and the pipe fluid outlet end. Thus, the sample may be charged at the nanogap by applying a voltage to the connecting element. The voltage may be in particular in the range of 800V to 1300V, preferably in the range of 1000V to 1200V. In particular, the voltage may be 1100V. The nanogap may have a gap of such a size that disturbance of the incoming flow of the sample, in particular of the incoming nanoflow of the sample, is at least largely prevented or reduced, in particular by, for example, inducing vortices to mix the sample with the adjacent transport liquid. In particular, the spacing between the fluid inlet end of the electrospray emitter and the fluid outlet end of the pipe and / or the diameter of the gap may have dimensions in the nanometer range."Adjacent transport liquid" refers to the liquid that sandwiches the sample on either side.

[0049] In a further aspect of the present invention, a mass spectrometer system is disclosed. The mass spectrometer system comprises at least one automation system as described above or as further described in more detail below. Furthermore, the mass spectrometer system comprises at least one mass spectrometry analyzer having at least one ion source. The emitter end of the electrospray emitter is disposed within the ion source.

[0050] In a further aspect of the present invention, a method for providing and ionizing at least one sample in a mass spectrometer system by electrospray ionization is disclosed.

[0051] The method includes using an automated system as described above or as further described in more detail below.

[0052] The method includes the method steps as given in the independent claims and as listed below. The method steps may be performed in a given order. However, the method steps can be performed in other orders. Furthermore, one or more of the method steps may be performed in parallel and / or overlapping in time. Furthermore, one or more of the method steps may be performed repeatedly. Furthermore, there may be additional method steps not listed.

[0053] The method includes the following steps: a) providing at least one sample to at least one sample tray of an autosampler; b) transferring the sample from the sample tray to the autosampler outlet, whereby the sample is further transferred to the electrospray emitter via a pipe and a liquid junction; and c) charging at least one component of the sample by applying a voltage to a connecting element of the liquid junction via a voltage source.

[0054] The term "provide" as used herein is a broad term and is to be given its ordinary and accustomed meaning to those of skill in the art and should not be limited to a specific or customized meaning. The term can specifically, but is not limited to, refer to the process of making available one or more needed objects.

[0055] The term "movement" or "transport" may generally refer to the active transport of any material from one location to another or vice versa. In that regard, the term "active transport" generally means that the transport is assisted by external forces and / or actuation means such as pumps or valves that are used for the directed transport of the material. Thus, the term "active transport" may also refer to the defined manipulation of the material.

[0056] The autosampler may specifically comprise at least one first valve having at least one sample loop, at least one fluid loading device, and at least one pump. For further details regarding the first valve, the sample loop, the fluid loading device, and the pump, reference is made to the above description.

[0057] Step b) may further comprise the steps of: b1) loading a sample into a sample loop of a first valve via a fluid loading device; and b2) Transferring the sample from the sample loop of the first valve to the autosampler outlet via a pump, whereby the sample is further transferred to the electrospray emitter via a pipe and a liquid junction. Transferring the sample from the sample tray to the autosampler outlet.

[0058] Specifically, the first valve may further comprise at least one sample port, at least one fluid loading device port, at least one pump port, and at least one outlet port. In step b1), a fluid connection between the fluid loading device port and the sample port via the sample loop may be established. In step b2), a fluid connection between the pump port and the outlet port via the sample loop may be established. In step b2), the pump may be configured to push the sample from the sample loop of the first valve. The pump may be connected to a solvent bottle. The pump may be configured to aspirate the sample and push the sample through the first valve.

[0059] Furthermore, the autospray can include at least one second valve, specifically, before performing step b), a first washing step can be performed, during which the second valve can fluidly connect the outlet port of the first valve to a waste container, and the fluidic connection can be formed between the pump port and at least one waste container via the sample loop and the outlet port.For further details regarding the second valve, reference is made to the above description.The first washing step can also be called sample loop washing step.

[0060] The autosampler may further comprise at least one third valve and at least one washing liquid container that may be configured to supply at least one washing liquid. As outlined above, the third valve may also be referred to as a fluid loading device valve or a syringe valve. For further details regarding the third valve and the washing liquid container, reference is made to the above description. At least one second washing step may be performed, in particular before performing step b). The second washing step may also be referred to as a fluid loading device priming step or a syringe priming step. The second washing step may include the following steps: i. fluidly connecting the fluid loading device to a washing fluid container via a third valve, whereby washing fluid is transferred, particularly aspirated, into an interior volume of the fluid loading device, particularly into a syringe; and ii. fluidly connecting the fluid loading device to a waste container via a third valve, whereby washing fluid is transferred from an internal volume of the fluid loading device to the waste container.

[0061] Furthermore, at least one third washing step may be performed, in particular before performing step b). The third washing step may also be called a sample needle washing step. The third washing step may include the following steps: i. fluidly connecting the fluid loading device to a washing fluid container via a third valve, whereby washing fluid is transferred, particularly aspirated, into an interior volume of the fluid loading device, particularly into a syringe; ii. placing the autosampler needle in the needle seat; and iii. fluidly connecting a fluid loading device to the needle via a third valve, whereby a wash fluid is transported from an internal volume of the fluid loading device to the needle receiving seat via the fluid loading device port of the first valve and the sample port of the first valve.

[0062] The terms "first wash step," "second wash step," and "third wash step" can be considered as mere names, without numbering or ranking the named steps, without specifying an order, and without excluding the possibility that more than one first wash step, second wash step, or third wash step may be performed. Additionally, additional wash steps, such as one or more fourth wash steps, may be performed.

[0063] The pressure inside the electrospray emitter during electrospray ionization may be atmospheric pressure. The term "atmospheric pressure" as used herein is a broad term and is to be given its ordinary and customary meaning to those skilled in the art and should not be limited to a special or customized meaning. The term may specifically refer to, but is not limited to, the pressure of a surrounding medium, such as a gas or liquid, in contact with the subject. Specifically, the pump may return to the waste container as soon as the sample is transferred from the sample tray to the pipe fluid inlet end of the pipe. After the transfer of the sample, the pump may be used to wash the sample loop. Specifically, the pump may be configured to idle as soon as the sample is transferred from the sample tray to the pipe fluid inlet end of the pipe and, specifically, finally to the electrospray emitter.

