Methods for determining ms reagent parameters via leachable compounds

Abstract

The present invention relates to a method for determining a mass spectrometry (MS) reagent stored in a storage container for a period of time and / or at least one storage parameter thereof, said method comprising (a) determining at least one leachable compound of said storage container; (b) comparing the at least one leachable compound determined in step (a) to a reference, and (c) determining said MS reagent and / or at least one storage parameter thereof based on the comparing in step (b); The present invention also relates to a method of quality assurance of an MS analysis, to a data carrier, an MS system, and a kit related thereto.

Description

[0001] Methods for determining MS reagent parameters via leachable compounds

[0002] The present invention relates to a method for determining a mass spectrometry (MS) reagent stored in a storage container for a period of time and / or at least one storage parameter thereof, said method comprising (a) determining at least one leachable compound of said storage container; (b) comparing the at least one leachable compound determined in step (a) to a reference, and (c) determining said MS reagent and / or at least one storage parameter thereof based on the comparing in step (b); The present invention also relates to a method of quality assurance of an MS analysis, to a data carrier, an MS system, and a kit related thereto.

[0003] Mass spectrometry (MS) has been widely used for chemical analysis for its sensitivity and specificity. More recently, methods using MS instrumentation were provided also for clinical and diagnostic applications, e.g. for measuring biomarker concentrations in complex samples. In order to maintain sensitivity and specificity of MS measurement, high quality reagents have to be used. The necessary quality standards often are not reached by copycat reagents, so there is a need to identify MS reagents provided e.g. by an originator, and to differentiate them from copycat versions, which may be of unknown quality.

[0004] To that end, several methods comprising addition of tracer compounds to MS reagents were proposed, e.g. in WO 2018 / 115237 Al, WO 2018 / 115247 Al, WO 2018 / 115243 Al. Also, addition of identifier devices was proposed, e.g. in US 2017 / 0336428 Al. However, addition of tracers or identifiers may not always be possible or desirable in manufacture of MS reagents.

[0005] Thus there is still a need for improved methods for determining MS reagent parameters, in particular MS reagent identity. The technical problem underlying the present invention can be seen as the provision of means and methods for complying with the aforementioned needs. The technical problem is solved by the embodiments characterized in the claims and herein below. In accordance, the present invention relates to a method for determining a mass spectrometry (MS) reagent stored in a storage container for a period of time and / or at least one storage parameter thereof, said method comprising

[0006] (a) determining at least one leachable compound of said storage container, in an embodiment by MS;

[0007] (b) comparing the at least one leachable compound determined in step (a) to a reference, and

[0008] (c) determining said MS reagent, period of time, and / or at least one storage parameter thereof based on the comparing in step (b).

[0009] In general, terms used herein are to be given their ordinary and customary meaning to a person of ordinary skill in the art and, unless indicated otherwise, are not to be limited to a special or customized meaning. As used in the following, the terms “have”, “comprise” or “include” or any arbitrary grammatical variations thereof are used in a non-exclusive way. Thus, these terms may both refer to a situation in which, besides the feature introduced by these terms, no further features are present in the entity described in this context and to a situation in which one or more further features are present. As an example, the expressions “A has B”, “A comprises B” and “A includes B” may both refer to a situation in which, besides B, no other element is present in A (i.e. a situation in which A solely and exclusively consists of B) and to a situation in which, besides B, one or more further elements are present in entity A, such as element C, elements C and D or even further elements. Also, as is understood by the skilled person, the expressions "comprising a" and "comprising an" in an embodiment refer to "comprising one or more", i.e. are equivalent to "comprising at least one". In accordance, expressions relating to one item of a plurality, unless otherwise indicated, in an embodiment relate to at least one such item, in a further embodiment a plurality thereof; thus, e.g. identifying "a cell" relates to identifying at least one cell, in an embodiment to identifying a multitude of cells. Also, the term "plurality" relates to a multitude, in an embodiment at least two, in a further embodiment, in a further embodiment at least three in a further embodiment at least for, in a further embodiment at least five, of the indicated items.

[0010] Further, as used in the following, the terms "preferably", "more preferably", "most preferably", "particularly", "more particularly", "specifically", "more specifically" or similar terms are used in conjunction with optional features, without restricting further possibilities. Thus, features introduced by these terms are optional features and are not intended to restrict the scope of the claims in any way. The invention may, as the skilled person will recognize, be performed by using alternative features. Similarly, features introduced by "in an embodiment" or similar expressions are intended to be optional features, without any restriction regarding further embodiments of the invention, without any restrictions regarding the scope of the invention and without any restriction regarding the possibility of combining the features introduced in such way with other optional or non-optional features of the invention.

[0011] The methods specified herein below are in vitro methods. The method steps may, in principle, be performed in any arbitrary sequence deemed suitable by the skilled person, but in an embodiment are performed in the indicated sequence; also, one or more, in an embodiment all, of said steps may be assisted or performed by automated equipment. Moreover, the methods may comprise steps in addition to those explicitly mentioned above.

[0012] As used herein, the term "standard conditions", if not otherwise noted, relates to IUPAC standard ambient temperature and pressure (SATP) conditions, i.e. in an embodiment, a temperature of 25°C and an absolute pressure of 100 kPa; also in an embodiment, standard conditions include a pH of 7. Moreover, if not otherwise indicated, the term "about" relates to the indicated value with the commonly accepted technical precision in the relevant field, in an embodiment relates to the indicated value ± 20%, in a further embodiment ± 10%, in a further embodiment ± 5%. Further, the term "essentially" indicates that deviations having influence on the indicated result or use are absent, i.e. potential deviations do not cause the indicated result to deviate by more than ± 20%, in a further embodiment ± 10%, in a further embodiment ± 5%. Thus, “consisting essentially of’ means including the components specified but excluding other components except for materials present as impurities, unavoidable materials present as a result of processes used to provide the components, and components added for a purpose other than achieving the technical effect of the invention. For example, a composition defined using the phrase “consisting essentially of’ encompasses any known acceptable additive, excipient, diluent, carrier, and the like. In an embodiment, a composition consisting essentially of a set of components will comprise less than 5% by weight, in a further embodiment less than 3% by weight, in a further embodiment less than 1% by weight, in a further embodiment less than 0.1% by weight of non-specified component(s).

[0013] The term “determining” is understood by the skilled person and in an embodiment relates to ascertaining the identity and optionally quantity of a chemical compound, composition, or a parameter referred to herein. Thus, determining may comprise qualitatively, semi quantitatively, or quantitatively determining a compound, composition, or parameter by measuring at least one characteristic feature thereof by a method as specified herein. Characteristic features as referred to herein are features which characterize physical and / or chemical properties including biochemical properties of a composition of matter, in particular a chemical compound or composition, such as an MS reagent. Such properties include in particular molecular weight, ion formation, one or more m / z value(s), and / or fragmentation, as e.g. determined by mass spectrometry. Values for the aforesaid properties may serve as characteristic features and can be determined by MS techniques well known in the art and as indicated herein below. Moreover, a value determined may be any value which is derived from the values of the aforesaid parameter(s) or the aforesaid properties of a chemical compound or composition by standard operations comprising, e.g., mathematical calculations such as multiplication, division, or logarithmic calculus. In an embodiment, the value determined is a normalized value, e.g. normalized to a control, calibrator, and / or standard. Thus, in an embodiment, determining may comprise identifying a chemical compound or composition and / or may comprise quantifying a chemical compound or composition, or determining a value of a parameter, in an embodiment as specified herein below. In an embodiment, determining may be identification and / or measurement of a parameter e.g. a storage parameter such as a period of time and / or a storage condition, optionally comprising quantifying said parameter to obtain a value thereof.

[0014] In view of the description herein, the term "determining a leachable compound" is understood by the skilled person. In an embodiment, the term relates to detecting the presence of a leachable compound, in an embodiment in an amount above the detection limit of the detection system. It will be understood that the leachable compound may not have to be identified to be determined. In particular, in case a leachable compound determined in an MS reagent is a compound excluded from a pre-determined composition of the MS reagent, it may be sufficient if the presence of the leachable compound is detected, e.g. by its m / z value. The method and equipment used for determining the leachable compound is determined by the skilled person depending in particular on the identity of the leachable compound. Thus, determining a leachable compound may be performed by any method deemed appropriate by the skilled person, e.g. by an optical method such as by US / VIS spectroscopy, e.g. using a diode array detector, or by MS. In an embodiment, the leachable compound is detected by MS, in particular in case the identity of the leachable compound is unknown. In a further embodiment, the term determining relates to determining the identity of a leachable compound, i.e. in an embodiment is identifying a leachable compound, wherein said identifying may provide an identifier of a compound class the leachable compound belongs to, and / or, in an embodiment provides an identification of the compound as such. In a further embodiment, the term determining relates to quantifying a leachable compound; in accordance with the description herein above, the skilled person understand that a leachable compound may also be quantified but not identified, although, in an embodiment, the leachable compound may be quantified and identified. Thus, in an embodiment, the term relates to determining a correlation plot of a semi quantitative or quantitative measure of one or more signals obtained from a leachable compound comprised in an MS reagent by an MS detector including the m / z value of the ions that cause said signals. A graphical representation of a mass spectrum may be provided e.g. as a centroid graph and / or as a continuum graph. As the skilled person understands in view of the description herein, determining a leachable compound may be determining a group of leachable compounds, in an embodiment providing the same or similar signal(s) in the detection method used; thus, determining a leachable compound may be determining a multitude of leachable compounds and does not necessarily have to comprise determining a single species of leachable compound. Thus, a leachable compound may be determined e.g. as an (apparent) increase of background, e.g. in a chromatogram.

