Device and method for analysis of reference elements in a sample by laser induced breakdown spectroscopy

Abstract

A device (1) for use in analyzing reference elements in a sample (400) by laser induced breakdown spectroscopy comprises a laser irradiation arrangement (100) configured to provide one or more focused laser beam pulses (140) at a predefined ablation position (130) in a predefined ablation plane (131) of the laser irradiation arrangement (100), at which a target surface (401) of the sample (400) is to be positioned, wherein the one or more laser beam pulses (140) are configured to ablate material from the target surface (401) when positioned at the ablation position (130), thereby creating a respective expanding plasma plume (300) including one or more reference elements. The device (1) further comprises an analysis arrangement (200) including light-collecting optics (210) configured to collect emission light (525) from the respective plasma plume (300) for each of the one or more laser beam pulses (140). An optical axis (211) of the light-collecting optics (210) along which emission light (525) is collected has a viewing angle (φ) to the main expansion axis (311) in a range between 0° and 60°, and a focal point (213) of the light-collection optics (210) is located above the ablation plane (131) at a detection distance (215) to the ablation position (130) in order to collect emission light (525) from the plasma plume (300) at a distance above the target surface (401). The emission light (525) is collected from the plasma plume (300) over a sampling time window (250) starting with a time delay (Δt) relative to the provision of the laser beam pulse (140) to the ablation position (130).

Description

New International Patent Application Raylution AGAttorney ref.: P6844PC00DESCRI PTIONTitleDEVICE AND METHOD FOR ANALYSIS OF REFERENCE ELEMENTS IN A SAMPLE BY LASER INDUCED BREAKDOWN SPECTROSCOPYTechnical Field

[0001] The present invention relates to a device for use in analyzing reference elements in a sample, in particular in a biological tissue, by laser induced breakdown spectroscopy. The invention further relates to a method of analyzing reference elements in a sample, in particular in a biological tissue, by laser induced breakdown spectroscopy, preferably by using such a device.Background Art

[0002] Laser induced breakdown spectroscopy (LIBS) has become a well-established method for a rapid and effective analysis of materials in the field of metal processing or geology, e.g. for the analysis of material mix-ups, the analysis of inclusions in steel, the analysis of slags in secondary metallurgy or the analysis of combustion processes minerals.

[0003] In LIBS, a short, highly energetic laser pulse (femtosecond to nanosecond) is provided to a target surface of a sample to ablate small amounts of material and convert it into an expanding plasma plume which breaks down into excited ionic, atomic and molecular species. As the expanding plasma cools down, it reveals characteristic emission lines of the elements contained therein, which can be analyzed with a spectrometer. From the specific emissions lines, it is possible to identify specific elements, such as specific atoms, which can be used as reference elements to characterize the sample. It is also possible to quantify the amount of a specific atom (reference element) relative to others.

[0004] While the specific emissions lines allow for a reasonable qualitative analysis, quantitative analysis of the exact element ratio within a given sample remains a challenge and can only yield a rough estimate of the element ratio. Quantitative analysis is even more difficult for “wet” biological samples due to the uncontrolled plasma generation process. Here, quantitative analysis is in particular impaired by the formation of strong oxygen, nitrogen and hydrogen emission lines resulting from plasma reactions with air and water. Also, inhomogeneities of the tissue and a strong quenching effect caused by the water content of the sample impede a quantitative analysis of biological samples. In addition, the Stark effect may lead to a strong peak broadening accompanied by a decrease of the peak intensity for higher element concentrations. All these factors adversely affect the quality of the LIBS signals and lead to a lack of linearity and poor reproducibility.

[0005] To compensate for these effects, averaging over a large number of spectra recorded for up to hundred or even thousand laser shots is often the only solution to obtain more robust and meaningful data. However, for inhomogeneous samples, averaging over a large number of spectra taken for different laser shots and thus for different sample depth blurs the information about the locally varying sample composition. Hence, to generate a depth profile across an inhomogeneous sample, single laser shot sensitivity is almost prerequisite in order to allow for quantitative analysis. The same would also be advantageous to generate a one or two-dimensional sample profile across an inhomogeneous sample in a plane parallel to the target surface.

[0006] Therefore, there is a need for a LIBS device and a method allowing for an improved, at least semi-quantitative sample analysis, in particular for an improved least semi-quantitative sample analysis of biological samples.Disclosure of the Invention

[0007] According to the invention this need is settled by a device and a method for analyzing reference elements in a sample, in particular in a biological tissue, by laser induced breakdown spectroscopy as it is defined by the features of the independent claims. Preferred embodiments are subject of the dependent claims.

[0008] In particular, the invention relates to a device for use in analyzing reference elements in a sample, in particular in a biological tissue, by laser induced breakdownspectroscopy. According to the invention, the device comprises a laser irradiation arrangement configured to provide one or more focused laser beam pulses at a predefined ablation position in a predefined ablation plane of the laser irradiation arrangement, at which a target surface of the sample is to be positioned. In particular, the laser irradiation arrangement may be configured to provide the one or more focused laser beam pulses onto a target surface of the sample in a predefined ablation plane of the laser irradiation arrangement such that an ablation focal point of the focused laser beam pulses is at a predefined ablation position in the ablation plane, at which the target surface of the sample is to be positioned.

[0009] The one or more laser beam pulses are configured to ablate material from the target surface when positioned at the ablation position. As a result, a respective plasma plume including one or more reference elements is created for each of the one or more laser beam pulses. The plasma plume including the one or more reference elements expands away from the target surface along main expansion axis which substantially corresponds to a normal to the target surface at the ablation position or, with reference to the device, to a normal to the ablation plane at the ablation position. In addition, the device comprises an analysis arrangement including light-collecting optics configured to collect emission light from the respective plasma plume for each of the one or more laser beam pulses, in particular to collect emission light emitted by the one or more reference elements within the respective plasma plume.

[0010] The terms “predefined ablation position” and “predefined ablation plane” as used herein refer to a predefined reference point and a predefined reference plane of the laser irradiation arrangement defining a position to be assumed by the sample, such that a target site of the target surface to be analyzed is to be placed at the ablation position and the target surface around the target site is to be arranged substantially in the ablation plane. The ablation plane may be defined by a structural element of the laser irradiation arrangement, such as a contact surface of the laser irradiation arrangement. Likewise, the predefined ablation position may be defined by a focal point of the laser irradiation arrangement and / or by a structural feature of the laser irradiation arrangement, such as an opening or an aperture in a contact element of the laser irradiation arrangement.

[0011] As used herein, the term “reference element” refers to any kind of element, in particular to atoms, molecules, ions, electrolytes and / or functional groups in a chemical compound, that may be indicative for a specific property or state of the sample. Forexample, with respect to biological tissue samples, the reference elements may comprise one or more specific members of the electrolyte and elemental system in a cell (Mg, Fe, K, Na, Ca, CN, CaO, CaOH, CaF, CaCI, etc.), for which it is known that it is changing when the healthy cell converts to a tumor cell. For example, it is known that in mandible bone tumor cells the calcium content is reduced, while potassium is increased. Accordingly, potassium and calcium may be used as reference elements for the analysis of a biological tissue sample, in particular as an indicator for the presence of a cancerous tissue.

[0012] Furthermore, the term “emission light” as used herein refers to any kind of electromagnetic radiation emitted during expansion of the plasma plume. In particular, the term “light” is not confined to the visual / optical part of the electromagnetic radiation spectrum but may in particular include electromagnetic radiation in the UV, optical and infra-red spectrum and also electromagnetic radiation beyond the UV and infra-red spectrum.