[0064] Furthermore, the pressure inside the electrospray emitter during electrospray ionization may be 1 bar to 50 bar, preferably 2 bar to 20 bar, and most preferably 5 bar to 15 bar. Conversely, in liquid chromatography, pressures of 300 bar to 400 bar may be applied depending on the column dimensions and the mode of chromatography.

[0065] The method may specifically be a method for providing at least two samples in a mass spectrometer system and ionizing by electrospray ionization, the method may include the steps of: (I) performing steps (a)-(c) on a first sample; (II) removing the first sample and cleaning at least the electrospray emitter, the pipe, and the liquid junction; (III) performing steps (a)-(c) on a second sample; and Optionally, repeating steps (II) and (III) for any additional samples.

[0066] After step (II) has been performed, one or more, or all of a first washing step, a second washing step, and a third washing step may also be performed.

[0067] The terms "first sample" and "second sample" may be considered as mere names, without numbering or ranking the named elements, without specifying an order, and without excluding the possibility that there may be more than one type of first sample or second sample. Furthermore, there may be additional samples, such as one or more third samples.

[0068] In a further aspect of the invention, a method for optimizing at least one mass spectrometry (MS) parameter is disclosed.

[0069] The term "MS parameters" is understood by those skilled in the art to relate to any MS parameters known or expected to affect the results of analytical MS performed on a sample. In one embodiment, said MS parameters may be selected from ionization method, type of detector, number of fragmentations, etc.

[0070] The term "optimize at least one MS parameter" is understood in principle by those skilled in the art. In one embodiment, the term relates to analyzing the influence of MS parameters. Furthermore, in one embodiment, the term relates to a procedure that includes modifying the parameters of an analytical MS method in such a way that the analyte of interest can be measured essentially without interference. Furthermore, in one embodiment, the term relates to investigating different characteristics of the analyte. In particular, data can be collected that can suggest different optimal conditions depending on the pursued goal of the assay to be optimized, especially in the case of determining the collision energy.

[0071] The method for optimizing at least one mass spectrometry (MS) parameter comprises the method steps as given in the independent claims and as listed below. The method steps may be performed in a given order. However, the method steps can be performed in other orders. Furthermore, one or more of the method steps may be performed in parallel and / or overlapping in time. Furthermore, one or more of the method steps may be performed repeatedly. Furthermore, there may be additional method steps not listed.

[0072] The method includes the following steps: (A) providing at least one sample and ionizing it by electrospray ionization, particularly by static electrospray ionization; (B) recording at least two MS spectra under conditions including at least two different values ​​for at least one MS parameter described above; (C) comparing the MS spectra recorded in step (B); and (D) thereby optimizing at least one MS parameter above based on the comparison in step (C).

[0073] Before step (B) is performed, the ions may optionally be separated in the gas phase by field asymmetric ion mobility spectrometry (FAIMS). In particular, a FAIMS device may be installed between the ion source and the inlet of the mass spectrometry analyzer. In this case, the optimization of at least one MS parameter may include, for example, optimizing the parameters for a given target peptide / molecule that should be optimally transferred to the mass spectrometry analyzer.

[0074] In a further aspect of the present invention, a method for detecting at least one analyte in a sample by mass spectrometry (MS) is disclosed.

[0075] A method for detecting at least one analyte in a sample by mass spectrometry (MS) comprises the method steps as given in the independent claims and as listed below. The method steps may be performed in a given order. However, the method steps can be performed in other orders. Furthermore, one or more of the method steps may be performed in parallel and / or overlapping in time. Furthermore, one or more of the method steps may be performed repeatedly. Furthermore, there may be additional method steps not listed.

[0076] The method includes the following steps: (A) providing and ionizing at least one sample using an automated system as described above or as further described in more detail below, preferably by a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system as described above or as further described in more detail below; and (B) Recording at least one MS spectrum of the ionized sample of step (A).

[0077] Furthermore, the method may include the steps of: (C) comparing at least one MS spectrum of step (B) with a reference, thereby detecting at least one analyte.

[0078] The term "reference" is understood in principle by those skilled in the art. In one embodiment, the term relates to a reference MS spectrum. Moreover, in one embodiment, the term relates to an expected value that can be calculated or experimentally determined. Other embodiments can also be implemented. In particular, the at least one MS spectrum of step (B) can be compared with an extracted spectral signal that is characteristic of the analyte. The spectral signal can be computationally determined. Thus, the detection of the analyte can be confirmed.

[0079] In a further aspect of the present invention, the use of an automated system as described above or as further described in more detail below and / or the use of a mass spectrometer system as described above or as further described in more detail below, for optimizing at least one MS parameter, preferably with a method for optimizing at least one mass spectrometry (MS) parameter as described above or as further described in more detail below, is disclosed.

[0080] In a further aspect of the present invention, the use of an automated system as described above or as further described in more detail below and / or the use of a mass spectrometer system as described above or as further described in more detail below is disclosed by a method for detecting at least one analyte in a sample, preferably by mass spectrometry (MS) parameters as described above or as further described in more detail below.

[0081] The present invention further discloses and proposes a computer program comprising computer executable instructions for carrying out the method according to the present invention in one or more of the embodiments enclosed herein when said program is executed on a computer or a computer network. In particular, the computer program may be stored on a computer readable data carrier. In particular, one or more or even all of the method steps a), b) and c) of the method for providing and ionizing at least one sample, the method steps (A), (B), (C) and (D) of the method for optimizing at least one mass spectrometry (MS) parameter, and / or one or even all of the method steps (A), (B) and (C) of the method for detecting at least one analyte in a sample by mass spectrometry (MS) as shown above, may be carried out by using a computer or a computer network, preferably by using a computer program. In particular, a table of scan parameters may be provided in the software of the instrument.

[0082] The present invention further discloses and proposes a computer program product having program code means for performing, when the program is executed on a computer or a computer network, several or all of the method steps a), b) and c) of the method for providing and ionizing at least one sample, the method steps (A), (B), (C) and (D) of the method for optimizing at least one mass spectrometry (MS) parameter, and / or one or all of the method steps (A), (B) and (C) of the method for detecting at least one analyte in a sample by mass spectrometry (MS) as indicated above, according to one or more of the embodiments of the present invention described herein. In particular, the program code means may be stored on a computer readable data carrier.