[0015] As will be appreciated by the skilled person in view of the description herein, the methods specified may be used to detect deviations in composition of MS reagents from pre-determined requirements, and / or deviations of MS reagent storage conditions and / or storage period of time from pre-determined requirements. Thus, the methods may in particular be auxiliary methods of quality assurance in MS analysis of samples. In accordance, the methods in an embodiment are performed using the same equipment as is used for sample analysis. As a consequence, the skilled person in an embodiment may select leachable compounds and fragments and / or ions thereof such that they are detectable by the specific equipment used also for sample analysis. It is, however, also envisaged that the methods are implemented on dedicated equipment, e.g. in incoming goods inspection departments.

[0016] The term "mass spectrometry", which may be abbreviated as "MS", is known to the skilled person. In mass spectrometry, analytes in a sample are ionized in order to generate charged molecules or molecule fragments. Afterwards, the mass-to-charge value of the ionized analyte or fragments thereof is measured. Ionization of molecules can be carried out by any method deemed appropriate, in particular by electron impact ionization, fast atom bombardment, electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI), matrix assisted laser desorption ionization (MALDI). Mass spectrometry as referred to herein encompasses all techniques which allow for the determination of the molecular weight (i.e. the mass) or a mass variable corresponding to an analyte to be determined in accordance with the present invention. In an embodiment, mass spectrometry is used in combination with a chromatographic separation step, in particular as gas chromatography mass spectrometry (GC-MS), liquid chromatography mass spectrometry (LC-MS), direct infusion mass spectrometry or Fourier transform ion-cyclotrone-resonance mass spectrometry (FT-ICR-MS), capillary electrophoresis mass spectrometry (CE-MS), high-performance liquid chromatography coupled mass spectrometry (HPLC-MS), quadrupole mass spectrometry, any sequentially coupled mass spectrometry, such as MS-MS or MS-MS-MS, inductively coupled plasma mass spectrometry (ICP-MS), pyrolysis mass spectrometry (Py-MS), ion mobility mass spectrometry, or time of flight mass spectrometry (TOF). How to apply these techniques is well known to the person skilled in the art. Moreover, suitable devices are commercially available.

[0017] The term "mass spectrometry reagent", which may also be referred to as "MS reagent", includes each and every reagent used in a mass spectrometry analysis of a sample. The MS reagent may be liquid, gaseous, or solid, in an embodiment is liquid or gaseous, in a further embodiment is liquid. In an embodiment, the MS reagent is a calibration reagent, e.g. a calibration solution; a standardization reagent, e.g. a standardization solution, an eluent, a diluent, or a sample pretreatment reagent. However, also other MS reagents may be determined, e.g. mass axis check solutions, internal or external standard stock solutions, and the like. In an embodiment, the MS reagent is a sample container, thus a sample, e.g. a biological sample, may be an MS reagent as well. In an embodiment, the MS reagent is an MS sample reagent, i.e. an MS reagent applied together or instead of a sample to the MS, in an embodiment the chromatography-MS, device. Thus, in an embodiment, the MS reagent is a calibration reagent, e.g. a calibration solution; a standardization reagent, e.g. a standardization solution, a sample or standard diluent; a sample pretreatment reagent; a mass axis check solution; or an internal or external standard stock solution. In an embodiment, the MS reagent is an in vitro diagnostics reagent, in particular one as specified herein above.

[0018] In an embodiment, a pre-determined reference composition has been defined which the MS regent is required to have; i.e., in an embodiment, the method of the present invention is used to verify whether the composition of the MS reagent is in conformity with pre-defined requirements, e.g. quality requirements. Such pre-defined requirements may be presence, and optionally concentration, or absence, of specific constituents, in particular at least one solvent, impurity, and / or contaminant. Pre-defined requirements in an embodiment identify all ingredients of a reference composition, and optionally may specify chemical compounds which are excluded from the composition. In a further embodiment, pre-defined requirements identify all ingredients of a reference composition as well as their concentration(s), and in an embodiment further specify maximum concentration(s) of one or more chemical compounds, e.g. contaminants. Said concentrations may be provided as target concentrations, in an embodiment with indication of tolerable deviations, as concentration ranges, as upper and / or lower concentration limits, and the like. Said concentrations may also be provided as relative concentrations, e.g. as ratio of a leachable compound to a constituent of the MS reagent, as a ratio of two leachable compounds, as a maximal percentage of a leachable compound, and the like.

[0019] The term "solvent" is understood by the skilled person and includes any and all chemical compounds deemed suitable for solubilization of a solute, e.g. an analyte or sample constituents in general. In an embodiment, the solvent is water or an organic solvent like methanol, ethanol, acetonitrile, acetone, or a mixture of any of the aforesaid compounds. For the avoidance of doubt, the solvent as referred to herein does not necessarily have to comprise a solute, i.e. the solvent may also be a pure solvent or mixture thereof.

[0020] The MS reagent may further comprise any auxiliary compound(s) deemed appropriate by the skilled person, e.g. pH regulators such as one or more acid(s), base(s) and / or buffer compound(s), one or more pH indicator(s), complexing agents such as EDTA and EGTA, one or more salt(s), and the like. The pH regulator may define the pH of the MS reagent, e.g. in a range of from pH 1 to pH 14, in an embodiment of from pH 1 to pH 4, in a further embodiment of from pH 4 to pH 8, in a further embodiment of from pH 8 to 14.

[0021] The term "impurity" is known to the skilled person; in an embodiment, the term relates to a chemical substance present in a chemical compound as a result of the production process which is not the chemical compound itself. Typically, concentrations of impurities are low; however, depending on the production process of a chemical compound and the care it is performed with, impurities may be substantial. "Contaminants", as the term is referred to herein, are chemical substances present in a chemical compound or composition which are not impurities and which are not included intentionally. In accordance, leachable compounds determined according to the instant invention are contaminants; contaminants may, however, also be e.g. spoilage microorganisms and / or their metabolic products, chemical compounds included by lack of care in production and / or handling, by intentional inclusion, e.g. as a replacement for a more costly ingredient, and the like.

[0022] The term "storage container" is used herein in its conventional meaning attributed to it by the skilled person. Thus, the storage container may be any object deemed suitable for storing at least one MS reagent by the skilled person. Typically, the storage container comprises a container body having a cavity to receive the MS reagent. Optionally, the storage container further comprises a fastening element, such as a lid, a valve, or the like. The storage container will be comprised of at least one solid material (storage container material), wherein the container body and the optional fastening element may be of the same or different material(s). The storage container may also be comprised of more than one material, e.g. in case of a layered structure of the storage container. In accordance, the storage container may be comprised of a plurality of layers of materials, and under normal use, e.g. only the innermost layer may contact the MS reagent; however, in case the integrity of the innermost layer is disturbed, e.g. by corrosion or by an impact, the one or more non-innermost layer(s) may be contacted by the MS reagent as well. In an embodiment, the storage container is a flask, a bottle, a tube, a syringe, a plastic bag, or any storage container deemed appropriate by the skilled person.

[0023] As used herein, the term "storage container material" relates to any and all materials deemed appropriate by the skilled person for production of a storage container. As used herein, the term, when used in the context of a specific storage container, relates to the specific material(s) the specific container is comprised of. In an embodiment, the storage container material is a polymer, in an embodiment an organic polymer, in a further embodiment is a polyethylene (PE), e.g. a high-density PE (HDPE) or a low-density PE (LDPE), a polycarbonate, a polypropylene, or a halogenoalkane-polymer, such as polyvinylchloride (PVC), polytetrafluoroethylene (PTFE) or a perfluoroalkoxy alkane (PF A) polymer. The storage container material may also be a metal, such as aluminum, steel, or any other material deemed appropriate by the skilled person. The storage container material may also be a glass, in an embodiment borosilicate glass, in a further embodiment lime glass, in a further embodiment soda-lime-silica glass. In particular, layered storage containers may also comprise a plurality of the aforesaid storage container materials, e.g. layers of two or more different types of plastic, plastic-coated aluminum, or the like. Appropriate storage container material(s) is / are selected by the skilled person as deemed appropriate and suitable containers and their materials for MS reagents are known in the art; also, for specific MS reagents, e.g. comprising solvents or mixtures thereof, recommendations are available as to which containers and their materials are recommended for use.

[0024] The MS reagent is stored in the storage container under a set of storage parameters, in particular for a period of time and under storage conditions.

[0025] The term "storage parameter" is used herein to relate to any and all, in an embodiment measureable, parameters an MS reagent in a storage container is exposed to; thus, a storage parameter may be a period of time of storage or a storage condition, both as specified herein below.

[0026] The term "period of time", which may also be referred to as "period of time of storage", includes, in principle, any time frame starting with contacting the MS reagent with the storage container, e.g. by infilling, and ending with withdrawal of the MS reagent or an aliquot thereof from the storage container. In an embodiment, said period of time is at least 1 day, in an embodiment at least 3 days, in a further embodiment five days, in a further embodiment at least 7 days, in a further embodiment at least 2 weeks. The maximum period of time will be determined, among other factors, by the stability of the MS reagent in the storage container and by the stability of the storage container material when contacted with the MS reagent; thus, the maximal period of time may be several weeks, as may be the case for some organic solvent / plastic combinations, several months, or may be essentially unlimited. In an embodiment, the maximal period of time is defined by the use by date of the MS reagent.