[0013] According to the invention, it has been found that if reference elements, e.g. atoms and molecules, within the inner center zone of the plasma plume have sufficient time to reach an excited equilibrium state due to the homogenization of the plasma plume, such reference elements will emit clear emission lines with an appropriate light intensity that is correlated with their actual amount present in the sample. This requires that the plasma expands for a certain amount of time, thereby moving away from the ablation position on the target surface. Moreover, it has been found that the signal intensity for LIBS applications can be further improved if as much emission light as possible can be collected from elements in the excited equilibrium state, in particular for a single laser beam pulse. In doing so, it has been found that even single-shot sensitivity can be achieved, enabling at least for a semi-quantitative analysis of biological samples.

[0014] As used herein, the term “semi-quantitative analysis” refers to an analysis of the sample that at least allows to compare the amounts of one or more specific reference elements within a single individual sample, in particular to compare the amounts of one or more specific reference elements at specific position relative to the respective amounts at other positions within the same sample. Accordingly, the term “semi-quantitative analysis” may refer to an analysis of the sample that yields a spatial profile of the relative amounts of one or more specific reference elements within a specific sample, i.e. a change of the amounts of one or more specific reference elements across a specificsample. Such a spatial profile may in particular be a depth profile and / or a one or two- dimensional profile in a plane parallel to the target surface for the (relative) amounts of one or more specific reference elements within an individual sample, e.g. a biological tissue of a specific individuum. As such, “semi-quantitative analysis” may in particular allow for a specific individual to generate a one or two-dimensional spatial profile of the relative amounts of K (potassium) across a biological tissue of that individuum. For example, such a spatial profile of the (relative) K amount across a biological sample of a specific individuum may enable to identify the spatial expansion or margins of potentially cancerous tissue in the sample. The spatial expansion or margins may be given by the parts of the sample where the (relative) K content for the specific sample of the individuum exceeds a certain threshold value.

[0015] As a consequence of the above findings, the present invention suggests to configure and arrange the light-collecting optics such that an optical axis of the lightcollecting optics along which emission light is collected has a viewing angle to the main expansion axis in a range between 0° and 60° and such that a focal point of the lightcollection optics is located above the ablation plane at a detection distance to the ablation position.

[0016] Having the focal point of the light-collection optics located above the ablation plane at a detection distance to the ablation position ensures that the light-collecting optics collects emission light from the plasma plume emitted at a distance above the target surface when passing the focal point or a light-collecting area around the focal point. Thus, the detection distance between the focal point and the ablation position ensures that the light-collecting optics mainly collects emission light from atoms and molecules within a region of the plasma plume that has expanded long enough to homogenize and thus to allow the reference elements of interest within that region, such as specific atoms and molecules, to reach the preferred excited equilibrium state.

[0017] As used herein, the term “above the ablation plane” refers to that side of the ablation plane into which the plasma plume expands, and from which the one or more laser beam pulses are provided to the ablation position / target surface.

[0018] Having the optical axis of the light-collecting optics to form a viewing angle to the main expansion axis in a range between 0° and 60° advantageously allows to collect more light from the respective reference elements in the excited equilibrium state thanwith a larger viewing angle, e.g. a viewing angle of 90°. This is due to the fact that for smaller viewing angles the plasma plume expands more into the colleting direction of the light-collecting optics, thus allowing the light emitting reference elements within the expanding plasma plume to stay longer within the viewing field of the light-collecting optics, or vice versa enabling the light-collecting optics to track the light emitting reference elements within the expanding plasma plume over a longer sampling time. This proves particularly beneficial since light emission occurs stochastically which requires the sampling time to be as long as possible in order to collect more emission light. A longer sampling time not only permits to collect more emission light but also more reproducibly, as fluctuations in light emission (due to its stochastic nature) average out over a longer sampling time. As a result, a smaller viewing angle helps to increase the sensitivity of the device and thus to improve (semi-)quantitative sample analysis. A smaller viewing angle also makes the sensitivity of the device less susceptible to the exact positioning of the focal point of the light-collecting optics relative to the ablation position.

[0019] In general, the smaller the viewing angle, the larger the amount and the better the reproducibility of the collected emission light, and thus the better the overall sensitivity of the device and the robustness of the optical alignment. Vice versa, the greater the viewing angle, the smaller the detectable signal, as the expanding plasma plume then moves out of the viewing field of the light-collecting optics more quickly. Therefore, it may be preferred that the viewing angle is in a range between 0° and 45°, in particular in a range between 0° and 30° or between 2° and 30° or between 0° and 20° or between 2° and 20° or between 0° and 15° or between 2° and 15°, more particularly in a range between 0° and 10° or between 5° and 15° or between 10° and 20°.

[0020] Preferably, the light-collecting optics may comprise at least one collector lens that may face the ablation position. The at least one collector lens may have a focal length defining the focal point of the light-collecting optics along the optical axis of the lightcollecting optics. The light-collecting optics may comprise additional optical elements, in particular at least one of: one or more additional lenses, one or more optical filters, one or more beam splitters, one or more apertures, one or more optical mirrors, and an optical fiber to direct emission light collected for each of the one or more focused laser beam pulses to an analysis device, such as a spectrometer.

[0021] According to the invention, the analysis arrangement is configured to collect emission light from the plasma plume over a sampling time window starting with a timedelay relative to the provision of the respective laser beam pulses to the ablation position. The time delay between the respective provision of the laser beam pulse and the respective start of emission light collection advantageously serves to fade out emission light that does not result from the reference elements of interest in the desired equilibrium state, in particular to fade out emission effects resulting from processes that typically occur prior to the desired emission of the reference elements. Such emission effects may comprise continuum emission, in particular electron cyclotron emission due to free electrons in the plasma, emission due to bremsstrahlung radiation and recombination reactions, or emission of ionic emission lines. Most important, the time delay serves to start collecting emission light only after the fast-moving shock wave induced by the expanding plasma plume has passed the light-collecting area surrounding the focal point. Thus, the time delay helps to avoid strong emission lines for nitrogen, oxygen and hydrogen in the LIBS spectra. These emission lines result from reactions of the hot plasma plume front with ambient air right after application of the laser beam pulse and can therefore typically be observed during an early phase of the plasma expansion (0 - 0.8 ps after application of the laser beam pulse).

[0022] In general, the time delay may be chosen such that the fast-moving shock wave induced by the expanding plasma plume has passed the light-collecting area surrounding the focal point of the light-collecting optics and such that the expanding plasma plume had sufficient time to properly homogenize. To this extent, the appropriate time delay to be chosen may depend on various factors, in particular on the energetic parameters of the plasma plume, such as the plasma temperature, on the properties of the sample, as well as on the position of the light-collecting area surrounding the focal point of the lightcollecting optics relative to the ablation position. Accordingly, the time delay may be chosen, in particular adjustable, based on at least one of the following parameters: the detection distance, an energy of the laser beam pulse, a spot diameter of the laser beam pulse at the ablation position, a material composition of the sample at the target surface, a density of the sample at the target surface, a water content of the sample at the target surface, and a maximum temperature within the plasma plume.

[0023] In principle, the same applies to the detection distance, which also strongly depends on the energetic parameters of the plasma plume and the properties of the sample. Of course, the detection distance also depends on the time delay, and vice versa. That is, the time delay also depends on the detection distance. Hence, the detectiondistance and the time delay preferably are matched to each other such that the analysis arrangement collects emission light from the plasma plume at the right time and in the right place.

[0024] Accordingly, the detection distance may be chosen, in particular adjustable, based on at least one of the following parameters: the time delay, an energy of the laser beam pulse, a spot diameter of the laser beam pulse at the ablation position, a material composition of the sample at the target surface, a density of the sample at the target surface, a water content of the sample at the target surface, and a maximum temperature within the plasma plume.