[0083] Furthermore, the present invention discloses and proposes a data carrier having a data structure stored thereon, which is capable, after being loaded into a computer or a computer network, such as into a working or main memory of a computer or a computer network, of performing, according to one or more of the embodiments disclosed herein, several or all of the method steps a), b) and c) of a method for providing and ionizing at least one sample, the method steps (A), (B), (C) and (D) of a method for optimizing at least one mass spectrometry (MS) parameter, and / or of performing one or all of the method steps (A) and (B) of a method for detecting at least one analyte in a sample by mass spectrometry (MS) as indicated above.

[0084] The present invention further discloses and proposes a computer program product having program code means stored on a machine-readable carrier for performing, when the program is executed on a computer or a computer network, several or all of the method steps a), b) and c) of the method for providing and ionizing at least one sample, the method steps (A), (B), (C) and (D) of the method for optimizing at least one mass spectrometry (MS) parameter, and / or one or all of the method steps (A) and (B) of the method for detecting at least one analyte in a sample by mass spectrometry (MS) as indicated above, according to one or more of the embodiments disclosed herein. As used herein, a computer program product refers to a program as a tradeable product. The product can generally be present in any format, such as a paper format or on a computer-readable data carrier. In particular, the computer program product may be distributed through a data network.

[0085] Finally, the present invention proposes and discloses a modulated data signal comprising instructions readable by a computer system or a computer network for performing several or all of method steps a), b) and c) of a method for providing and ionizing at least one sample, method steps (A), (B), (C) and (D) of a method for optimizing at least one Mass Spectrometry (MS) parameter, and / or for performing one or all of method steps (A) and (B) of a method for detecting at least one analyte in a sample by Mass Spectrometry (MS) as set out above, according to one or more of the embodiments disclosed herein.

[0086] Preferably, referring to the computer-implemented aspects of the present invention, according to one or more of the embodiments disclosed herein, multiple or all of the method steps a), b) and c) of the method for providing and ionizing at least one sample, the method steps (A), (B), (C) and (D) of the method for optimizing at least one mass spectrometry (MS) parameter, and / or one or all of the method steps (A) and (B) of the method for detecting at least one analyte in a sample by mass spectrometry (MS) as shown above, can be performed by using a computer or a computer network. Thus, in general, any of the method steps including providing and / or manipulating data can be performed by using a computer or a computer network. In general, these method steps can include any of the method steps that generally exclude method steps that require manual operations, such as certain aspects of providing the sample and / or performing the actual measurement.

[0087] Specifically, the present invention further comprises: a computer or computer network comprising at least one processor configured to perform multiple or all of method steps a), b) and c) of a method for providing and ionizing at least one sample, method steps (A), (B), (C) and (D) of a method for optimizing at least one mass spectrometry (MS) parameter, and / or to perform one or all of method steps (A) and (B) of a method for detecting at least one analyte in a sample by mass spectrometry (MS) according to one of the embodiments described herein; a computer loadable data structure configured, when executed on a computer, to perform multiple or all of method steps a), b) and c) of a method for providing and ionizing at least one sample, method steps (A), (B), (C) and (D) of a method for optimizing at least one mass spectrometry (MS) parameter, and / or to perform one or all of method steps (A) and (B) of a method for detecting at least one analyte in a sample by mass spectrometry (MS) according to one of the embodiments described herein; A computer program configured to perform, when the program is run on a computer, several or all of the method steps a), b) and c) of a method for providing and ionizing at least one sample, the method steps (A), (B), (C) and (D) of a method for optimizing at least one mass spectrometry (MS) parameter, and / or one or all of the method steps (A) and (B) of a method for detecting at least one analyte in a sample by mass spectrometry (MS) according to one of the embodiments described herein, A computer program comprising program means for performing, while said computer program is run on a computer or on a computer network, several or all of the method steps a), b) and c) of a method for providing and ionizing at least one sample, the method steps (A), (B), (C) and (D) of a method for optimizing at least one mass spectrometry (MS) parameter, and / or one or all of the method steps (A) and (B) of a method for detecting at least one analyte in a sample by mass spectrometry (MS) according to one of the embodiments described herein, A computer program comprising program means according to the above embodiment, the program means being stored in a computer-readable recording medium; A recording medium, on which a data structure is stored, the data structure being configured, after being loaded into a main and / or working storage of a computer or a computer network, to perform several or all of the method steps a), b) and c) of a method for providing and ionizing at least one sample, the method steps (A), (B), (C) and (D) of a method for optimizing at least one mass spectrometry (MS) parameter, and / or to perform one or all of the method steps (A) and (B) of a method for detecting at least one analyte in a sample by mass spectrometry (MS), according to one of the embodiments described herein; and a computer program product having program code means storable or stored on a storage medium for performing, when said program code means execute on a computer or on a computer network, more than one or all of method steps a), b) and c) of a method for providing and ionizing at least one sample, method steps (A), (B), (C) and (D) of a method for optimizing at least one mass spectrometry (MS) parameter, and / or one or all of method steps (A) and (B) of a method for detecting at least one analyte in a sample by mass spectrometry (MS) according to one of the embodiments described herein; Disclose.

[0088] The terms "second automation system," "second mass spectrometer system," "third mass spectrometer system," "second method for detecting at least one analyte in a sample by mass spectrometry (MS)," and "third method for detecting at least one analyte in a sample by mass spectrometry (MS)" may be considered as mere names, without numbering or ranking the named elements, without specifying an order, and without excluding the possibility that there may be multiple first / second / third automation systems, multiple first / second / third mass spectrometer systems, and multiple first / second / third methods for detecting at least one analyte in a sample by mass spectrometry (MS).