[0027] The term "storage condition", as used herein, relates to a condition the MS reagent is exposed to over a period of time. In an embodiment, a storage condition is any measureable parameter which is known or assumed to influence the composition of the MS reagent in the storage container. Thus, the storage condition in an embodiment is selected from the list consisting of a storage container material, a storage temperature, a light exposure, a humidity, a vibration or other movement, a pressure, ionizing radiation, and a magnetic field(s). It will be understood from the description herein that a storage condition may have an effect on the MS reagent, e.g. on solubility of leachable compounds therein, on the storage container material, e.g. diffusibility of leachable compounds, or on both the MS reagent and the storage container material; e.g. an increase in temperature may increase leachability of leachable compounds in the storage container material, but may also increase the solubility of leachable compounds in the MS reagent. Storage conditions may e.g. be standard conditions or SATP conditions, all of which are known to the skilled person.

[0028] The term "leachable compound" is, in principle, known in the art. In an embodiment, the term relates to a compound which can be extracted from a storage container material into a solvent, in an embodiment into an MS reagent. In accordance, the leachable compound may, in principle, be any organic or inorganic compound. The leachable compound may be a compound known to be leachable from a storage container during storage of an MS reagent; the leachable compound may, however, also be a compound newly identified in an MS reagent after storage being indicative of incorrect storage conditions, e.g. by being absent from said MS reagent before storage and / or after correct storage. In an embodiment, the leachable compound is detectable by MS, in a further embodiment, the leachable compound is a compound extractable by the MS reagent from the storage container at an amount above the detection limit of MS, in an embodiment within the specified period of time. In a further embodiment, the leachable compound is a low molecular mass compound, in an embodiment with a molecular mass of at most 2 ku, in a further embodiment at most 1 ku, in a further embodiment at most 0.5 ku. The leachable compound may be present in a storage container material as a result of its production process. The leachable compound may, however, also be added to the storage container material, e.g. for identification purposes and / or for the purpose of the applications described herein. As the skilled person understands, such a leachable compound added to a storage container material may be a compound present in a storage container material which is added to increase its concentration. A leachable compound added to a storage container material may, however, also be a compound not present in the storage container material as a result of its production process; in such case, the leachable compound may be a chemical compound not present in the educts used in the production process of the storage container material; it may, however, also be an isotopically labeled variant of a compound present in the storage container material as a result of its production process, e.g. an isotopologue or an isotopomer. As the skilled person understands, the leachable compound may also be an isotopically labeled variant of a compound not present in the storage container material as a result of its production process. As will also be understood by the skilled person, in the context of adding a leachable compound to a storage container material, the term "adding" relates to any activity causing the leachable compound to be present in and / or associated with the storage container material, i.e. the leachable compound may be admixed to the storage container material or an educt thereof at any time during the production process deemed appropriate by the skilled person, may be included by diffusion, e.g. by contacting the storage container material or parts thereof with an admixture comprising the leachable compound, may be added as a layer on the storage container material, e.g. by impregnation, sputtering, or the like, or, in the case of a storage container having more than one layer, by including the leachable compound between two, identical or non-identical, layers of storage container material.

[0029] In an embodiment, in case the storage container is comprised of a plurality of layers, the leachable compound in an embodiment is not comprised in an innermost layer of the storage container; i.e. in such case the leachable compound may be a compound comprised only in a non-innermost layer. In a further embodiment, in case the storage container comprises a fastening element comprised of a material non-identical to the storage container body, the leachable compound may be a compound comprised only in the storage container body or only comprised in the storage container fastening element; the latter case may e.g. allow to identify storage in an incorrect orientation of the storage container, e.g. tilted or upside down. Thus, in an embodiment the storage container is comprised of a plurality of elements and the leachable compound is not comprised in at least one element of the storage container. The leachable compound in an embodiment depends on the storage container material(s). In case the storage container material is a metal or an alloy, the leachable compound may in particular be a metal ion comprised in the metal or alloy or generated therefrom by e.g. corrosion. In an embodiment, in case the storage container material is a polymer, in particular an organic polymer, the leachable compound comprises at least one monomer or oligomer from which the polymer was produced, the term oligomer in an embodiment relating to compounds comprising at least two monomer units, in an embodiment of from two to ten monomer units, in a further embodiment of from two to seven monomer units, in a further embodiment of from two to five monomer units. The leachable compound may, however, also be a softener, processing aid, or other adjuvant comprised in the storage container material. Standard polymers for production of storage containers, as well as softeners, processing aids, or other adjuvants, if used, as well as corresponding monomers used for their production, are known to the skilled person from standard textbooks. Typical storage container materials have been described herein above. The term “reference” refers to data of characteristic features of at least one leachable compound, which can be correlated to an MS reagent and / or storage parameter. Such a reference is, in an embodiment, obtained from a reference reagent stored under a set of pre-determined storage parameters, in particular for a period of time under a pre-determined set of known storage conditions in a storage container made of a pre-determined storage container material. In an embodiment, the reference reagent is a reagent having essentially the same composition as the MS reagent and / or the same composition the MS reagent is presumed to have. In an embodiment, the reference reagent comprises, in an embodiment consists of, the same solute(s) as the MS reagent and / or the MS reagent is presumed to consist of. Thus, in case the MS reagent is a solution of a chemical compound in a solvent or mixture thereof, the reference reagent may comprise an isotopologue of said chemical compound, or may lack the chemical compound. How to provide a suitable reference is in principle well known in the art, so the skilled person knows how to establish a reference. The data comprised in the reference in an embodiment, are at least one m / z value of at least one leachable compound, or a value derived therefrom, such as a molecular mass and / or a compound identifier, and optionally a quantitative parameter thereof, such as an intensity, in an embodiment allocated to a period of time and / or a set of environmental parameters. It will be understood that references for leachable compounds in an embodiment are determined under the same conditions as the leachable compounds in the MS reagent, i.e. in particular the same amount of sample is analyzed, the same sample pretreatment^) is(are) applied, the same sample separation, if any, is applied, and the MS system and conditions in an embodiment are the same. This in particular in case the leachable is not identified as such. In cases where the leachable compound is identified, and / or wherein the MS system is calibrated, the aforesaid conditions may deviate from those used for determining the leachable compound in the MS reagent. In an embodiment, the reference is a determination of the at least one leachable compound made with an MS reagent (i) stored in said storage container for less than 1 d, in an embodiment for less than 12 h, (ii) stored in an inert container essentially free of leachable compounds, in an embodiment a glass storage container, and / or (iii) stored in a reference container known or suspected to be comprised of the same material as the storage container.

[0030] It will be understood that the specific reference will, among others, be selected by the skilled person depending on the specific determining intended: in an embodiment, in case correct production, packaging, and / or storage of an MS reagent shall be verified, e.g. in quality assurance applications, it may be sufficient to provide a reference comprising data on permissible leachable compounds and, optionally, their maximum amounts. In a further embodiment, e.g. in case a storage container material shall be identified, it may be envisaged to provide a reference based on a plurality of candidate storage container materials for storage. In a further embodiment, e.g. in case a storage temperature shall be identified, it may be envisaged to provide a reference based on a plurality of candidate storage parameters, e.g. periods of time and / or temperatures. In a further embodiment, e.g. in case an MS reagent shall be identified, it may be envisaged to provide a reference based on a plurality of candidate MS reagents.

[0031] In the reference or references, the leachable compound may be identified, e.g. via an m / z value of the leachable compound or a fragment thereof, by a compound identifier, or the like. However, in some embodiments, identification of a leachable compound may be dispensable; e.g. in verification of correct production, packaging, and / or storage of an MS reagent, it may be sufficient to specify one or more m / z value(s) or range(s) thereof (i) which are required or (ii) which are excluded in order for the MS reagent or an analysis result obtained therewith to be acceptable. Also, in e.g. identification of a storage container material, it may be sufficient to identify one or more m / z value(s) of a leachable compound or a pattern of such m / z signals, without identifying the leachable compound(s) causing said m / z signals as such.

[0032] The term “comparing” is understood by the skilled person. In an embodiment, the term encompasses comparing a value of a parameter of a leachable compound determined in an MS reagent with a value of said parameter in a reference as specified elsewhere herein. It is to be understood that comparing as used herein refers to any kind of comparison made between the values determined in the MS reagent and the reference. However, in an embodiment, identical types of values are compared with each other, e.g., if an absolute amount is determined, the reference shall also be an absolute amount, if a relative amount is determined, the reference shall also be a relative amount, if an ion is determined after fragmentation, it shall be compared to a reference ion determined after fragmentation, etc. As set forth above, it may also be envisaged to calculate a score based on one or more parameter value(s), in particular a single score, and to compare this score to a reference score. The calculated score in an embodiment combines information on the amounts of leachable compound(s), may, however, also be calculated based e.g. on a multitude of m / z values, i.e. may comprise, exclusively or in addition, qualitative information on the leachable compound. Moreover, in the score, the parameter values may be weighted in accordance with their contribution to the establishment of the determining, wherein the weighting factors of the individual parameters may be different. When using a scoring system as described herein, values of different dimensions or units for the leachable compound(s) may be used, since the values will be mathematically transformed into the score. Accordingly, e.g. values for absolute concentrations may be combined in a score with peak area ratios and / or intensity values. In view of the description herein above, comparing may also comprise comparing presence or absence of one or more signals of a leachable compound to a reference, e.g. by comparing a pattern of m / z peaks determined in the MS reagent to a reference pattern. As specified in more detail herein above, comparing may neither require quantifying nor identifying a leachable compound.