[0025] In order to enable an adaption of the time delay and / or the detection distance to different conditions, the analysis arrangement preferably is configured such that the time delay and / or the detection distance is adjustable. In principle, it is also possible that the time delay and / or the detection distance is fixed.

[0026] As stated above, the time delay should preferably be chosen long enough to wait until the early phase of the plasma expansion (0 - 0.8 ps after application of the laser beam pulse) has passed and the plasma has been homogenized. Accordingly, the time delay may preferably be at least 0.5 ps and at least 0.8 ps, in particular at least 1.0 ps, more particularly at least 1 .3 ps, preferably at least 1 .5 ps or 2.0 ps or at least 2.0 ps or at least 2.5 ps or at least 3.0 ps or at least 3.5 ps or at least 4.0 ps or at least 4.5 ps or at least 5.0 ps or at least 5.5 ps or at least 6.0 ps or at least 6.5 ps or at least 7.0 ps or at least 7.5 ps or at least 8.0 ps or at least 8.5 ps or at least 9.0 ps or at least 9.5 ps or at least 10.0 ps or at least 12.5 ps or at least 15.0 ps or at least 17.5 ps. Vice versa, the time delay should preferably be chosen short enough so that the plasma plume is not diluted in ambient air or in any other ambient gas. Accordingly, the time delay may preferably be in a range between 0.5 ps and 30.0 ps or between 0.5 ps and 20.0 ps or between 0.8 ps and 30.0 ps or between 0.8 ps and 20.0 ps, in particular between 1 .0 ps and 25.0 ps, more particularly between 1.3 ps and 20.0 ps or between 1.5 ps and 20.0 ps or between 2.0 ps and 20.0 ps or between 2.5 ps and 20.0 ps or between 1 .3 ps and 15.0 ps or between 1.5 ps and 15.0 ps or between 3.0 ps and 15.0 ps or between 5.0 ps and 15.0 ps or between 5.0 ps and 12.0 ps or between 5.0 ps and 12.0 ps or between 5.0 ps and 10.0 ps or between 6.0 ps and 10.0 ps or between 4.0 ps and 9.0 ps or between 5.0 ps and 10.0 ps.

[0027] Start of the time delay may be triggered by detecting the application of the laser beam pulse. For this, the analysis arrangement may comprise a light detector, such as a photodiode, for detecting a portion of the light of a laser beam pulse applied to the sample. In response to detecting light, i.e. the onset of a laser beam pulse, the detector may generate a detection single that may be used, e.g. by a controller of the device, as trigger for start of the time delay. For detecting a portion of the light of a laser beam pulse, the device may further comprise a beam splitter, such as beam splitter cube a glass plate, in the optical path of the laser irradiation arrangement that is configured to split off / reflect off a small portion of the light of a laser beam pulse applied to the sample and to direct it towards the light detector.

[0028] Likewise, the detection distance should preferably be chosen large enough to allow the expanding plasma to homogenize. Accordingly, the detection distance may preferably be at least 2 mm, in particular at least 5 mm, more particularly at least 10 mm or at least 15 mm. Vice versa, the detection distance should not be too large to avoid dilution effects. Accordingly, the detection distance may preferably be in a range between 2 mm and 30mm, in particular between 5 mm and 20 mm or between 5 mm and 15 mm, more particularly in a range between 7.5 mm and 15 mm or between 5 mm and 12 mm or between 6 mm and 12 mm or between 7 mm and 12 mm or between 6 mm and 11 mm or between 7 mm and 11 mm, for example 8 mm or 8.5 mm or 9 mm or 9.5 mm or 10 mm.

[0029] In order to achieve sufficient signal intensity for LIBS applications, a length of the sampling time window (integration time) should be chosen long enough to collect a sufficient amount of emission light. Accordingly, a length of the sampling time window may preferably be at least 0.5 ps, in particular at least 0.75 ps, more particularly at least 1 .0 ps or at least 1 .5 ps or at least 2.0 ps. However, the shorter the sampling time window the less ambient light is collected and the less the LIBS data are affected by ambient light. For this reason, it may prove advantageous that a length of the sampling time window is in a range between 0.5 ps and 10.0 ps, in particular between 0.75 ps and 5.0 ps, more particularly between 1 .0 ps and 3.0 ps.

[0030] Inherently, the light-collecting optics defines a detection volume above the ablation plane traversed by the expanding plasma plume from which it is able to effectively collect emission light. That is, the light-collecting optics is configured to collect emission light from a detection volume above the ablation plane traversed by the expanding plasma plume. Preferably, this detection volume should be as large aspossible in order to collect as much emission light as possible for a single laser beam pulse. In particular, the detection volume should have a largest possible extension along the main expansion axis of the plasma plume so that the light-collecting optics can track the light emitting atoms within the expanding plasma plume over a longest possible expansion distance. The detection volume may have an extension along the main expansion axis, or alternatively, along the optical axis of the light-collecting optics, of at least 2 mm, in particular at least 5 mm, more particularly at least 10 mm or at least 15 mm or at least 20 mm or at least 25 mm. Likewise, the detection volume may have an extension along the main expansion axis, or alternatively, along the optical axis of the light-collecting optics, in a range between 2 mm and 40 mm, in particular between 5 mm and 30 mm, more particularly in a range between 7.5 mm and 25 mm or in range between 10 mm and 20 mm, for example 10 mm or 12 mm or 15 mm.

[0031] Preferably, the light-collecting optics, in particular a collector lens of the lightcollecting optics facing the ablation position, may have a small F-number, which is defined by the following equation: F-number = 1 / (2 n sin 9), wherein n is the refractive index of space in which the source (light emitting reference elements) is located, and 6 is the half angle of the cone of radiation. For small angles (6 <15°), the F-number a collector lens in the paraxial approximation is given by F-number = f / D, wherein f is the focal length of the collector lens, and D is the clear aperture D of the collector lens. The smaller the F- number the greater the radiant flux collected by the collector lens. Accordingly, usage of a shorter focal length in principle increases the radiant flux collected by the collector lens. However, in order to track the light emitting atoms within the expanding plasma plume over a longest possible expansion distance, a longer local distance is desirable. Preferably, the light-collecting optics, in particular a collector lens of the light-collecting optics facing the ablation position, may have a focal length (defining the focal point of the light-collecting optics along the optical axis of the light-collecting optics) in a range between 50 mm and 500 mm, in particular between 100 mm and 400 mm or between 100 mm and 300 mm or between 150 mm and 300 mm, more particularly between 180 mm and 250 mm, for example 200 mm. A focal length within these ranges constitutes a good comprises between a sufficiently high collected radiant flux and a sufficiently long detection volume along the optical axis of the light-collecting optics. As stated above, the latter becomes effectively even longer, the smaller the viewing angle between the optical axis of the light-collecting optics and the main expansion axis of the plasma plume.

[0032] In order to avoid signal fluctuations due to sample inhomogeneities on a micrometer scale, it has proven to be beneficial that the spot size of the focused laser beam pulse in the ablation plane at the ablation position is sufficiently large such that sample inhomogeneities within an area of the laser beam spot will average out for each pulse. In particular when analyzing biological tissues, a larger laser spot may facilitate to ablate and thus to analyze a cell ensemble. Thus, even a single laser beam pulse may provide enough sensitivity, while signal fluctuations can be reduced at the same time. Additionally, a larger spot size of the ablating laser beam pulse advantageously results in a larger plasma plume. In a larger plasma plume, the ratio between the outer surface area of the expanding plasma shock wave and the plasma volume is smaller than for smaller plasma plumes. Advantageously, a smaller surface-to-volume ratio of the plasma plume helps to keep the plasma temperature at a high level over a longer period of time, so that the plasma homogenizes more efficiently. This in turn results in a better sensitivity and thus in an improved (semi-)quantitative sample analysis. For a given plasma temperature, a larger plasma plume also allows the inner plasma processes to stabilize over a longer period of time and thus to allow the reference elements within the plasma to better reach the preferred excited equilibrium state.