[0089] In a further aspect, a second automated system for providing at least one sample for electrospray ionization in a mass spectrometer system is described. The second automated system comprises at least one electrospray emitter comprising at least one emitter end having at least one emitter tip and at least one fluid inlet end. Furthermore, the second automated system comprises at least one autosampler. The autosampler comprises at least one autosampler outlet. The autosampler is configured to provide at least one sample having at least one analyte. Furthermore, the second automated system comprises at least one pipe, specifically at least one fused silica capillary, having at least one pipe fluid outlet end and at least one pipe fluid inlet end. The pipe fluid inlet end is fluidly connected to the autosampler outlet. The pipe fluid outlet end is fluidly connected to the fluid inlet end of the electrospray emitter. Furthermore, the second automated system comprises at least one device for applying a voltage to the sample. The emitter tip comprises an opening having a diameter of 1 μm to 10 μm.

[0090] Regarding the electrospray emitter, the pipe and the autosampler, reference is made to the above descriptions.

[0091] The device for applying a voltage to the sample may be a metal coated emitter, although other embodiments are also possible.

[0092] In a further embodiment, a second mass spectrometer system is described. The second mass spectrometer system comprises at least one second automated system as described above or as described in more detail below. Furthermore, the second mass spectrometer system comprises at least one mass spectrometry analyzer having at least one ion source. The emitter end of the electrospray emitter is disposed inside the ion source.

[0093] In a further embodiment, a second method is described for providing at least one sample in a mass spectrometer system and ionizing it by electrospray ionization.The method includes using a second automated system as described above or as further described in more detail below.The method includes the following steps: a) providing at least one sample into at least one sample tray of an autosampler; b) transferring the sample from the sample tray to an autosampler outlet, whereby the sample is further transferred to the electrospray emitter via a pipe and a liquid junction; and c) charging at least one component of the sample by applying a voltage to the sample.

[0094] In a further embodiment, a second method for detecting at least one analyte in a sample by mass spectrometry (MS) is described. The method comprises the steps of: (A) providing and ionizing a sample using a second automated system as described above or as further described in more detail below, preferably by the method for optimizing at least one mass spectrometry (MS) parameter as described above or as further described in more detail below; and (B) Recording at least one MS spectrum of the ionized sample of step (A).

[0095] In a further aspect, the use of a second automated system as described above or as further described in more detail below and / or the use of a second mass spectrometer system as described above or as further described in more detail below is disclosed for optimizing at least one MS parameter, preferably by a method for optimizing at least one mass spectrometry (MS) parameter as described above or as further described in more detail below.

[0096] In a further embodiment, the use of a second automated system as described above or as further described in more detail below, and / or the use of a second mass spectrometer system as described above or as further described in more detail below, with a second method for detecting at least one analyte in a sample, preferably by mass spectrometry (MS) parameters as described above or as further described in more detail below, is disclosed.

[0097] In a further aspect, a system for providing, and in particular for transporting, at least one sample for electrospray ionization in a mass spectrometer system is disclosed. The system comprises at least one electrospray emitter comprising at least one emitter end having at least one emitter tip and at least one fluid inlet end. Furthermore, the system comprises at least one autosampler. The autosampler comprises at least one autosampler outlet. The autosampler is configured to provide at least one sample having at least one analyte. Furthermore, the system comprises at least one pipe, in particular at least one fused silica capillary, having at least one pipe fluid outlet end and at least one pipe fluid inlet end. The pipe fluid inlet end is fluidly connected to the autosampler outlet. Furthermore, the system comprises at least one liquid junction. The liquid junction comprises at least one connection element made of at least one conductive material. The connecting element receives a fluid inlet end of the electrospray emitter and a pipe fluid outlet end of the pipe such that a fluid connection is established between the electrospray emitter and the pipe. The connecting element is electrically connectable to at least one voltage source. The emitter tip includes an opening having a diameter of 1 μm to 10 μm.

[0098] With regard to the electrospray emitter, the pipe and the autosampler, reference is made to the above descriptions, in particular to the above descriptions regarding the automated system according to the first aspect of the invention.

[0099] In a further embodiment, a third mass spectrometer system is described. The third mass spectrometer system comprises at least one system as described above or as further described in more detail below. Furthermore, the mass spectrometer system comprises at least one mass spectrometry analyzer having at least one ion source. The emitter end of the electrospray emitter is disposed inside the ion source.

[0100] In a further aspect, a third method is described for providing at least one sample in a third mass spectrometer system and ionizing by electrospray ionization. The method includes using a system as described above or as further described in more detail below. The method includes the following steps: a) providing at least one sample into at least one sample tray of an autosampler; b) transferring the sample from the sample tray to an autosampler outlet, whereby the sample is further transferred to the electrospray emitter via a pipe and a liquid junction; and c) charging at least one component of the sample by applying a voltage to the sample.

[0101] In a further embodiment, a third method for detecting at least one analyte in a sample by mass spectrometry (MS) is described. The method comprises the steps of: (A) providing and ionizing a sample using a system as described above or as further described in more detail below, preferably by a third method for optimizing at least one mass spectrometry (MS) parameter as described above or as further described in more detail below; and (B) Recording at least one MS spectrum of the ionized sample of step (A).

[0102] In a further aspect, the use of a system as described above or as further described in more detail below, and / or a third mass spectrometer system as described above or as further described in more detail below, for optimizing at least one MS parameter, preferably by a method for optimizing at least one mass spectrometry (MS) parameter as described above or as further described in more detail below, is disclosed.

[0103] In a further aspect, the use of a system as described above or as further described in more detail below, and / or the use of a third mass spectrometer system as described above or as further described in more detail below, by a third method for detecting at least one analyte in a sample, preferably by mass spectrometry (MS) parameters as described above or as further described in more detail below, is disclosed.

[0104] Methods and apparatus in accordance with the present invention offer numerous advantages over known methods and apparatus.

[0105] The automated system can be easily plugged and unplugged from the ion source of the mass spectrometry analyzer. The automated system can be configured to transfer at least one sample, particularly at low nanoliter per minute flow rates. The automated system allows for stable ionization when performing a method for providing and ionizing at least one sample by electrospray ionization in a mass spectrometer system.

[0106] Using static spray instead of the common LC-MS method to achieve ultra-high sensitivity, efficient procedures for optimizing various detection parameters may be implemented, which may in particular be automated. Multiple LC-MS runs, including multiple back and forth steps through multiple software platforms, followed by cleaning, may be avoided.