[0033] The method comprises step (a) determining at least one leachable compound of said storage container, in an embodiment by MS. Typical methods of such determining and exemplary parameters which may be determined have been described herein above. In an embodiment, a leachable compound or a multitude thereof is determined by performing one or more MS measurement(s) on the MS reagent, typically on an aliquot thereof. The MS reagent or aliquot thereof may be pre-treated before being used in the determining step. Said pre-treatment may include treatments required to separate or enrich a leachable compound or to remove excessive material or waste. Suitable techniques are known in the art and in particular comprise centrifugation, extraction, fractioning, ultrafiltration, ultrasonic treatment, precipitation, optionally followed by filtration and purification and / or enrichment of leachable compounds. Moreover, other pre-treatments may be carried out in order to provide the leachable compound in a form or concentration suitable for the intended determination. Suitable and necessary pretreatments depend on the means used for carrying out the method and are well known to the person skilled in the art. MS determination may be carried out in any embodiment deemed appropriate by the skilled person. Thus, positive ion mode and / or negative ion method may be used, m / z windows may be adjusted to expected leachable compounds, optional collision energies may be selected to optimize detection of one or more leachable compounds, and the like. As the skilled person understands, the determining in step (a) in an embodiment is performed essentially identical, in a further embodiment identical, to the determination of the reference.

[0034] The method also comprises step (b) comparing the at least one leachable compound determined in step (a) to a reference. Methods for performing the comparing step have been described herein above. The comparison may be carried out manually or computer assisted. The comparing step may also encompasses comparing a calculated score with a suitable reference score. The value of the leachable compound and the reference can be, e.g., compared to each other and the said comparison can be automatically carried out by a computer program executing an algorithm for the comparison. The computer program carrying out the said evaluation will provide the desired assessment in a suitable output format.

[0035] The method further comprises step (c) determining said MS reagent and / or at least one storage parameter thereof based on the comparing in step (b). The term "determining said MS reagent and / or at least one storage parameter", as used herein, relates to establishing the identity and / or quantity of at least one of said parameters. In an embodiment, the MS reagent is determined, the term "determining an MS reagent" relating to establishing or verifying the identity and / or composition of an MS reagent. Thus, in an embodiment, determining an MS reagent comprises confirming, or not, the identity of an MS reagent, i.e. by determining at least one leachable compound and comparing to a reference, it may be confirmed, or not, that the MS reagent is the MS reagent it is presumed to be, e.g. based on a labeling of the storage container. In a further embodiment, determining an MS reagent comprises establishing or verifying a composition of an MS reagent; thus, by determining at least one leachable compound and comparing to a reference, it may be established that the composition of the MS reagent conforms with a predetermined specification.

[0036] In a further embodiment, at least one storage parameter is determined, the term "determining at least one storage parameter" relating to determining, in an embodiment quantitatively, the period of time of storage and / or at least one storage condition of an MS reagent. Thus, by determining at least one leachable compound and comparing to a reference, a time period of storage may be determined; or a storage condition, in an embodiment as specified herein above, may be determined. Thus, e.g. at least one of a storage container material, storage temperature, light exposure, humidity, vibration or other movement, pressure, ionizing radiation, and magnetic field(s) during storage may be determined. In an embodiment, determining a storage container material is identifying said storage container material; determining a storage container material may, however, also only be determining that the storage container material does not correspond to a pre-defined requirement. In an embodiment, determining a storage parameter is qualitatively or semi-quantitatively determining said storage parameter, in an embodiment is determining whether said storage parameter was potentially detrimental to said MS reagent. Thus, a storage parameter does not necessarily have to be determined quantitatively, it may be sufficient to determine that the storage parameter was beyond a pre-determined threshold. Also, it may not even be necessary to determine which storage parameter was beyond a predetermined threshold; i.e., determining at least one leachable compound may be used as an integrated indicator for determining non-compliance with at least one of a multitude of predetermined storage parameters, which may potentially be detrimental to said MS reagent. In an embodiment, however, the storage parameter is determined quantitatively.

[0037] As shown herein in the Examples, e.g. Fig. 1 and 10, leachable compounds may be determinable already from data obtained in the course of determining an analyte, e.g. from a UV / VIS or MS chromatogram, e.g. an MRM intensity over time chromatogram. Thus, in an embodiment, the determining in step (a) is comprised in a step of determining an analyte in a sample. In accordance, determining at least one leachable compound may be based on the same data, e.g. chromatogram, which are used to determine an analyte. Thus, in an embodiment, a method for determining as specified herein may be comprised in and / or performed concomitantly to a determination of an analyte, i.e. may not require a dedicated MS run. Thus, in an embodiment, the leachable compound is selected such that the signal used for determining is close to, but in an embodiment not overlapping, the signal used for determining the analyte(s).

[0038] As is also evident from the Examples provided herein, a leachable compound comprised in a chromatography eluent, depending on chromatography conditions, in particular on the selected stationary phase and optional further eluent(s) or gradient(s) thereof, may elute from a chromatography column without binding to the stationary phase, i.e. may be washed over a stationary phase or may be retarded by the stationary phase; in case a second eluent used to generate an elution gradient does not comprise the leachable compound or comprises the leachable compound at a significantly different concentration, this may cause a baseline shift over the eluent gradient (cf. e.g. Fig. 4). The leachable compound may, however, also become transiently bound, e.g. adsorbed, to the stationary phase, and may cause a discernible peak in a chromatogram when eluted, e.g. by an eluent gradient (cf. e.g. Fig. 7 B,E). Also, as discussed herein above, the leachable compound(s) does / do not necessarily have to be identified. Thus, in particular in case determining the MS reagent and / or at least one storage parameter thereof is verifying that a pre-defined chromatography eluent was used and that pre-defined storage conditions were obeyed, the aforesaid baseline drift and / or additional peak(s) caused by a leachable compound may be compared to a corresponding reference for verification. Thus, the aforesaid baseline drift and / or one or more additional peak(s) caused by one or more leachable compound(s) may be used as a "method fingerprint", said method fingerprint, if being essentially identical to a pre-determined fingerprint from a reference method, being indicative that a chromatography eluent has the expected composition and / or was stored under pre-defined conditions. As will be understood by the skilled person in view of the description herein, in particular a chromatogram or a subsection thereof may be used as such a method fingerprint. As will also be understood by the skilled person, a reference range for chromatograms or sections thereof may be defined, e.g. defining minimal and / or maximal baseline values at one or more given points of a chromatogram and / or presence, and optionally minimal and / or maximal peaks heights, or absence of pre-defied peaks may be defined. As specified in more detail elsewhere herein, such a method fingerprint may be used in quality assurance to reject measurement results if pre-defined criteria are not met.

[0039] Advantageously, it was found in the work underlying the present invention that, depending on storage conditions and time of storage, leachable compounds from storage containers leach into MS reagents and thus can be used to identify storage container materials, MS reagent composition, and / or storage parameters. Thus, on an exemplary basis and with no limitation of the subject matter of the invention, the following embodiments may in particular be envisaged:

[0040] - With a pre-determined MS reagent and storage container combination, a determinable set of leachable compounds in an embodiment leach into the MS reagent. Thus, determining leachable compounds may be used to determine whether the correct storage container material was used and / or whether the MS reagent has the required composition. Thus, the method may be used to identify non-originator products.

[0041] - The method may be used to detect incorrect storage of an MS reagent, e.g. (i) in case the concentration of one or more leachable compounds exceeds a pre-determined threshold(s), it may be concluded that the MS reagent was stored for a too long period of time of storage and / or under inappropriate storage conditions, such as too high temperature; (ii) in case a leachable compound which is only comprised in a lid of storage container is determined in the MS reagent, it may be concluded that inappropriate storage was applied, e.g. in a tilted position instead of upright and / or under excessive vibration or other movement; (iii) in case leachable compounds from a non-innermost layer of the storage container material are detected, it may be concluded that the structural integrity of the storage container material was lost before or during storage, and / or (iv) in case a leachable compound which should not be present is detected and / or the concentration of one or more leachable compounds exceeds a pre-determined threshold(s), it may be concluded that the MS reagent was stored in an inappropriate container, e.g. a container not originally supplied or prescribed by the manufacturer.

[0042] - By quantifying leachable compounds, storage parameters may be quantified; e.g. by quantitatively determining leachable compounds, a (maximum) storage temperature may be determined. For mutually influencing parameters, e.g. period of time and temperature of storage, in an embodiment one may be pre-determined in order to determine the other. E.g. in case a (maximum) storage temperature shall be determined, it may be envisaged to determine period of time of storage by other means, e.g. by transport and storage protocols.

[0043] - In view of the above, the method may be an in-line method of quality assurance; i.e. during an analytical run, signals of leachable compounds within a range of signals determined for determining the analyte, may be used for detecting potentially detrimental storage conditions of one or more MS reagents. Thus, the method may e.g. included in an in vitro method of determining an analyte, in particular an in vitro diagnostic method, in an embodiment an automated in vitro diagnostic method.

[0044] The definitions made above apply mutatis mutandis to the following. Additional definitions and explanations made further below also apply for all embodiments described in this specification mutatis mutandis.

[0045] The present invention also relates to a method of verifying a composition of an MS reagent stored in a storage container for a period of time, the method comprising the steps of the method for determining an MS reagent and / or at least one storage parameter as specified herein above, wherein the reference in step (b) is a determination of at least one leachable compound in a reference reagent of predetermined composition stored in a container comprised of the same material as the storage container for said period of time, and wherein step (c) is verifying the composition of said MS reagent based on the comparing in step (b).