[0033] Accordingly, the laser irradiation arrangement preferably is configured to provide the one or more focused laser beam pulses such that each one of the laser beam pulses has a spot diameter at the ablation position in the ablation plane of at least 30 pm, in particular at least 50 pm, more particularly at least 100 pm, preferably at least 125 pm; or wherein the laser irradiation arrangement is configured to provide the one or more focused laser beam pulses to have a spot diameter at the ablation position in the ablation plane in a range between 30 pm and 400 pm, in particular between 50 pm and 200 pm or between 80 pm and 200 pm, more particularly between 100 pm and 200 pm, for example 125 pm or 150 pm.

[0034] Preferably, the laser irradiation arrangement is configured to focus the laser beam pulses to have a focal point at the ablation position in the ablation plane. Advantageously, this facilitates to provide a sufficiently high photon density at the ablation position. In this configuration, the ablation position may be defined by the position of the focal point of the laser irradiation arrangement, and the above discussed spot diameters correspond to the spot diameter of the focused laser beam pulse at their focal point. Forfocusing the laser beam pulses, the laser irradiation arrangement may comprise focusing means, such as at least one focusing lens facing the ablation position.

[0035] In order to have more flexibility in positioning the focal point of the laser irradiation arrangement along the optical axis relative to the ablation position, it may be advantageous that the laser irradiation arrangement comprises focusing means, such as the at least one focusing lens, with a large focal length, resulting in a longer Rayleigh length of the laser beam pulses. In this way, the photon density in the ablation position is more or less the same independent from the exact position of the focal point of the laser irradiation arrangement relative to the ablation position. A longer Rayleigh length is also desirable if the laser beam pulses shall be able to drill a tunnel into the sample without any widening or shrinking of the tunnel diameter. Preferably, the laser irradiation arrangement, in particular focusing means of the laser irradiation arrangement, more particularly the at least one focusing lens facing the ablation position, may have a focal length in a range between 50 mm and 500 mm, in particular between 100 mm and 400 mm or between 100 mm and 300 mm or between 150 mm and 300 mm, more particularly between 180 mm and 250 mm, for example 200 mm.

[0036] In principle, the laser beam pulses may be provided onto the target surface in a direction normal to the target surface and the ablation plane, respectively, or with an oblique angle of incidence. Accordingly, the optical axis of the laser irradiation arrangement along which the laser beam pulse is provided onto the target surface may form an irradiation angle with a normal to the ablation plane at the ablation position in a range between 0° and 60°, in particular between 0° and 45°, more particularly in a range between 2° and 30° or between 5° and 20° or between 5° and 15° or between 10° and 20° or between 0° and 15° or between 0° and 10° or between 0° and 5° or between 0° and 2°. Preferably, the irradiation angle is as small as possible such that the laser beam pulses hit the target surface perpendicular to it, i.e. with an irradiation angle of 0°. In this configuration, the photon intensity is highest and most homogeneous at the ablation position. With respect to the irradiation angle, it is to be noted that the main expansion axis / direction of the plasma is substantially independent from the irradiation angle. That is, the plasma plume will more or less always expand substantially along a normal to the target surface and thus along a normal to the ablation plane at the ablation position, irrespective of whether the laser beam pulses hit the target surface perpendicular to the target surface or at an oblique angle of incidence.

[0037] Moreover, it may be preferred that the optical axis of the light-collecting optics along which emission light is collected and an optical axis of the laser irradiation arrangement along which the laser beam pulse is provided onto the target surface form an acute angle to each other, in particular an acute angle in a range between 1 ° and 45°, more particularly between 1 ° and 30° or between 1 ° and 20° or between 1 ° and 15° or between 5° and 20° or between 5° and 15° or between 1 ° and 10° or between 1 ° and 5° or between 1 ° and 2°. Likewise, the optical axis of the light-collecting optics along which emission light is collected and an optical axis of the laser irradiation arrangement along which the laser beam pulse is provided onto the target surface may be parallel to each other. In the latter case, the optical axis of the light-collecting optics, along which emission light is collected, and an optical axis of the laser irradiation arrangement, along which the laser beam pulse is provided onto the target surface, may coincide over a common optical path of a certain length and somewhere be separated, e.g. by means of a beam splitter, such as beam splitter cube a glass plate or a dichromatic mirror.

[0038] In addition, the laser irradiation arrangement may comprise at least one pulsed laser source, in particular at least one pulsed laser source with Q-switching, more particularly at least one pulsed laser source with passive Q-switching. Advantageously, passive Q-switching provides a constant pulse energy, which enhances the reproducibility of the sample analysis. For example, the laser source may comprise a Nd: YAG laser, in particular a high-power Q-switched Nd:YAG laser. Advantageously, the Nd:YAG laser may provide a 1064 nm or a 532 nm laser beam pulses or laser beam pulses with wavelengths between 200 nm and 1200 nm.

[0039] The one or more laser beam pulses may each have a pulse duration of a few nanoseconds, e.g. in a range between 2 ns to 10 ns, in particular between 2 ns and 8 ns or between 2 ns and 5 ns or between 5 ns and 8 ns. The energy per pulse should be high enough to achieve a sufficiently high photon density at the ablation position that is required to generate a hot plasma. Accordingly, the one or more laser pulses may each have an energy per pulse in a range between 50 mJ and 150 mJ, in particular between 70 mJ and 100 mJ. Preferably, the pulsed laser source may be configured to adjust the pulse duration and / or the energy per pulse of the or more laser beam pulses.

[0040] As used herein, the term “one or more laser beam pulses" in general refers to such laser beam pulses that are used to ablate material from the target surface so that aplasma plume to be analyzed is generated. Accordingly, these laser beam pulses may also be denoted as analysis laser beam pulses.

[0041] In addition to that, the laser irradiation arrangement may be configured to generate one or more preparation laser beam pulses in advance to an analysis laser beam pulse, in particular one or more preparation laser beam pulses in advance to each analysis laser beam pulse. The one or more preparation laser beam pulses may comprise / may be one comparably long preparation laser beam pulse or a series of shorter preparation laser beam pulses. For example, the one or more preparation laser beam pulses may be a predefined number of pulses identical to the analysis laser beam pulses and preceding an analysis laser beam pulse. The one or more preparation laser beam pulses may be used to clean and / or dry the target surface at the ablation position, in particular, where the sample is a biological tissue, to remove liquid from the target surface where it is aimed to be hit by the analysis laser beam pulses. This proves particularly to reduce undesired quenching effects due to a larger water content of biological samples. Further, the preparation laser beam pulses may provide a certain ablation depth such that the analysis pulse hits the sample at a lower layer. A temporal width of the preparation laser beam pulses preferably is bigger than a temporal width of a subsequent analysis laser beam pulse. The laser irradiation arrangement may preferably be configured to provide an analysis laser beam pulse less than 1 millisecond, less than 1 microsecond, less than 100 nanoseconds or less than 50 nanoseconds after the one or more preceding preparation laser beam pulses. Such a time gap allows for preventing liquid, e.g. blood and / or water, from flowing to the ablation position where the analysis laser beam pulse is aimed to hit the target surface. Moreover, such time gaps allow to achieve that the preparation effect is still existing when the analysis laser beam pulse is provided. In particular, at least one pulsed laser source may be embodied for fast switching or a Q- switch crystal for slow switching between the preparation laser beam pulses and the analysis laser beam pulses.