[0107] An automated system according to the invention may be based on a liquid junction between a tube, in particular a fused silica capillary, and an electrospray emitter, with a controlled size of the tip opening, which may also be called an orifice. This configuration makes it possible (i) to achieve a much more stable electrospray, and (ii) to open up the perspective of automation for transferring the sample to the electrospray emitter. Static spray mass spectrometry based on a liquid junction has never been combined with an autosampler before.

[0108] A clear advantage of the automated system according to the invention is that it can achieve very low flow rates. While the TriVersa Nano Mate® can only achieve 200 nl / min (comparable to nano-LC, but without the obvious benefit of chromatographic separation), the automated system according to the invention can achieve 10 nl / min or less. A low electrospray flow rate can be expected to increase the ionization efficiency up to 100%. A lower flow rate can be expected to provide a wider linear dynamic range, which is important for the analysis of complex mixtures-complexes due to the number of analytes that compete for ionization and their concentration range.

[0109] The automated system according to the present invention has a wide range of applications in the research fields of proteomics, peptidomics, and metabolomics. In particular, the automated system according to the present invention can be configured to transform the current extremely slow and low throughput neoepitope discovery to a higher throughput, for example, to rapidly test a library of predicted neoepitopes for a wide range of tumors. It can therefore help to facilitate the design of rational bed-to-bench-and-back-to-bed immunotherapies. Besides neoepitopes, the automated system according to the present invention can be applied to metabolomics, analysis of small molecules in drug discovery, and will certainly prove useful for analysis of therapeutic antibodies by intact protein mass spectrometry.

[0110] Further optional features and embodiments will be disclosed in more detail in the following description of the embodiments, preferably in conjunction with the dependent claims, in which each optional feature may be realized alone or in any possible combination, as will be understood by those skilled in the art. The scope of the present invention is not limited by the preferred embodiments. The embodiments are depicted diagrammatically in the figures, in which the same reference symbols in these figures refer to identical or functionally similar elements. [Brief description of the drawings]

[0111] [Figure 1A] 1 illustrates an exemplary embodiment of a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system according to the present invention. [Figure 1B] 1 illustrates an exemplary embodiment of a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system according to the present invention. [Figure 1C] 1 illustrates an exemplary embodiment of a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system according to the present invention. [Figure 1D] 1 illustrates an exemplary embodiment of a method for providing at least one sample in a mass spectrometer system according to the present invention and ionizing it by electrospray ionization. [Figure 1E] 1 illustrates an exemplary embodiment of a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system according to the present invention. [Figure 1F] 1 illustrates an exemplary embodiment of a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system according to the present invention. [Figure 1G] 1 illustrates an exemplary embodiment of a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system according to the present invention. [Figure 1H] 1 illustrates an exemplary embodiment of a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system according to the present invention. [Figure 2A] 1 illustrates various embodiments of data analysis and visualization. [Figure 2B] 1 illustrates various embodiments of data analysis and visualization. [Figure 2C] 1 illustrates various embodiments of data analysis and visualization. [Figure 2D] 1 illustrates various embodiments of data analysis and visualization. [Figure 2E] 1 illustrates various embodiments of data analysis and visualization. [Figure 3A] 1 shows the stability of the spray current at different settings of the static spray. [Figure 3B] The corresponding quality of the MS data is shown. [Figure 4] Optimization of the normalized collision energy NCE is shown. [Diagram 5] 1 shows cell plots illustrating the effect of optimizing field asymmetric ion mobility spectrometry (FAIMS) parameters on the sensitivity of peptide detection. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0112] 1A-1F show in schematic diagrams an exemplary embodiment of a method for providing and ionizing by electrospray ionization at least one sample in a mass spectrometer system 110 according to the present invention.

[0113] 1A, an exemplary embodiment of a mass spectrometer system 110 is shown. The mass spectrometer system 110 comprises at least one automation system 112 and at least one mass spectrometry analyzer 114. The mass spectrometry analyzer 114 has at least one ion source 116, specifically at least one ionization chamber 118. An emitter end 120 of an electrospray emitter 122 of the automation system 112 is disposed within the ion source 116.

[0114] The automation system 112 includes at least one electrospray emitter 122 including at least one emitter end 120 having at least one emitter tip and at least one fluid inlet end 126. The electrospray emitter 122 may specifically include at least one fused silica capillary 128. Specifically, the emitter tip 124 may be conically tapered at the emitter end 120. The emitter tip 124 includes an opening having a diameter of 1 μm to 10 μm. Specifically, the opening of the emitter tip 124 has a diameter of 3 μm or 5 μm. The electrospray emitter 122 may specifically be a CoAnn emitter provided by CoAnn Technologies. The electrospray emitter 122 may specifically be held by a holder 130. The holder 130 may specifically be a very high pressure zero dead volume (ZDV) union, specifically a ZDV union provided by IDEX Health & Science LLC.

[0115] Additionally, the automation system 112 includes at least one autosampler 132. The autosampler 132 includes at least one autosampler outlet 134. The autosampler is configured to provide at least one sample 136 having at least one analyte.

[0116] Additionally, the automated system 112 comprises at least one pipe 138, specifically at least one fused silica capillary 140, having at least one pipe fluid outlet end 142 and at least one pipe fluid inlet end 144. The pipe fluid inlet end 142 is fluidly connected to the autosampler outlet 134.

[0117] Furthermore, the automation system 112 comprises at least one liquid junction 146. The liquid junction 146 comprises at least one connection element 148 made of at least one electrically conductive material. The connection element 148 receives the fluid inlet end 126 of the electrospray emitter 122 and the pipe fluid outlet end 142 of the pipe 138 such that a fluid connection between the electrospray emitter 122 and the pipe 138 is established. The connection element 148 is electrically connectable to at least one voltage source 149. The connection element 148 may specifically comprise at least one hollow cylinder 150 having at least one first end 152 and at least one opposite second end 154. The fluid inlet end 126 of the electrospray emitter 122 may be received at the first end 152, and the pipe fluid outlet end 142 may be received at the second end 154. The first end 152 and the second end 154 are each sealed by a screw 156. The screw 156 may have a through hole 158 for receiving the electrospray emitter 122 and the pipe 138, respectively. The fluid inlet end 126 of the electrospray emitter 122 and the pipe fluid outlet end 142 of the connecting element 148 may be received in the connecting element 148, specifically such that a nanogap 160 is formed between the pipe fluid outlet end 142 and the fluid inlet end 126 of the electrospray emitter 122. Thus, the sample 136 may be charged by or at the nanogap 160 by applying a voltage to the connecting element 148 via a voltage source 149.