[0046] As discussed herein above, the means and methods described herein enable identifying a solvent or mixture of solvents by determining at least one leachable compound leached from a storage container by said solvent or mixture of solvents. As shown herein in the Examples, different solvents leach different leachable compounds and / or leach leachable compounds at different rates from the material of a storage container. Thus, e.g. in quality assurance, determining leachable compounds may be used to verify a pre-determined composition of an MS reagent. As discussed herein above, said verifying in an embodiment comprises determining presence or absence of one or more signal(s) of a leachable compound, compared to a reference. Thus, in case for an MS reagent a required leachable compound fails to be determined, and / or in case an excluded leachable compound is determined, the MS reagent in an embodiment fails composition verification.

[0047] As the skilled person will understand in view of the description herein, the above may be applied mutatis mutandis to verify a storage container material. Thus, the present invention also relates to a method of verifying a storage container material of a storage container of an MS reagent wherein said MS reagent is stored for a period of time, the method comprising the steps of the method for determining an MS reagent and / or at least one storage parameter as specified herein above, wherein the reference in step (b) is a determination of at least one leachable compound in a reference MS reagent of predetermined composition stored in a container comprised of reference storage container material for said period of time, and wherein step (c) is verifying the storage container material based on the comparing in step (b).

[0048] Also, the present invention relates to a method of identifying at least one storage container material of an MS reagent, the method comprising the steps of the method for determining an MS reagent and / or at least one storage parameter as specified herein above, wherein determining said storage condition of said MS reagent in step (c) is identifying said at least one storage container material.

[0049] Also, the present invention relates to a method of quality assurance of an MS analysis, comprising determining a storage condition of an MS reagent according to the method for determining an MS reagent and / or at least one storage parameter as specified herein above, verifying a composition of an MS reagent as specified herein above; identifying a storage container material of an MS reagent according to the method as specified herein above; and / or determining at least one storage parameter; and the further step of rejecting a result of an MS measurement in case it is determined that said composition of the MS reagent, storage container material, and / or storage parameter does not fulfill a predetermined specification.

[0050] The term "quality assurance", which may also be referred to as "QA", is understood by the skilled person. In an embodiment, the term includes any and all measures taken to ensure that a product provided meets pre-determined specifications, such as reliability, performance, design, and maintainability. In case of medical applications, in particular diagnostics, QA in particular incudes measures to ensure reliability of results, in particular correctness, sensitivity, specificity, and reproducibility. One of said measures in an embodiment is ensuring identity and quality of MS reagents. Thus, by the aforesaid method, e.g. copycat MS reagents, and / or MS reagents stored in storage container of inappropriate material and / or quality may be detected and rejected. However, also storage under inappropriate conditions or structural failure within a storage container may be detected.

[0051] Further, the present invention relates to a method of fingerprinting a storage container material, said method comprising

[0052] (A) incubating said storage container material with an MS reagent under a set of pre-defined environmental conditions for a period of time,

[0053] (B) determining at least one leachable compound of said storage container material, in an embodiment by mass spectrometry (MS);

[0054] (C) allocating data on leachable compounds obtained in step (B) to the storage container material and optionally the environmental conditions and / or duration of said period of time of step (A), thereby fingerprinting said storage container material.

[0055] The term "fingerprinting" is used herein in its conventional meaning. In an embodiment, the term includes any and all proceedings making a storage container material identifiable, i.e. in an embodiment, comprises providing data enabling identification of a storage container material, said data also being referred to as "storage container material fingerprint". In view of the description elsewhere herein, different storage container materials have different storage container material fingerprints; in an embodiment also different MS reagents may have different storage container material fingerprints, even if obtained with the same storage container material; and, in a further embodiment, different storage parameter values may have different storage container material fingerprints, even if obtained with the same storage container material. As referred to herein, fingerprinting a storage container material in an embodiment does not comprise identifying a storage container material based on said storage container material fingerprint, e.g. in cases where it only needs to be decided whether the storage container material is according to a pre-determined specification. In a further embodiment, fingerprinting a storage container material comprises identifying a storage container material based on said storage container material fingerprint. Also, the method of fingerprinting a storage container material may be a method of providing a reference in the method for determining an MS reagent and / or at least one storage parameter thereof described herein above. Thus, the storage container material fingerprint may be a reference as described herein above, in particular in case a storage container material shall be determined. In a further embodiment, the fingerprinting a storage container material may be used in a method of identifying a storage container material based on said storage container material fingerprint.

[0056] The term "data on leachable compounds" is understood by the skilled person in view of the description herein. Data on leachable compounds may in particular be data on ions generatable from said leachable compound(s) in MS, e.g. m / z values of leachable compound(s) and / or at least one fragment generatable from a leachable compound, at least one MS intensity value of at least one leachable compound, identity of at least one leachable compound, and the like; thus, data on leachable compounds may include data obtainable directly from an MS system, such as m / z or intensity values, but also data drivable therefrom, such as identity and / or chemical structure of a leachable compound, or concentrations thereof.

[0057] The method of fingerprinting comprises step (C) allocating data on leachable compounds obtained in step (B) to the storage container material and optionally the environmental conditions and / or duration of said period of time of step (A), thereby fingerprinting said storage container material. How data can be allocated, i.e. correlated, with e.g. a storage container material is known to the skilled person. In an embodiment, allocating comprises making a correlation between said data on leachable compounds and a storage container material discernible. Thus, said allocating may be put into practice in the form of a database, such as a database as specified herein below. However, any other allocating method deemed appropriate by the skilled person may be used.

[0058] Furthermore, the present invention relates to a database, in an embodiment tangibly embedded on a data carrier, comprising (i) at least one identifier of a reference reagent; and / or (ii) at least one identifier of a storage container material; allocated to data on at least one leachable compound and optionally a set of environmental conditions and / or a length of a period of time of incubation used to obtain said data on at least one leachable compound. The present invention also relates to a data carrier comprising tangibly embedded the aforesaid database.

[0059] The term "identifier" is used in its conventional meaning relating to any data making a composition of matter identifiable; said identifier may be a name, an indication of composition such as a recipe, a reference number, or the like providing sufficient information to allow identification. The identifier may, however, also be a pointer to further information comprising the compound identified, such as a hyperlink, a barcode, or the like.

[0060] The term "reference reagent" relates to a reagent, in an embodiment a solution, of known, i.e. in an embodiment pre-determined, composition which was used to establish the data on at least one leachable compound. In concurrence, the storage container material data included such as its identifier, are those of the storage container material used to establish the data on at least one leachable compound. Thus, in an embodiment, the data comprised in the database may e.g. be established by the method of fingerprinting a storage container material described herein above.

[0061] The database, in an embodiment, comprises further data, such as upper and / or lower detection limits, upper and / or lower acceptable amounts of one or more leachable compounds, acceptable ratios of at least one compound to at least one leachable compound, data related to further storage container materials, data relevant for plausibility checks, and the like. Thus, the database in an embodiment further comprises at least one predetermined specification which has to be fulfilled in order for a result of an MS measurement to be acceptable. In a further embodiment, the database comprises data on one or more assay methods to use, lot-specific data, e.g. storage parameters, and the like. In an embodiment, the database may be implemented in a single data storage medium or in physically separated data storage media being operatively linked to each other. In an embodiment, the database comprises a data collection on a suitable storage medium, in an embodiment tangible embedded thereon. Moreover, the database, in an embodiment, further comprises a database management system. The database management system is, in an embodiment, a network-based, hierarchical or object-oriented database management system. Furthermore, the database may be a federal or integrated database. In a further embodiment, the database will be implemented as a distributed (federal) system, e.g. as a Client-Server-System. In a further embodiment, the database is structured as to allow a search algorithm to compare a test data set with the data sets, in particular the data on leachable compounds, comprised by the data collection. Specifically, by using such an algorithm, the database can be searched for similar or identical data sets being indicative for, e.g., a storage container material (e.g. a query search). Thus, in an embodiment, if a data set of at least one leachable compound fulfilling the comparison criteria as detailed elsewhere herein can be identified in the database, the test data set will be associated with the e.g. the storage container material. Consequently, the information obtained from the database can be used, e.g., as a reference for the methods described elsewhere herein. The present invention also relates to an MS system comprising an MS measuring unit and a control unit, wherein said MS system further comprises a data carrier as specified herein above and / or wherein said control unit comprises tangibly embedded an executable code which, when executed on the control unit, causes the system to perform at least steps (a) and (b) of the method for determining an MS reagent and / or at least one storage parameter as specified herein above.

[0062] The term "mass spectrometry system", abbreviated as "MS system", is understood by the skilled person. In an embodiment, the term relates to a system configured for performing a mass spectrometry (MS); thus, the system, in an embodiment, comprises at least one MS unit. As used herein, the term “mass spectrometry unit”, in an embodiment, relates to a mass analyzer configured for detecting at least one analyte based on a mass to charge ratio of a leachable compound or a fragment thereof. In an embodiment, the MS unit is a tandem mass spectrometry (MS / MS) unit, in a further embodiment a triple quadrupole MS (QqQ-MS), in a further embodiment in Multiple Reaction Monitoring (MRM) mode. The MS system may further comprise at least one ionization source configured for generating molecular ions and for transferring the molecular ions into the gas phase. Ionization methods and appropriate ionization units are known in the art and have been described herein above.