[0042] The beam profile of the laser beam pulses may be an important factor to create a given photon density and / or to make the usage of long focusing lenses possible. In particular, the laser beam pulses may preferably have a homogeneous beam profile to provide equal distributions of photons onto the target surface at the ablation position.

[0043] The analysis arrangement may be configured to analyze emission lines in the emission light collected for each of the one or more focused laser beam pulses. Inparticular, the analysis arrangement may comprise a spectrometer for generating a respective emission light spectrum of the emission light collected for each of the one or more focused laser beam pulses. For this, the light-collecting optics may be configured to transfer the emission light collected for each of the one or more focused laser beam pulses to the spectrometer. The light transfer can be accomplished at least partially in free space, in particular by means for directing / deflecting the collected emission light, such as deflecting mirrors. Alternatively or additionally, the light transfer can be accomplished at least partially fiber-based, in particular by means of an optical fiber.

[0044] The spectrometer may comprise a spectral separator, such as a prism, a movable slit or a diffraction grating, to separate the collected emission light into its spectral components. Furthermore, the spectrometer may comprise a detector or a detector array to detect an intensity of the separated spectral components. The spectrometer may be a low-resolution spectrometer, in particular a low-resolution spectrometer having a resolving power of at most 6’000 and / or having a resolution of more than 0.2 nm per detector pixel, in particular more than 0.4 nm per detector pixel or more than 0.45 nm per detector pixel. The latter may ensure that a single plasma (laser shot) may provide enough emission light to a respective detector pixel in order to allow for single-shot sensitivity. Likewise, the spectrometer may have a resolution of less than 5 nm, in particular of less than 3 nm or less than 2 nm, for example about 1.8 nm. Furthermore, a detector or detector array of the spectrometer may be configured in a way to be sensitive enough that a single plasma (laser shot) provides enough emission light, also of low abundant elements, to obtain a full and intensive LIBS spectrum. The spectrometer may have a wavelength sensitivity range between 100 nm and 1200 nm, in particular between 130 nm and 870 nm or between 190 nm and 1100 nm. Preferably, the spectrometer is a gated spectrometer. Moreover, the spectrometer may have entrance slit with a slit width in the range between 10 pm and 30 pm, for example 12.5 pm. The width of the slit in a spectrometer impacts its resolution (and, conversely, its sensitivity) because it affects how light spreads outward. As light passes through the entrance slit, it spreads outward, and the extent of this diffraction depends on the slit width. The slit width also controls the total amount of light that enters the spectrometer which will also affect the spectral resolution.

[0045] In general, the analysis arrangement may include a single light-collecting optics and further a single spectrometer associated with the single light-collecting optics whichtransfers the collected emission light to the single spectrometer. Alternatively, the analysis arrangement may include a plurality of light-collecting optics and further a plurality of spectrometers, each associated with one of the light-collecting optics. Each one of the light-collecting optics and associated spectrometers may be configured to collect and analyze emission light of a specific wavelength range. Thus, the analysis arrangement may be selectively sensitive to different wavelength ranges and thus to different reference elements having spectra lines in different wavelength ranges.

[0046] Furthermore, the analysis arrangement or a controller of the device may be configured to evaluate spectral data provided by the spectrometer, wherein evaluation of the spectral data comprises at least one of: (i) discarding spectra having a maximum signal below a predefined value; (ii) discarding spectra saturating the spectrometer module; (iii) discarding spectra associated to air by analyzing emission peak intensities of N, of O, and / or of H, which may stem from photon density breakthrough in front of the sample surface; (iv) discarding spectra generated outside a predefined plasma electron temperature and / or plasma electron density; (v) filtering inappropriate spectra; and (vi) summing up or integrating peak areas of the emission lines. Each of the listed evaluation steps may improve the results of the sample analysis of. For example, by summing up or integrating the peak areas, sensitivity of the device can be increased or improved. Most preferably, plural or particularly all of the listed steps are combined in order to achieve the best results. In particular, the analysis arrangement may be configured to carry out the steps of analyzing emission light spectra, in particular the steps of pre-processing and applying statistical analysis, as defined further below with respect to the method of the present invention. For this, the analysis arrangement may comprise computational means, to evaluate / analyze the spectral data provided by the spectrometer.

[0047] The device may further comprise a sample stage, in particular a one- or multi- dimensionally movable sample stage, that is configured to support a sample at the ablation position in the ablation plane and preferably to position and to move the sample within one or multiple dimensions, in particular relative to the laser irradiation arrangement and the analysis arrangement. For example, the sample stage may be a xyz sample stage configured to position and move the sample in an x-direction, in a y-direction perpendicular to the x-direction and in a z-direction perpendicular to the x-direction and the y-direction. A movable sample stage may also prove advantageous to analyze a spatial distribution of one or more reference elements within a sample volume. This is ofparticular interest to analysis a heterogeneous sample, such as a heterogeneous which shows a heterogeneity in one or more dimensions.

[0048] In particular in combination with the achieved single-shot sensitivity, the sample stage may thus enable to determine a sample profile in one or more dimensions, e.g. a one or two-dimensional sample profile in a plane parallel to the target surface / sample surface and / or a depth profile in a direction perpendicular to the target surface / sample surface.

[0049] For the latter, the sample may, for example, be irradiated with several subsequent laser beam pulses, wherein the new target surface created by each pulse is repositioned after each laser beam pulse into the ablation position in the ablation plane in order to compensate for the ablation-related material removal.

[0050] For a one or two-dimensional sample profile in a plane parallel to the target surface / sample surface, the sample may, for example, be moved parallel to the ablation plane into several position, wherein for each position at least one, in particular only one laser beam pulse is provided onto the respective portion of the sample surface (that is currently in the ablation position) in order to generate a plasma plume that can be analyzed to determine the local sample composition or a specific sample property at this position.

[0051] Alternatively or in addition, the device may further comprise a one- or multi- dimensionally movable device stage that is configured to support the laser irradiation arrangement and the analysis arrangement as well as to position and to move the laser irradiation arrangement and the analysis arrangement within one or multiple dimensions, in particular relative to the sample. Preferably, the device stage may comprise a one- or multi-dimensionally movable robot or robot arm. For example, the device stage may be a xyz device stage, in particular a xyz robot, a xyz robot arm or a three-axes robot or three- axes robot arm, configured to position and move the sample in an x-direction, in a y- direction perpendicular to the x-direction and in a z-direction perpendicular to the x- direction and the y-direction. As described above with respect to the sample stage, the device may also allow in a similar way to determine a sample profile in one or more dimensions, e.g. a one or two-dimensional sample profile in a plane parallel to the target surface / sample surface or a depth profile in a direction perpendicular to the target surface / sample surface.

[0052] The sample stage and / or the device stage may also be used to determine and thus to assign a respective sampling position in 3D, i.e. the 3D coordinates (xyz coordinates) for / to each laser beam pulse, i.e. for / to each spectrum taken at that specific sampling position. This may be important, for example, for the recognition of tumor cells distributions within a given tissue. That is, knowing the 3D coordinates (xyz coordinates) of each LIBS spectrum, advantageously supports the recognition confidence.

[0053] As mentioned above, the device may further comprise a controller to control and coordinate operation of the several components of the device, in particular of the laser irradiation arrangement, the analysis arrangement, the sample stage (of present) and the device stage (if present).