[0118] The autosampler 132 may include at least one sample tray 162 configured to provide at least one sample 136. The sample tray 162 may have a plurality of flasks 164 each configured to provide a sample 136 and, more particularly, to store a sample 136.

[0119] Additionally, the autosampler 132 may specifically include at least one first valve 166 having at least one sample loop 168 configured to receive the sample 136. Specifically, the first valve 166 may include at least one stationary element 170 including the sample loop 168 configured to receive the sample 136. Additionally, the first valve 166 may include at least one rotatable element 172 having a channel 174 configured to flow and / or direct the sample 136. Each port 176 may be connected to adjacent ports 176, specifically to the immediate right and to the immediate left, via one of the multiple channels 174.

[0120] Further, the autosampler 132 may specifically include at least one fluid loading device 178, specifically at least one syringe 180, configured to load the sample 136 provided by the sample tray 162 into the sample loop 168 of the first valve 166. Further, the autosampler 132 may specifically include at least one pump 182, specifically at least one nanopump 184. The pump 182 may be configured to transport the sample 136 from the sample loop 168 of the first valve 166 to the autosampler outlet 134 and into the electrospray emitter 122.

[0121] The first valve 166 may include at least one sample port 186, at least one fluid loading device port 188, at least one pump port 190, and at least one outlet port 192. The sample tray 162 may be fluidly connectable to the sample loop 168 of the first valve 166 via the sample port 186. The fluid loading device 178 may be fluidly connectable to the sample loop 168 of the first valve 166 via the fluid loading device port 188. The pump 182 may be fluidly connectable to the sample loop 168 of the first valve 166 via the pump port 190. The autosampler outlet 134 may be fluidly connectable to the sample loop 168 of the first valve 166 via the outlet port 192 and an inlet port 194 of a second valve 196, which may be further described in more detail below. The automation system 112 may be configured to load the sample 136 into the sample loop 168 of the first valve 166 by establishing a fluid connection between the fluid loading device port 188 and the sample port 186 via the sample loop 168. Additionally, the automation system 112 may be configured to inject the sample 136 into the pipe 138 by establishing a fluid connection between the pump port 190 and the outlet port 192 via the sample loop 168.

[0122] Additionally, the autosampler 132 may include at least one second valve 196. The second valve 196 may be configured to selectively fluidly connect the outlet port 192 of the first valve 166 to the autosampler outlet 134 or to at least one waste container 198. The second valve 196 may be configured to return the pump outlet 190 to the waste container 198 upon transfer of the sample 136 from the sample loop 168 of the first valve 166 to the pipe fluid inlet end 144 of the pipe 138. The second valve 196 may have a rotatable element 200 having a channel 202 configured to flow the sample 136. Each port 204 may be connected to adjacent ports 204, specifically to the immediate right and to the immediate left, via one of the multiple channels 202.

[0123] Additionally, the autosampler 132 may include at least one third valve 206. Additionally, the autosampler 132 may include at least one wash fluid container 208 configured to provide at least one wash fluid. The third valve 206 may be configured to selectively connect the fluid loading device 178 to the wash fluid container 208 or to the fluid loading device port 188 of the first valve 166.

[0124] As shown in FIG. 1A, in a first step of a method for providing at least one sample 136 in a mass spectrometer system 110 and ionizing it by electrospray ionization, the at least one sample 136 is provided to at least one sample tray 162 of the autosampler 132, specifically to at least one flask 164.

[0125] As shown in Figures 1B and 1C, in a second step of the method for supplying at least one sample 136 in a mass spectrometer system 110 and ionizing it by electrospray ionization, the sample 136 is transferred from the sample tray 162 to the autosampler outlet 134, whereby the sample 136 is further transferred to the electrospray emitter 122 via a pipe 138 and a liquid junction 146.

[0126] 1B, the sample loop 168 of the first valve 166 can be loaded with the sample 136 by the fluid loading device 178, as indicated by arrow 210. Specifically, a fluid connection can be established between the fluid loading device port 188 and the sample port 186 by the sample loop 168.

[0127] Specifically, as shown in FIG. 1C, the sample 136 may then be transferred from the sample loop 168 of the first valve 166 to the autosampler outlet 134. The pump 182 may be configured to push the sample as soon as the first valve 166 switches from the configuration depicted in FIG. 1B, in which the sample loop 168 is connected to ports 188-186, to the configuration depicted in FIG. 1C, in which the sample loop 168 is connected to ports 190-192. Thereby, as shown by arrow 212, the sample 136 may be further transferred via the pipe 138 and the liquid junction 146 to the electrospray emitter 122. Specifically, a fluid connection may be established between the pump port 190 and the outlet port 192 via the sample loop 168.

[0128] Additionally, different types of washing steps may be performed as shown in Figures 1D to 1H.

[0129] As shown in FIG. 1D, a first wash step is shown. The first wash step may also be referred to as a sample loop wash step. During the first wash step, the second valve 196 may fluidly connect the outlet port 192 of the first valve 166 to a waste container 198, and a fluid connection may be formed between the pump port 190 and the waste container 198 via the sample loop 168 and the outlet port 192.

[0130] In Figures 1E and 1F, a second washing step is shown. The second washing step may also be referred to as a fluid loading device priming step or a syringe priming step. As shown in Figure 1E, the fluid loading device 178 may be fluidly connected to a washing fluid container 208 via a third valve 206, as indicated by arrow 214, whereby washing fluid is transferred to an internal volume 216 of the fluid loading device 178. Furthermore, as shown in Figure 1F, the fluid loading device 178 may be fluidly connected to a waste container 222 via a third valve 206, whereby washing fluid is transferred from the internal volume 216 of the fluid loading device 178 to the waste container 222.