[0063] The term "control unit" relates to any module comprised in an MS system adapted to perform at least steps (a) and (b) of the method for determining an MS reagent and / or at least one storage parameter as specified herein above. In an embodiment, the control module comprises an executable code which, when executed on the control unit, causes the system to perform said method steps. In an embodiment, the control unit comprises at least one microprocessor and optionally a memory unit and the aforesaid executable code is tangibly embedded on said microprocessor or optionally on said memory unit. The control unit may also comprise tangibly embedded the database as specified herein above, e.g. in the memory unit. The system may, however, also have a networking unit, such that the aforesaid database may also be accessed via a network connection on a server device.

[0064] In an embodiment, the MS system is a chromatography MS system, in particular a gas chromatography MS (GC-MS) system or a liquid chromatography MS (LC-MS) system, as specified herein above. Thus, in an embodiment, the system is configured for performing a combination of chromatography (e.g. LC or GC)) with mass spectrometry (MS). Thus, the system, in an embodiment, comprises at least one LC and / or GC unit, and at least one MS unit, wherein the LC and / or GC unit(s) and the MS unit are coupled via at least one interface. As used herein, the term “liquid chromatography (LC) unit”, in an embodiment, relates to an analytical module configured to separate one or more analytes of interest of a sample from other components of the sample via liquid chromatography, in an embodiment for detection of the one or more analytes with the mass spectrometry system. The term "gas chromatography" is understood by the skilled person; in an embodiment the same separation principles as for LC are applicable, however, the mobile phase being a gas in GC.

[0065] The MS system also may comprise further devices and units deemed appropriate by the skilled person, such as input and / or output units, sample receiving and / or sample handling units, reagent storage units, fluid connectors, and the like. In particular, the MS system may be a fully automated system. In an embodiment, the MS system is an in vitro diagnostic system, in an embodiment a fully automated vitro diagnostic system.

[0066] The present invention also relates to a kit comprising an MS reagent and a data carrier, both as specified herein above.

[0067] The term “kit” as used herein refers to a collection of the aforementioned components. The means are, in an embodiment, provided in a single container (i.e. a housing), in a further embodiment enabling common translocation, e.g. transport, of the components. The container also typically comprises instructions for carrying out at least one of the methods of the present invention. These instructions may be in the form of a manual or may be provided by a computer program code which is capable of carrying out or supports the performing the methods of the present invention when implemented on a computer or a data processing device, in particular on a control unit of an MS system. The computer program code may be provided on a data storage medium or device such as an optical storage medium (e.g., a Compact Disc) or directly on a computer or data processing device or may be provided in a download format such as a link to an accessible server or cloud, in an embodiment, the computer program code is comprised on the data carrier. The kit may also comprise further components which are necessary for carrying one of the methods described herein, may assist in doing so, or may provide further functions. The present invention also relates to a use of an MS system as specified herein above or a kit as specified herein above for detecting an analyte in a sample, wherein, in an embodiment, said detecting an analyte in a sample is an aid in diagnosing and / or surveilling treatment of disease.

[0068] The invention further discloses and proposes a computer program including computerexecutable instructions for performing the method according to the present invention in one or more of the embodiments enclosed herein when the program is executed on a computer or computer network. Specifically, the computer program may be stored on a computer-readable data carrier. Thus, specifically, one, more than one or even all of method steps a) to d) as indicated above may be performed by using a computer or a computer network, in an embodiment by using a computer program.

[0069] The invention further discloses and proposes a computer program product having program code means, in order to perform the method according to the present invention in one or more of the embodiments enclosed herein when the program is executed on a computer or computer network. Specifically, the program code means may be stored on a computer-readable data carrier.

[0070] Further, the invention discloses and proposes a data carrier having a data structure stored thereon, which, after loading into a computer or computer network, such as into a working memory or main memory of the computer or computer network, may execute the method according to one or more of the embodiments disclosed herein.

[0071] The invention further proposes and discloses a computer program product with program code means stored on a machine-readable carrier, in order to perform the method according to one or more of the embodiments disclosed herein, when the program is executed on a computer or computer network. As used herein, a computer program product refers to the program as a tradable product. The product may generally exist in an arbitrary format, such as in a paper format, or on a computer-readable data carrier. Specifically, the computer program product may be distributed over a data network.

[0072] Finally, the invention proposes and discloses a modulated data signal which contains instructions readable by a computer system or computer network, for performing the method according to one or more of the embodiments disclosed herein. In an embodiment, referring to the computer-implemented aspects of the invention, one or more of the method steps or even all of the method steps of the method according to one or more of the embodiments disclosed herein may be performed by using a computer or computer network. Thus, generally, any of the method steps including provision and / or manipulation of data may be performed by using a computer or computer network. Generally, these method steps may include any of the method steps, typically except for method steps requiring manual work, such as providing the samples and / or certain aspects of performing the actual measurements.

[0073] Specifically, the present invention further discloses:

[0074] A computer or computer network comprising at least one processor, wherein the processor is adapted to perform the method according to one of the embodiments described in this description, a computer loadable data structure that is adapted to perform the method according to one of the embodiments described in this description while the data structure is being executed on a computer, a computer program, wherein the computer program is adapted to perform the method according to one of the embodiments described in this description while the program is being executed on a computer, a computer program comprising program means for performing the method according to one of the embodiments described in this description while the computer program is being executed on a computer or on a computer network, a computer program comprising program means according to the preceding embodiment, wherein the program means are stored on a storage medium readable to a computer, a storage medium, wherein a data structure is stored on the storage medium and wherein the data structure is adapted to perform the method according to one of the embodiments described in this description after having been loaded into a main and / or working storage of a computer or of a computer network, and a computer program product having program code means, wherein the program code means can be stored or are stored on a storage medium, for performing the method according to one of the embodiments described in this description, if the program code means are executed on a computer or on a computer network. The present invention also relates to a method of verifying and / or controlling a chromatography gradient, said method comprising

[0075] (I) determining at least one leachable compound comprised in an eluate, in an embodiment by MS, at least at a first time point in said chromatography;

[0076] (II) comparing the at least one leachable compound determined in step (I) to a reference; and

[0077] (III) thereby verifying said chromatography gradient and / or, if necessary, re-adjusting eluent feed, thereby controlling said chromatography gradient.

[0078] The terms "verifying" and "controlling" are used herein in their meanings known to the skilled person. In an embodiment, verifying is confirming that pre-determined specifications are met, e.g. confirming that a chromatography gradient meets pre-defined specifications, in an embodiment of composition and optionally its change over time. Also in an embodiment, controlling is exercising directing influence, if necessary, e.g. on a chromatography gradient; thus, controlling typically comprises measuring a state of a gradient, comparing said state to a pre-defined reference state, i.e. desired state, and taking measures to direct the state of the gradient to a desired state if necessary.

[0079] The terms "eluent" and "eluate" are understood by the skilled person. In an embodiment, the eluent is the mobile phase of a chromatography system, but also a solvent used to generate such a gradient may be referred to as eluent. Thus, e.g. in case a gradient of from 0 to 100% methanol in water is used, both water and methanol, as well as their mixtures in the gradient are eluents. The eluate, in an embodiment, is the composition of matter leaving the column after chromatography, comprising eluent, at least one leachable compound, and optionally one or more sample constituent s).

[0080] The term reference has been specified herein above and was discussed also for the case of verifying that a pre-defined chromatography eluent was used and that pre-defined storage conditions were obeyed. Thus, the baseline drift and / or one or more additional peak(s) caused by one or more leachable compound(s) as discussed herein above, determined in a predetermined reference method may be used as reference(s), wherein essential concurrence of the determined leachable compound(s) with the reference leads to verification of the chromatography gradient, and / or wherein non-concurrence leads to the finding of a deviation from the pre-determined chromatography gradient. Similarly, essential concurrence of the determined leachable compound(s) with the reference in an embodiment leads to the conclusion that no directing influence is required, and / or non-concurrence may lead to the conclusion that directing influence is required. As indicated herein above, in particular a chromatogram or a subsection thereof may be compared to a reference chromatogram in step (II). As will also be understood by the skilled person, a reference range for chromatograms or sections thereof may be defined, e.g. defining minimal and / or maximal baseline values at one or more given points of a chromatogram and / or presence, and optionally minimal and / or maximal peaks heights, or absence of pre-defied peaks may be defined. As specified in more detail elsewhere herein, such a method for verifying a chromatography gradient may be used in quality assurance to reject measurement results if pre-defined criteria are not met; also, the method of controlling a chromatography gradient may be used to control or to assist in controlling a chromatography gradient. As the skilled person understands, the eluents used in chromatography in an embodiment are verified to contain a pre-defined amount of leachable compound(s) used for determination at a pre-determined concentration. In view of the description herein, the skilled person understands that said verification of content may be accomplished by measuring the leachable compound in the eluent by one of the methods described herein, or by storing the eluent in a pre-determined storage container under pre-determined storage conditions.