[0054] In another aspect, the present invention relates to a method of analyzing reference elements in a sample, in particular in a biological tissue, by laser induced breakdown spectroscopy, preferably by using a device according to the present invention and as described herein. The method comprises: providing one or more focused laser beam pulses at a predefined ablation position on a target surface of the sample, wherein the one or more focused laser beam pulses are configured to ablate material from the target surface, thereby creating a respective plasma plume including one or more reference elements, the plasma plume expanding away from the target surface along a main expansion axis substantially corresponding to a normal to the target surface at the ablation position; collecting emission light from the plasma plume for each of the one or more focused laser beam pulses for spectral analysis; wherein collecting emission light for each of the one or more focused laser beam pulses comprises collecting emission light from the plasma plume at a viewing angle relative to the main expansion axis in a range between 0° and 60° at a distance, in particular a detection distance offset from the target surface and over a sampling time window starting with a time delay relative to the provision of the respective focused laser beam pulse onto the target surface.

[0055] In general, the device and the method according to the present invention may be used for analyzing reference elements in a sample of any kind. For example, the sample may be a metallic sample, a salt sample, a ceramic sample or a plastic sample. Thus, the device and the method according to the present invention may, for example, be used forthe analysis of elemental concentrations in alloys, ores, brines and powders. Likewise, the device and the method according to the present invention may be used for the analysis of material mix-ups, the analysis of inclusions in steel, the analysis of slags in metallurgy or the analysis of combustion processes minerals.

[0056] In particular, the sample may be a biological tissue, such as a potentially cancerous tissue. In this regard, the device and the method according to the present invention may be used for analyzing specific reference elements in a biological tissue which may be indicative for a specific state of the biological tissue, in particular for the presence of a cancerous tissue. This may be possible due to the fact that the electrolyte and elemental system (Mg, Fe, K, Na, Ca, CN, CaO, CaOH, CaF, CaCI, etc.) of a healthy cell is changing when the healthy cell converts to a tumor cell. This change has been measured by indirect methods, like Raman and fluorescent microscopy systems. Many tumor cells show a strong increase of potassium. For example, in mandible bone tumor cells the calcium content is reduced, while potassium is increased. Also, the amount and presence of other reference elements, such as for example CN, may change during conversion form healthy to the tumor state.

[0057] As regards biological tissue samples, the sample may in particular be an extracted biological tissue such that the method is an ex-vivo method. Accordingly, the method may be no method for treatment of the human or animal body by surgery or therapy, nor a diagnostic method practiced on the human or animal body.

[0058] The step of collecting emission light for each of the one or more focused laser beam pulses may in particular comprise using light-collecting optics, wherein the emission light is collected along an optical axis of the light-collecting optics orientated at the viewing angle relative to the main expansion axis, and wherein a focal point of the light-collection optics is located above the target surface at the detection distance to the ablation position.

[0059] As stated above with respect to the device according to the present invention, the time delay may be chosen, in particular may be adjustable, based on at least one of the following parameters: the detection distance, an energy of the laser beam pulse, a spot diameter of the laser beam pulse at the ablation position, a material composition of the sample at the target surface, a density of the sample at the target surface, a water content of the sample at the target surface, and a maximum temperature within theplasma plume. Preferably, the time delay is at least 0.8 ps, in particular at least 1.0 ps, more particularly at least 1 .3 ps, preferably at least 1 .5 ps or 2.0 ps or at least 2.5 ps or at least 3.0 ps or at least 4.0 ps or at least 5.0 ps or at least 7.5 ps or at least 10.0 ps. Likewise, the time delay may preferably be in a range between 0.8 ps and 30.0 ps, in particular between 1 .0 ps and 25.0 ps, more particularly between 1 .3 ps and 20.0 ps or betweenl .5 ps and 20.0 ps or between 2.0 ps and 20.0 ps or between 2.5 ps and 20.0 ps or between 1.3 ps and 15.0 ps or between 1.5 ps and 15.0 ps or between 3.0 ps and 15.0 ps.

[0060] Analogous to the device of the present invention, the detection distance for the method may be chosen, in particular adjustable, based on at least one of the following parameters: the time delay, an energy of the laser beam pulse, a spot diameter of the laser beam pulse at the ablation position, a material composition of the sample at the target surface, a density of the sample at the target surface, a water content of the sample at the target surface, and a maximum temperature within the plasma plume. Preferably, the detection distance may be at least 2 mm, in particular at least 5 mm, more particularly at least 10 mm or at least 15 mm; or wherein the detection distance is in a range between 2 mm and 30mm, in particular between 5 mm and 20 mm, more particularly in a range between 7.5 mm and 15 mm, for example 10 mm.

[0061] Furthermore, a length of the sampling time window (integration time) may be in a range between 0.5 ps and 10.0 ps, in particular between 0.75 ps and 5.0 ps, more particularly between 1 .0 ps and 3.0 ps; or wherein a length of the sampling time window is at least 0.5 ps, in particular at least 0.75 ps, more particularly at least 1 .0 ps or at least 1.5 ps or at least 2.0 ps. A length of the sampling time window within these ranges advantageously allows to collect a sufficient amount of emission light.

[0062] As also already outlined above with respect to the device of the present invention, a smaller viewing angle increases the amount of collected emission light. Therefore, it may be preferred that the viewing angle is in a between 0° and 45°, in particular in a range between 0° and 30° or between 2° and 30° or between 0° and 20° or between 2° and 20° or between 0° and 15° between 2° and 15°, more particularly in a range between 0° and 10° or between 5° and 15° or between 10° and 20°.

[0063] Preferably, each one of the laser beam pulses has a spot diameter at the ablation position on the target surface of at least 50 pm, in particular at least 100 pm, moreparticularly at least 125 pm. Likewise, each one of the laser beam pulses may have a spot diameter at the ablation position on the target surface in a range between 50 pm and 400 pm, in particular between 80 pm and 200 pm, more particularly between 100 pm and 200 pm, for example 125 pm or 150 pm.

[0064] Preferably, the one or more focused laser beam pulses are provided onto the target surface such that their focal point is at the ablation position on the target surface.

[0065] Furthermore, as also explained with respect to the device of the present invention, an optical axis along which the laser beam pulse is provided onto the target surface may form an irradiation angle with a normal to the ablation plane at the ablation position in a range between 0° and 60°, in particular between 0° and 45°, more particularly in a range between 2° and 30° or between 5° and 20° or between 5° and 15° or between 10° and 20°. Likewise, the optical axis (of the light-collecting optics) along which emission light is collected and an optical axis along which the laser beam pulse is provided onto the target surface may form an acute angle to each other, or alternatively, may be parallel to each other. The acute angle may be in a range between 1 ° and 45°, more particularly between 1 ° and 30° or between 1 ° and 20° or between 1 ° and 15° or between 5° and 20° or between 5° and 15° or between 1 ° and 10° or between 1 ° and 5° or between 1 ° and 2°.

[0066] The method may further comprise the step of generating a respective emission light spectrum of the emission light collected for each of the one or more focused laser beam pulses, in particular by directing the collected emission light to a spectrometer, resulting in a set of one or more emission light spectra.

[0067] In addition, the method may comprise the step of analyzing the set of the one or more emission light spectra, in particular analyzing emission lines in the set of one or more emission light spectra.

[0068] In order to allow for a reasonable comparison of the emission light spectra, analyzing the set of the one or more emission light spectra may preferably comprise the step of pre-processing the set of the one or more emission light spectra, resulting in a set of one or more pre-processed emission light spectra.

[0069] Preferably, pre-processing the set of the one or more emission light spectra may comprise at least one of:- applying a base line subtraction to each emission light spectrum, resulting in a respective baseline-free emission light spectrum;- normalizing each emission light spectrum, in particular each baseline-free emission light spectrum, preferably by applying a base peak intensity (BPI) normalization to each emission light spectrum, in particular to each baseline-free emission light spectrum;- applying a peak detection and quantification process to each emission light spectrum, in particular to each normalized emission light spectrum, more particularly to each baseline-free emission light spectrum, resulting in a respective emission light spectrum with detected and quantified peaks;- applying a selection process to each emission light spectrum, in particular to each normalized emission light spectrum, more particularly to each baseline-free emission light spectrum, even more particularly to each emission light spectrum with detected and quantified peaks, to exclude spectra:- comprising saturation of one or more peaks; and / or- comprising an intensity below a pre-defined intensity threshold; and / or- comprising hydrogen, oxygen and / or nitrogen emission lines above a predefined level; and / or- resulting from a plasma plume with a temperature outside a pre-defined temperature range.