[0131] In FIG. 1G, a third washing step is shown. The third washing step may also be referred to as a sample needle washing step. The fluid loading device 178 may be fluidly connected to a washing fluid container 208 via a third valve 206, so that the washing fluid is transferred to the internal volume 216 of the fluid loading device 178 as shown in FIG. 1E, and then the needle 224 of the autosampler 132 may be placed into the needle seat 226 as shown in FIG. 1G. As further shown in FIG. 1G, the fluid loading device 178 may be fluidly connected to the needle 224 via a third valve 206, so that the washing fluid is transferred from the internal volume 216 of the fluid loading device 178 to the needle seat 226 via the fluid loading device port 188 of the first valve 166 and the sample port 186 of the first valve 166.

[0132] Figures 2A-2E show various embodiments of data analysis and visualization. The embodiments according to Figures 2A-2C correspond to "hand-made" schematic diagrams intended to explain the concept. The embodiments according to Figures 2A-2C do not correspond to measured data. The embodiments according to Figures 2D and 2E are experimental results and correspond to measured data.

[0133] To take full advantage of the ability to analyze peptides in batches, a program was developed to extract, analyze, and visualize the optimization results using a static spray approach.

[0134] Advantageously, the above programs are customized for the applications described within the scope of this application and are therefore able to generate results from the instrument output without additional user input, whereas other software typically requires manual adjustments for each peptide.

[0135] In FIG. 2A, a mass spectrum resulting from a liquid column separation is shown. Intensity I in arbitrary units corresponds to retention time t r The signal can shift further as the time axis progresses as the parameters are adjusted for optimization. In FIG. 2B, the mass spectrum obtained as a result of the static spray approach is shown. The intensity I in arbitrary units is plotted against the retention time t r It is shown that the dependence of the time (min) is shown. In static spray (ss), it can be seen that the signal is ideally constant and is only affected by noise and instrument parameters. In Fig. 2C it is shown that the ss signal above can be further compressed to represent what is relevant for the workflow. The different textures represent different characteristic signals / transitions / fragments generated by collisional dissociation of the target precursors / analytes / species.

[0136] In Figures 2D and 2E, it is shown that by cycling instrument parameters such as FAIMS CV (Figure 2D) or collision energy CE, the visualization allows the optimal parameter values ​​to be clearly identified, as indicated by mark 220.

[0137] Data extraction is very similar to what is commonly done using Skyline software (https: / / skyline.ms) and other software in the field, where the developed program calculates the ions produced upon fragmentation of each analyte peptide, facilitates data extraction with a strong focus on LC-MS, and visualizes the data as an ion chromatogram (Figure 2A). It is possible to extract the data as it is generated, although some friction is added and manual input is required. Overall, the above workflow is common, although most modern software strongly expects LC separation.

[0138] Once the instrument parameters have been cycled for optimization in static spray, it can be further limited by the software used and whether it supports reading the parameters. While most software is not set up for this these days, software has also been released by others to facilitate this via R scripting (https: / / github.com / fgcz / rawDiag#343--how-to-get-all-scan-attributes-assosiated-to-each-scan).

[0139] For data analysis and visualization, the results can be visualized as shown in Figures 2D and 2E. For most parameters, the identification of optimal conditions can be easy according to these plots. For collision energy optimization (Figure 2E), there can be further nuances with the collision energy parameter, which not only affects the overall signal intensity but also the signal composition. In the present example, some ions appear only at high energies.

[0140] The data collected in our automated approach can be used to probe several interesting scientific questions about what the best collision energy is for various applications.

[0141] 3A and 3B show the spray current c at different settings of the static spray. s (Figure 3A), and the corresponding quality of the MS data (Figure 3B).

[0142] Figure 3A shows the spray current c as a function of time t (min) for different static spray settings. s 3A shows the results of the static spray setup for the coated emitter. The data marked with a circle correspond to data from the static spray setup for the liquid junction and the uncoated emitter. The data with the cross symbol correspond to data from the static spray setup for the liquid junction and the uncoated emitter. The data with the cross symbol are acquired by the mass spectrometer system according to the invention. In FIG. 3A it can be seen that the spray current is more stable for the uncoated emitter in the presence of the liquid junction compared to the coated glass capillary, which corresponds to the classical approach of static spray.

[0143] In Fig. 3B the intensity I at au is shown in dependence on the normalized collision energy NCE. Identifying a clear optimum for the collision energy proves to be less efficient for coated emitters (see upper panel) compared to uncoated capillaries (see lower panel) in the presence of a liquid junction.

[0144] Figure 4 shows the optimization of the normalized collision energy NCE. The intensity I in au is shown in dependence on the normalized collision energy NCE. In the upper panel the complete peptide is represented. In the lower panel the different fragments y4+, y5+, y5++, y6+, y7+, y7++ are shown. The HLA-I associated 9-mer peptide (2+) shows the optimal NCE value (dotted line) which is lower than the default value (26-30%, dotted band) routinely used in peptidomics. The optimized NCE could improve the sensitivity of the detection of peptides in biological samples.

[0145] FIG. 5 shows cell plots showing the effect of optimizing parameters of field asymmetric ion mobility spectrometry (FAIMS) on the sensitivity of peptide detection, specifically of HPV-derived HLA-2 restricted target peptides. The upper panel relates to data without FAIMS and the lower panel to data with FAIMS. The FAIMS interface cannot be used in target MS without prior optimization of compensation voltages (CV) for each target peptide. The higher sensitivity of peptide detection in the presence of FAIMS (analysis based on optimized CV for each peptide) is clearly shown. The code of the cell plot follows the dot product (dotp, varying between 0-1), which corresponds to the normalized spectral contrast angle. dopt is a scoring of the similarity of the detected peptide fragment pattern as compared to an external library (reference spectrum generated by synthetic peptides). [Explanation of symbols]