[0081] The leachable compound is determined at least at a first time point in the chromatography; i.e. at at least one point during a chromatography run is the method of verifying and / or controlling performed on the eluate or an aliquot thereof. The term "time point", in the context of chromatography described herein, is used in a wide sense as relating to a specific point in chromatography; thus, a time point may be an actual point in time after start of a chromatography run; it may, however, also be an elution volume. As the skilled person will understand, comparison to a reference in step (II) typically comprises comparing to said reference at a corresponding time point in the reference chromatography, in an embodiment, at the same time point. As will also be understood that the determining of a leachable compound and comparison to a reference may be performed a multitude of times for a chromatography, in a further embodiment is performed continuously over at least a part of a chromatography run. Steps (I) to (III) may, in principle be performed during or after a chromatographic run in case the method is a method of verifying a chromatography gradient. However, in case the method is a method of controlling a chromatography gradient, it is in particular foreseen that steps (I) to (III) are performed concomitant to the chromatographic run, in an embodiment as an in-line method. In view of the above, the following embodiments are particularly envisaged:

[0082] Embodiment 1 : A method for determining a mass spectrometry (MS) reagent stored in a storage container for a period of time and / or at least one storage parameter thereof, said method comprising

[0083] (a) determining at least one leachable compound of said storage container, in an embodiment by MS;

[0084] (b) comparing the at least one leachable compound determined in step (a) to a reference, and

[0085] (c) determining said MS reagent and / or at least one storage parameter thereof based on the comparing in step (b).

[0086] Embodiment 2: The method of embodiment 1, wherein said MS reagent is a liquid MS reagent.

[0087] Embodiment 3 : The method of embodiment 1 or 2, wherein said MS reagent is a calibration reagent, in an embodiment a calibration solution; a standardization reagent, in an embodiment a standardization solution, an eluent, a diluent, or a sample pretreatment reagent. Embodiment 4: The method of any one of embodiments 1 to 3, wherein determining an

[0088] MS reagent is verifying a composition of an MS reagent.

[0089] Embodiment s: The method of any one of embodiments 1 to 4, wherein said storage parameter is said period of time of storage or is a storage condition.

[0090] Embodiment 6: The method of any one of embodiments 1 to 5, wherein said storage condition is a material the storage container is comprised of (storage container material), storage temperature, light exposure, humidity, vibration or other movement, pressure, and / or magnetic field(s).

[0091] Embodiment 7: The method of embodiment 6, wherein said storage container material is a polymer, in an embodiment an organic polymer

[0092] Embodiment 8: The method of any one of embodiments 1 to 7, wherein said determining a storage parameter is qualitatively determining said storage parameter, in an embodiment is determining whether said storage parameter was potentially detrimental to said MS reagent. Embodiment 9: The method of any one of embodiments 1 to 8, wherein said determining is quantitatively determining said at least one storage parameter.

[0093] Embodiment 10: The method of any one of embodiments 1 to 9, wherein said period of time is at least 1 day, in an embodiment at least 3 days, in a further embodiment at least 7 days, in a further embodiment at least 2 weeks. Embodiment 11 : The method of any one of embodiments 1 to 10, wherein said leachable compound is a compound extractable by the MS reagent from the storage container at an amount above the detection limit of said MS within said period of time.

[0094] Embodiment 12: The method of any one of embodiments 7 to 11, wherein said leachable compound comprises at least one monomer from which the polymer was produced.

[0095] Embodiment 13: The method of any one of embodiments 1 to 11, wherein said leachable compound is a softener, processing aid, or other adjuvant.

[0096] Embodiment 14: The method of any one of embodiments 1 to 13, wherein said storage container is comprised of a plurality of layers and wherein said leachable compound is not comprised in an innermost layer of the storage container.

[0097] Embodiment 15: The method of any one of embodiments 1 to 14, wherein said storage container is comprised of a plurality of elements and wherein said leachable compound is not comprised in at least one element of the storage container.

[0098] Embodiment 16: The method of any one of embodiments 1 to 15, wherein said reference is a determination of said at least one leachable compound made with an MS reagent (i) stored in said storage container for less than 1 d, in an embodiment for less than 12 hours, (ii) stored in an inert container essentially free of leachable compounds, in an embodiment a glass storage container, and / or (iii) stored in a reference container known or suspected to be comprised of the same material as the storage container.

[0099] Embodiment 17: A method of verifying a composition of an MS reagent stored in a storage container for a period of time, the method comprising the steps of the method of any one of embodiments 1 to 16, wherein the reference in step (b) is a determination of at least one leachable compound in a reference reagent of predetermined composition stored in a container comprised of the same material as the storage container for said period of time, and wherein step (c) is verifying the composition of said MS reagent based on the comparing in step (b). Embodiment 18: A method of identifying at least one storage container material of an MS reagent, the method comprising the steps of the method of any one of embodiments 1 to 16, wherein determining said storage condition of said MS reagent in step (c) is identifying said at least one storage container material.

[0100] Embodiment 19: A method of fingerprinting a storage container material, said method comprising

[0101] (A) incubating said storage container material with an MS reagent under a set of environmental conditions for a period of time, (B) determining at least one leachable compound of said storage container material, in an embodiment by mass spectrometry (MS);

[0102] (C) allocating data on leachable compounds obtained in step (B) to the storage container material and optionally the environmental conditions and / or duration of said period of time of step (A), thereby fingerprinting said storage container material.

[0103] Embodiment 20: A method of quality assurance of an MS analysis, comprising determining a storage parameter of an MS reagent method of any one of embodiments 1 to 16, verifying a composition of an MS reagent according to the method of embodiment 17 and / or identifying a storage container material of an MS reagent according to the method of embodiment 18; and the further step of rejecting a result of an MS measurement in case it is determined that said storage condition, composition of the MS reagent, and / or storage parameter material does not fulfill a predetermined specification.

[0104] Embodiment 21 : A database, in an embodiment tangibly embedded on a data carrier, comprising (i) at least one identifier of a reference reagent; and / or (ii) at least one identifier of a storage container material; allocated to data on at least one leachable compound and optionally a set of environmental conditions and / or a length of said period of time of incubation used to obtain said data on at least one leachable compound.

[0105] Embodiment 22: The database of embodiment 21, wherein said database further comprises at least one predetermined specification which has to be fulfilled in order for a result of an MS measurement to be acceptable.

[0106] Embodiment 23 : A data carrier comprising tangibly embedded the database of embodiment 21 or 22.

[0107] Embodiment 24: An MS system comprising an MS measuring unit and a control unit, wherein said MS system further comprises a data carrier according to embodiment 23, and / or wherein said control unit comprises tangibly embedded an executable code which, when executed on the control unit, causes the system to perform at least steps (a) and (b) of the method of any one of embodiments 1 to 16.

[0108] Embodiment 25: The system of embodiment 24, wherein the database comprised on said data carrier is a database according to embodiment 22 and wherein a result obtained by the measuring unit is rejected in case at least one of said predetermined specifications is not fulfilled.

[0109] Embodiment 26: The MS system of embodiment 24 or 25, wherein said MS system is a gas chromatography MS (GC-MS) system or a liquid chromatography MS (LC-MS) system. Embodiment 27: The MS system of embodiment 26, wherein said MS system is a fully automated system.

[0110] Embodiment 28: The MS system of any one of embodiments 24 to 27 being an in vitro diagnostic system.

[0111] Embodiment 29: The MS system of any one of embodiments 25 to 28 configured to display a warning and / or not display said result obtained by the measuring unit in case said result is rejected.

[0112] Embodiment 30: A kit comprising an MS reagent and a data carrier according to embodiment 23.

[0113] Embodiment 31 : Use of an MS system according to any one of embodiments 24 to 29 or a kit according to embodiment 30 for detecting an analyte in a sample.

[0114] Embodiment 32: The use of embodiment 32, wherein said detecting an analyte in a sample is an aid in diagnosing and / or surveilling treatment of disease.

[0115] Embodiment 33: A method of verifying and / or controlling a chromatography gradient, said method comprising

[0116] (I) determining at least one leachable compound comprised in an eluate, in an embodiment by MS, at least at a first time point in said chromatography;

[0117] (II) comparing the at least one leachable compound determined in step (I) to a reference; and

[0118] (III) thereby verifying said chromatography gradient and / or, if necessary, re-adjusting eluent feed, thereby controlling said chromatography gradient.

[0119] Embodiment 34: The method of embodiment 33, having a feature of any one of embodiments 1 to 32.

[0120] All references cited in this specification are herewith incorporated by reference with respect to their entire disclosure content and the disclosure content specifically mentioned in this specification.

[0121] Figure Legends

[0122] Fig. 1 : MRM-MS determination of estradiol, using water stored in storage containers made of a variety of storage container materials as LC eluent: (A) A round H, (B) CB gray, (C) CB UVN SD, (D), Schott (i.e. borosilicate) glass, (E) control (water not stored); the expected MRM peak at 255.2 / 159.0 at 2.24 min retention time is indicated with a small black arrow on the x-axis, additional peaks are indicated by arrows, ranges with baseline shifts are indicated by circles; x- axes: retention time [min], y-axes: intensity [cps] .

[0123] Fig. 2: Full scan (m / z: 50.00 - 500.00) data in positive mode after long incubation time of water with signal as a function of time reveals additional peaks originating from polymer bottle material; data from LC-MS without sample application, x-axis: retention time [min], y-axis: intensity [cps]; P: storage in polymer bottle, G: storage in glass bottle.

[0124] Fig. 3: 2 D plot of m / z values of leachable compounds in water against retention time in min in negative ESI; (A) glass bottle, (B) polymer bottle (A round H); data from LC-MS without sample application.

[0125] Fig. 4: As in Fig. 2, but showing results of positive ESI mode (A) and negative ESI mode (B). Direct comparison of ‘inert’ material glass to respective polymer.

[0126] Fig. 5: Full scan data in positive ESI mode after long incubation time of methanol as eluent, analyzed at a specific retention time reveals high background peaks for plastic materials of bottles (A, CB UVN SD) and (B, A round H) compared to glass (C); data from LC-MS without sample application, x-axes: m / z, y-axes: intensity [cps].