[0070] Normalizing each emission light spectrum, in particular each baseline-free emission light spectrum advantageously helps to reduce the effect of signal quenching for biological sample that is caused by their relatively high water content. In particular, normalization may be a normalization with base peak intensity, wherein the largest peak is set to 100%.

[0071] The peak detection and quantification process may comprise one of peak binning, summing up or integrating of peak areas or Lorentzian peak fitting. For peak binning, summing up or integration, wavelength ranges for each peak may be defined in advance. The sum of all measured pixel intensities within a given peak represents a peak area. Advantageously, the peak detection and quantification process helps to overcome signal processing issues due to the Stark effect.

[0072] Finally, analyzing the set of the one or more emission light spectra may comprise applying statistical analysis to the set of the one or more emission light spectra, in particular to the set of the one or more pre-processed emission light spectra, to differentiate between a desired and an undesired state of the sample. For example, the analysis may allow to determine whether a given metallic sample has a desired specific composition. Alternatively, the analysis may allow to differentiate between non-cancerous tissue and cancerous tissue in a biological sample. As such, the present invention may allow for a direct, fast and independent investigation of tissue probes or biopsies for the fast differentiation of healthy and disease tissue areas of given tissues. In particular, the intracellular electrolyte content and some minor intercellular electrolytes, can be analyzed.Brief Description of the Drawings

[0073] The device and the method according to the invention is described in more detail hereinbelow by way of an exemplary embodiment and with reference to the attached drawings, in which:Fig. 1 a schematically shows an example of a LIBS setup involving an exemplary embodiment of a device according to the present invention for use in analyzing reference elements in a sample;Fig. 1 b schematically shows an alternative example of a LIBS setup involving another exemplary embodiment of a device according to the present invention;Fig. 1 c schematically shows another alternative example of a LIBS setup involving yet another exemplary embodiment of a device according to the present invention;Fig. 1 d schematically shows yet another alternative example of a LIBS setup involving still another exemplary embodiment of a device according to the present invention;Fig. 2 shows an illustration of the evolution of processes over time associated with the method and the use of the device according to present invention;Fig. 3 shows a flow diagram associated with an exemplary embodiment of a method according to the present invention;Fig. 4 illustrates an example of a LIBS spectrum obtained from performing the method according to the present invention using the device according to the present invention and as described herein;Fig. 5 shows a LIBS spectrum recorded with the same sample over the same spectral wavelength range using the same setup and the same method as in Fig. 4, yet with a detection distance of essentially zero, and a time delay At being adjusted accordingly; andFig. 6 shows the signal scatter over a total number of 30 subsequent ablation laser beam pulses taken under the conditions associated to the measurements according to Fig. 4 (lower line) and Fig. 5 (upper line).of Embodiments

[0074] In the following description, certain terms are used for reasons of convenience and are not intended to limit the invention. The terms Tight”, “left”, “up”, “down”, “under", and “above" refer to directions in the figures. The terminology comprises the explicitly mentioned terms as well as their derivations and terms with a similar meaning. Also, spatially relative terms, such as "beneath", "below", "lower", "above", "upper", "proximal", "distal", and the like, may be used to describe one element's or feature's relationship to another element or feature as illustrated in the figures. These spatially relative terms are intended to encompass different positions and orientations of the devices in use or operation in addition to the position and orientation shown in the figures. For example, if a device in the figures is turned over, elements described as "below" or "beneath" other elements or features would then be "above" or "over" the other elements or features. Thus, the exemplary term "below" can encompass both positions and orientations of above and below. The devices may be otherwise oriented (rotated 90 degrees or at other orientations), and the spatially relative descriptors used herein interpreted accordingly. Likewise, descriptions of movement along and around various axes include various special device positions and orientations.

[0075] To avoid repetition in the figures and the descriptions of the various aspects and illustrative embodiments, it should be understood that many features are common to many aspects and embodiments. Omission of an aspect from a description or figure does not imply that the aspect is missing from embodiments that incorporate that aspect. Instead, the aspect may have been omitted for clarity and to avoid prolix description. In this context, the following applies to the rest of this description: If, in order to clarify the drawings, a figure contains reference signs which are not explained in the directly associated part of the description, then it is referred to previous or following description sections. Further, for reason of lucidity, if in a drawing not all features of a part areprovided with reference signs it is referred to other drawings showing the same part. Like numbers in two or more figures represent the same or similar elements.

[0076] Fig. 1 a schematically shows an example of a LIBS setup involving an exemplary embodiment of a device 1 according to the present invention that is used for analyzing reference elements in a sample 400. In the present example, the sample 400 preferably is a biological tissue, such as a mandible bone which may contain cancerous tissue. It is known from scientific studies using other methods that in mandible bone tumor cells the calcium content is reduced, while potassium is increased. Also, the amount and presence of other reference elements, such as for example CN, may change during conversion form healthy to the tumor state. Accordingly, potassium, calcium or CN have already been used in prior art LIBS system as reference elements to indicate the eventual presence of cancerous tissue.

[0077] While such prior art LIBS systems already allow for a reasonable qualitative analysis of reference elements, quantitative analysis of the exact element ratio within a given sample remains a challenge and can only yield a rough estimate. This applies in particular for biological samples for which it is difficult to control the applied LIBS process. Here, a reasonable quantitative analysis is impaired inter alia by the formation of strong oxygen, nitrogen and hydrogen emission lines resulting from plasma reactions with air and water, by inhomogeneous material or tissue densities, by a strong quenching effect that is caused by the water content of the sample, as well as by the Stark effect that may lead to a strong peak broadening accompanied by a decrease of the peak intensity for higher element concentrations. All this is making LIBS of biological tissues often unpredictable. To overcome these difficulties, averaging over up to hundreds or even thousand laser shots often is the only possible measure to reduce the variability and differentiate between different tissues. However, averaging is only suitable for homogeneous samples. In contrast, for inhomogeneous biological tissue including cells giving a fine structure to the tissue, averaging may not be the appropriate way to analyze, as averaging over a large number of spectra taken for different laser shots and thus for different sample depth blurs the information about the locally varying sample composition.

[0078] The device and the method according to the present invention aim to overcome these issues and allow at least for an improved semi-quantitative sample analysis, in particular of biological samples, even with single-shot sensitivity.

[0079] The invention is based on the finding that an improved at least semi-quantitative sample analysis with single-shot sensitivity can be achieved, if substantially only such emission light is considered for the spectral analysis that is emitted from elements within the inner center zone of the expanding plasma plume which had sufficient time to reach an excited equilibrium state. Such elements will emit clear emission lines with an appropriate light intensity that is correlated with their actual amount present in the sample. In addition, it has been found that the signal intensity for LIBS analysis increases with the amount of collected emission light that is emitted from the respective elements in the excited equilibrium state in response to a single laser beam pulse.

[0080] Accordingly, the device and the method according to the present invention are designed in such a way to collect as much plasma emission light as possible at the right time and in the right place.

[0081] Fig. 1 a shows an exemplary embodiment of such a device 1 that can in particular be used to implement the method of the present invention, an exemplary embodiment of which is shown in Fig. 3.