[0146] 110 Mass Spectrometer System 112 Automation Systems 114 Mass Spectrometer Analyzer 116 Ion Source 118 Ionization Chamber 120 Emitter end 122 Electrospray Emitter 124 Emitter tip 126 Fluid inlet end 128 Fused Silica Capillary 130 Holder 132 Autosampler 134 Autosampler outlet 136 Samples 138 Pipe 140 Fused Silica Capillary 142 Pipe Fluid Outlet End 144 Pipe Fluid Inlet End 146 Liquid junction 148 Connection Elements 149 Voltage Source 150 Hollow Cylinder 152 First end 154 Second End 156 Screw 158 through hole 160 Nanogap 162 Sample Tray 164 flasks 166 First Valve 168 Sample Loops 170 Fixed elements 172 Rotatable Elements 174 channels 176 Port 178 Fluid Loading Device 180 Syringe 182 Pump 184 Nanopump 186 sample ports 188 Port for fluid loading device 190 Pump port 192 Exit Port 194 Inlet Port 196 Second Valve 198 Waste Container 200 Rotatable Elements 202 Channel 204 Port 206 Third Valve 208 Cleaning solution container 210 Arrow 212 Arrow 214 Arrow 216 Internal volume 218 Arrow 220 marks 222 Waste container 224 Needle 226 Needle seat

Claims

1. 1. An automated system (112) for providing at least one sample (136) for electrospray ionization in a mass spectrometer system (110), comprising: The automation system (112) at least one electrospray emitter (122) comprising at least one emitter end (120) having at least one emitter tip (124) and at least one fluid inlet end (126); at least one autosampler (132), said autosampler (132) comprising at least one autosampler outlet (134), said autosampler (132) configured to provide at least one sample (136) having at least one analyte; at least one pipe (138) having at least one pipe fluid outlet end (142) and at least one pipe fluid inlet end (144), said pipe fluid inlet end (144) being fluidly connected to said autosampler outlet (134); at least one liquid junction (146), the liquid junction (146) comprising at least one connecting element (148) formed of at least one electrically conductive material, the connecting element (148) receiving the fluid inlet end (126) of the electrospray emitter (122) and the pipe fluid outlet end (142) of the pipe (138) such that a fluid connection is established between the electrospray emitter (122) and the pipe (138), the connecting element (148) being electrically connectable to at least one voltage source; Equipped with the emitter tip (124) comprises an opening having a diameter of 1 μm to 10 μm; An automation system (112).

2. The automated system (112) of any preceding claim, wherein the opening in the emitter tip (124) has a diameter of between 3 μm and 5 μm.

3. 3. The automated system (112) of claim 1 or 2, wherein the pipe fluid inlet end (144) of the pipe (138) is directly fluidly connected to the autosampler outlet (134).

4. The automation system (112) of any one of claims 1 to 3, wherein the fluid inlet end (126) of the electrospray emitter (122) and the pipe fluid outlet end (142) of the pipe (138) are received in the connection element (148) such that a nanogap (160) is formed between the fluid inlet end (126) and the pipe fluid outlet end (142).

5. The connecting element (148) is at least one hollow cylinder (150) having at least one first end (152) and at least one opposing second end (154); Equipped with the fluid inlet end (126) of the electrospray emitter (122) is received in the first end (152); The pipe fluid outlet end (142) is received in the second end (154). The automation system (112) according to any one of claims 1 to 4.

6. The autosampler (132) at least one sample tray (162) configured to supply said at least one sample (136); at least one first valve (166) having at least one sample loop (168) configured to receive the sample (136); at least one fluid loading device (178), the fluid loading device (178) configured to load the sample (136) provided by the sample tray (162) into the sample loop (168) of the first valve (166); at least one pump (182) configured to transport the sample (136) from the sample loop (168) of the first valve (166) to the autosampler outlet (134) and to the electrospray emitter (122); Further comprising: The automation system (112) according to any one of the preceding claims.

7. the autosampler (132) includes at least one second valve (196); the second valve (196) is configured to selectively fluidly connect the outlet port (192) of the first valve (166) to the autosampler outlet (134) or to at least one waste container (198); The automation system (112) of claim 6.

8. the second valve (196) is configured to return the pump outlet of the first valve (166) to the waste container (198) once the sample (136) has been transferred from the sample loop (168) of the first valve (166) to the pipe fluid inlet end (144) of the pipe (138); The automation system (112) of claim 7.

9. A mass spectrometer system (110), comprising at least one automation system (122) according to any one of claims 1 to 8, further comprising at least one mass spectrometry analyzer (114) having at least one ion source (116), the emitter end (120) of the electrospray emitter (122) being disposed within the ion source (116).

10. A method for providing at least one sample (136) in a mass spectrometer system (110) and ionizing by electrospray ionization, the method comprising using an automated system (112) according to any one of claims 1 to 8, the method comprising: a) providing at least one sample (136) to at least one sample tray (162) of said autosampler (132); b) transferring the sample (136) from the sample tray (162) to an autosampler outlet (134), whereby the sample (136) is further transferred to the electrospray emitter (122) via the pipe (138) and the liquid junction (146); c) charging at least one component of the sample (136) by applying a voltage to the connection element (148) of the liquid junction (146) via a voltage source; Including, method.

11. the pressure inside the electrospray emitter (122) during electrospray ionization is atmospheric pressure, or The pressure inside the electrospray emitter (122) during electrospray ionization is between 5 bar and 15 bar. The subject matter of any one of claims 1 to 10.

12. 1. A method for optimizing at least one mass spectrometry (MS) parameter, the method comprising: (A) providing at least one sample (136) and ionizing it by electrospray ionization; (B) recording at least two MS spectra under conditions including at least two different values ​​for said at least one MS parameter; (C) comparing the MS spectra recorded in step (B); (D) thereby optimizing the at least one MS parameter based on the comparison in step (C); and Including, method.

13. 1. A method for detecting at least one analyte in a sample by mass spectrometry (MS), the method comprising: (A) providing and ionizing a sample (136) using an automated system (112) according to any one of claims 1 to 8 for a method for providing and ionizing at least one sample; (B) recording at least one MS spectrum of the ionized sample of step (A); Including, method.

14. an automated system (112) for optimizing at least one MS parameter; Use of an automation system (112) according to any one of claims 1 to 8 in relation to a mass spectrometer system (110) and / or use of a mass spectrometer system (110) according to claim 9 in relation to a mass spectrometer system (110).

15. an automated system (112) for detecting at least one analyte in a sample (136); Use of an automation system (112) according to any one of claims 1 to 8 in relation to a mass spectrometer system (110) and / or use of a mass spectrometer system (110) according to claim 9 in relation to a mass spectrometer system (110).

Images