[0127] Fig. 6: As in Fig. 3, but showing leachable compounds in methanol; data from LC-MS without sample application.

[0128] Fig. 7: As in Fig. 1, but using acetonitrile as LC eluent and performing MRM for a variety of analytes: (A)-(C) A round H, (D)-(F) CB type A, (G)-(I) glass, (J)-(L) control (not incubated); (A), (D), (G), (J): measurement of 13 C3 -progesterone [M+H]+; transition 318.2 / 99.9; (B), (E), (H), (K): measurement of 13C3-cortisol [M+H]+; transition 366.1 / 124.1 (C), (F), (I), (L): measurement of dehydroepiandrosterone; transition 289.2 / 253.2; x-axes: retention time [min], y-axes: intensity [cps].

[0129] Fig. 8: Effect of storage parameters on leaching: As in Fig. 2, using as LC eluent water stored at 45°C for 120h or at 45°C for 120h, followed by 3 weeks at 35°C; (A) in A round H additional incubation for 3 weeks does not increase leaching; (b) in CB UVN SD, the signal intensified with longer incubation; data from LC-MS without sample application, x-axes: retention time [min], y-axes: total ion intensity [cps].

[0130] Fig. 9: As in Fig. 7, separation of a steroid-mix (17a-Hydroxyprogesteron, Aldosterone, Estradiol, Progesterone, Testosterone, together with 13C-labeled derivatives) with methanol stored in CB UVN SD bottles for (A) 120h at 45°C, or (B) 120h at 45°C, followed by 3 weeks at 37°C as LC eluent; transitions: 315.2 / 109.1.

[0131] Fig. 10: Detection of leachable compounds in acetonitrile as LC eluent after incubation as indicated in Example 1 (A) by MS (full scan, neg. mode) and (B) by UV spectroscopy (diode array detector, DAD); data from LC-MS without sample application; x-axes: retention time [min], y-axes: intensity [cps]; P: storage in polymer bottle, G: storage in glass bottle.

[0132] The following Examples shall merely illustrate the invention. They shall not be construed, whatsoever, to limit the scope of the invention.

[0133] Example 1 : Leachable compounds and storage container materials

[0134] Unless indicated otherwise, the indicated solvents were kept at 120 h @ 45 °C + / - 2°C, followed by 3 weeks @ 35 °C + / - 2°C and then were used as eluents in LC-MS, either applying a sample as indicated or in runs without sample. Water samples were tested for microbial contamination after incubation: results for all samples: < 1 CFU (colony forming) / 100 mL was deemed to indicate no microbial contamination.

[0135] Container materials used were Schott glass bottles (borosilicate glass), Aicello bottle "A round H", Aicello bottle "CB type A", Aicello bottle "CB UVN SD", and Aicello bottle "CB grey".

[0136] Samples were analyzed by LC-MS according to standard protocols. LC gradients were 5-95% solvent (acetonitrile or methanol) + 0.1% Formic Acid in Water in 10 min. MS; MRM transitions used are indicated in the specific Examples and / or Figure legends.

[0137] Example 1.1 Water as eluent Results are shown in Fig. 1-3; Fig. 1 shows higher background MS noise after storage of water in polymer bottles, but also an additional peak for the A round H and CB grey bottle material.

[0138] Fig. 2 shows full scan MS data in positive mode after long incubation time with signal as a function of time; this Figure reveals additional peaks originating from the bottle material supporting the MRM findings.

[0139] As shown in Fig. 3, origin of additional peaks (leachable compounds) can be traced back to the polymer of the bottle dissolving into the solution by the 2 D plot of m / z value against retention time. From the full scan, the equidistant pattern is a clear indicator of polymer origin (marked with the ellipses in Fig. 3; notably, the signals are absent when using a glass bottle.

[0140] Further, Fig. 4 shows that the additional signals after storage in plastic bottles can be observed in positive (A) and negative (B) ESI mode.

[0141] Example 1.2 Methanol as eluent

[0142] As shown in Fig. 5, incubation of polymeric (plastic) bottles with methanol causes high background peaks in the range from 50-500 M / z at retention time 4,13 min compared to glass bottles. The data of the two plastic bottles (Fig. 5 (A): CB UVN SD; and (B): A round H) shows characteristic polymer fingerprint spectra that are unique to the bottle material in combination with the solvent, storage time and assay protocol, absent after incubation in glass bottles (C). This is confirmed by the 2D-plot shown in Fig. 6.

[0143] Example 1.3: Acetonitrile (ACN) as eluent

[0144] After storage of acetonitrile, as shown in Fig. 7, additional peaks originating from the polymer bottle are also visible when compared to glass bottles for different bottle types (but not for the control and glass) and depending on the monitored MRM signal.

[0145] Example 2: Storage parameters

[0146] The impact of storage temperatures and times on compound leaching was evaluated. Bottles consisting of a variety of materials were incubated with water or acetonitrile for 120h at 45 °C, 3 weeks at 35°C, or 2 weeks at room temperature. As shown in Figs. 8 and 9, with most polymeric storage container materials, signal intensities of leachable compounds increase after prolonged incubation. With some materials, equilibrium may already be reached after the initial incubation (Fig. 8(A)). Example 3 : Detection methods

[0147] After incubation of acetonitrile in A round H, CB UVN SD, CB type A (together referred to as "polymer" in Fig. 10), or glass bottles, samples were analyzed by LC-MS; leachable compounds were detected by MS (Fig. 10(A)) or by UV detection after LC (FIG. 10(B)), showing that also UV detection is applicable to detect leachable compounds.

Claims

Claims1. A method for determining a mass spectrometry (MS) reagent stored in a storage container for a period of time and / or at least one storage parameter thereof, said method comprising(a) determining at least one leachable compound of said storage container;(b) comparing the at least one leachable compound determined in step (a) to a reference, and(c) determining said MS reagent and / or at least one storage parameter thereof based on the comparing in step (b), wherein said storage parameter is said period of time of storage or is a storage condition, and wherein said storage condition is a material the storage container is comprised of (storage container material), storage temperature, light exposure, humidity, vibration or other movement, pressure, and / or magnetic field(s).

2. The method of claim 1, wherein step (c) is determining said MS reagent.

3. The method of claim 1 or 2, wherein said storage container material is a polymer, in an embodiment an organic polymer.

4. The method of any one of claims 1 to 3, wherein said determining is quantitatively determining said at least one storage parameter.

5. The method of any one of claims 1 to 4, wherein said leachable compound is a compound extractable by the MS reagent from the storage container at an amount above the detection limit of said MS within said period of time.

6. The method of any one of claims 3 to 5, wherein said leachable compound comprises at least one monomer used to produce the polymer, or is a softener, processing aid, or other adjuvant.

7. The method of any one of claims 1 to 6, wherein said storage container is comprised of a plurality of layers and wherein said leachable compound is not comprised in an innermost layer of the storage container, in an embodiment wherein in case at least one leachable compound from a non-innermost layer of the storage container is detected, it is concluded that the structural integrity of the storage container was lost before or during storage.

8. The method of any one of claims 1 to 7, wherein said storage container is comprised of a plurality of elements and wherein said leachable compound is not comprised in at least one element of the storage container, in an embodiment wherein said leachable compound is only comprised in a lid of said storage container.

9. The method of any one of claims 1 to 8, wherein determining an MS reagent is verifying a composition of an MS reagent by determining at least one leachable compound of said storage container.

10. The method of any one of claims 1 to 9, wherein said leachable compound is determined in step (b) by MS.

11. The method of any one of claims 1 to 10, wherein said mass spectrometry is used in combination with a chromatographic separation step, in an embodiment liquid chromatography mass spectrometry (LC-MS), in a further embodiment high- performance liquid chromatography coupled mass spectrometry (HPLC-MS).

12. The method of any one of claims 1 to 11, wherein said MS reagent comprises at least one organic solvent.

13. A method of fingerprinting a storage container material, said method comprising(A) incubating said storage container material with an MS reagent under a set of environmental conditions for a period of time,(B) determining at least one leachable compound of said storage container material, in an embodiment by mass spectrometry (MS);(C) allocating data on leachable compounds obtained in step (B) to the storage container material and optionally the environmental conditions and / or duration of said period of time of step (A), thereby fingerprinting said storage container material, wherein said fingerprinting is providing data enabling identification of a storage container material and / or enabling determining whether the storage container material is according to a pre-determined specification.

14. The method of any one of claims 1 to 13, wherein said leachable compound is an organic compound.

15. A method of quality assurance of an MS analysis, comprising determining a storage condition of an MS reagent method of any one of claims 1 to 12 or 14 and the further step of rejecting a result of an MS measurement in case it is determined that said MS reagent and / or storage parameter does not fulfill a predetermined specification.

16. A data carrier comprising tangibly embedded a database comprising at least one identifier of a storage container material; allocated to data on at least one leachable compound and optionally a set of environmental conditions and / or a length of said period of time of incubation used to obtain said data on at least one leachable compound.

17. The data carrier of claim 16, further comprising at least one identifier of a reference reagent allocated to said data on at least one leachable compound and optionally said set of environmental conditions and / or said length of said period of time of incubation used to obtain said data on at least one leachable compound.

18. An MS system comprising an MS measuring unit and a control unit, wherein said MS system further comprises a data carrier according to claim 16 or 17, and / or wherein said control unit comprises tangibly embedded an executable code which, when executed on the control unit, causes the system to perform the method of any one of claims 1 to 14.

19. A kit comprising an MS reagent and a data carrier according to claim 16 or 17.

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