[0082] Essentially, the device 1 according to Fig. 1 a comprises two main components, a laser radiation arrangement 100 and an analysis arrangement 200. While the laser irradiation arrangement 100 is used to ablate material from the target surface 401 of the sample 400 in order to create a plasma plume including the respective reference elements to be analyzed, the analysis arrangement 200 is used to collect and analyze the emission light emitted from these elements.

[0083] The laser irradiation arrangement 100 comprises a pulsed laser source 120 that is configured to generate one or more laser beam pulses 140 with a homogenous beam profile. In the present example, the laser source is a high-power Q-switched Nd:YAG laser providing laser beam pulses 140 having an adjustable pulse duration in a range between 2 ns to 10 ns and an adjustable energy per pulse in a range between 50 mJ and 150 mJ.

[0084] The laser irradiation arrangement 100 further comprises focusing means, such as a focusing lens 110 facing the target surface 401 that is configured to focus the laser beam pulses 140 along an optical axis 111 down to a predefined ablation position 130 in a predefined ablation plane 131 of the laser irradiation arrangement 100. There, the targetsurface 401 of the sample 400 is to be positioned. Preferably, the ablation position 130 is defined by the focal point 113 of the laser irradiation arrangement 100, which at hand is determined by the focal length of the focusing length 110. Preferably, the focal length of the focusing lens 110 in a range between 150 mm and 300 mm, which is rather large resulting in a large Rayleigh length of the laser beam pulses 140. In this way, the photon density in the ablation position 130 is more or less the same independent from the exact position of the focal point 113 of the laser irradiation arrangement 100 relative to the ablation position 130. A longer Rayleigh length is also desirable if the laser beam pulses 140 shall be able to drill a tunnel into the sample 400 without any widening or shrinking of the tunnel diameter. In the present embodiment, the spot diameter of the focused laser beam pulses 140 at the ablation position 130 is about 80 pm.

[0085] With the parameters discussed above, in particular with the given pulse duration, energy per pulse, and spot diameter at the ablation position 130, the focused laser beam pulses 140 are able to ablate material from the target surface 141 at the ablation position 130 causing the creation of a plasma plume 300 for each pulse 140. As shown in Fig. 1 a, the plasma plume 300 expands away from the target surface 401 together with a fastmoving shock wave front 330 induced by the plasma plume 300. More specifically, the plasma plume 300 and the shock wave front 330 expand along a main expansion axis 311 , which essentially corresponds to a normal to the target surface 401 at the ablation position 130, or to the ablation plane 131 at the ablation position 130, respectively.

[0086] The direction of the main expansion axis 311 of the plasma is substantially independent from the irradiation angle. That is, main expansion axis 311 is more or less always normal to the target surface 401 at the ablation position 130 irrespective of whether the laser beam pulses 140 hit the target surface 401 perpendicular to the target surface 401 as shown in Fig. 1 a or at an oblique angle of incidence as shown in Fig. 1 b. Preferably, the irradiation angle formed between a normal to the ablation plane 131 and the optical axis 111 of the laser irradiation arrangement 100 along which the laser beam pulses 140 are provided onto the target surface 401 is as small as possible as the photon intensity at the ablation position 130 becomes higher and more homogeneous with decreasing irradiation angle. Accordingly, Fig. 1 a shows a preferred embodiment of the laser irradiation arrangement 100 in which the irradiation angle formed between a normal to the ablation plane 131 and the optical axis 111 of the laser irradiation arrangement 100 is 0°. However, larger angles are also possible as shown in Fig. 1 b which schematicallyillustrates an alternative example of a LIBS setup involving an alternative, though quite similar embodiment of the device 1. More particularly, the device 1 according to Fig. 1 b comprises a laser irradiation arrangement 100, the optical axis 111 of which forms an irradiation angle [3 with a normal to the ablation plane 131 of about 15°.

[0087] The expanding plasma plume breaks down the ablated material into excited ionic, atomic and molecular species including the refence elements of interest to be analyzed. As the plasma cools down, electrons that were stripped from the ionic, atomic and molecular species recombine with the respective species, thereby emitting light in the UV, optical and infra-red spectrum with characteristic emission lines of the respective species, in particular with characteristic emission lines associated with the reference elements of interest.

[0088] The time course of this light emission process is illustrated in Fig. 2. As shown, the emission of ionic lines typically occurs in range 510 between a few tens of nanoseconds to a few tens of microseconds after provision of the respective laser beam pulse 140, whereas the emission of atomic and molecular lines occurs later. More specifically, the emission of atomic lines mainly occurs in an interval 520 between 150 ns to 200 ps after a laser beam pulse, whereas the emission of molecular lines occurs in range 530 between a few tens of microseconds and some hundred microseconds.

[0089] In addition to the ionic, atomic and molecular emission light, the plasma emission further comprises continuum emission, in particular electron cyclotron emission due to free electrons in the plasma, emission due to bremsstrahlung radiation and recombination reactions. This continuum emission typically occurs in the very first phase of the plasma expansion, mostly in the interval 540 between the first few nanoseconds up to about hundred nanoseconds.

[0090] Apart from the emission lines associated with the material composition of the sample, the light emitted from the plasma during an early phase of the plasma expansion (0 - 0.8 ps after application of a laser beam pulse) also includes emission lines for nitrogen (N), oxygen (O) and hydrogen (H), which result from reactions of the hot plasma plume 300 with ambient air right after application of the laser beam pulse 140.

[0091] In order to collect the plasma emission light for spectral analysis, the analysis arrangement 200 includes light-collecting optics 210. At hand, the light-collecting optics210 comprises at least one collector lens 212 defining an optical axis 211 of the lightcollecting optics 210 along which emission light 525 is collected.

[0092] As stated above, the device and method according to the present invention aim to collect and analyze emission light 525 resulting from reference elements within the inner center zone 310 of the expanding plasma plume 300 which had sufficient time to reach an excited equilibrium state. Typically, this state is only reached after a few hundred nanoseconds after application of the ablating laser beam pulse 140, depending on the laser energy converted into plasma temperature and the material properties of the sample. During this time period, the plasma plume 300 and the respective reference elements have moved away from the target surface 310. That is, depending on the laser energy converted into plasma temperature and the material properties of the sample, the desired center zone 310 of the expanding plasma plume 300 can typically be found a few millimeters above the target surface 401 , after an expansion time which is required to reach that position and during which the plasma homogenizes and the reference elements reach the desired excited equilibrium state.

[0093] Consequently, the light-collecting optics 210 is configured and arranged such that a focal point 213 of the light-collection optics 210 is located above the ablation plane 131 at a corresponding detection distance 215 to the ablation position 130 (see dashed double arrow 215 in Figs. 1 and 2) in order to collect emission light 525 from this zone 310 of the plasma plume 300 at a distance above the target surface 401 . For the given laser beam pulse energy and material properties of the sample 400 in the present example, the detection distance 215 may be about 5 mm.

[0094] To fade out emission light that does not result from the respective reference elements of interest in the desired equilibrium state, such as the continuum emission light and the emission lines for nitrogen (N), oxygen (O) and hydrogen (H), collection of the emission light 525 only starts with a time delay AT after the provision of the respective laser beam pulse (as indicated in Fig. 2). Referring to the time scales discussed above with respect to the background emission, the time delay preferably is at least 0.8 ps. After this period of time, the fast-moving shock wave front 330 induced by the expanding plasma plume has also typically passed the light-collecting area 214 around the focal point 213 of the light-collection optics 210. Of course, the time delay again depends on the expansion dynamics of the plasma plume which in turn depends on the laser energy converted into plasma temperature and the material properties of the sample. For thegiven laser beam pulse energy and material properties of the sample 400 in the present example, the time delay AT may be about 1 .5 ps.

[0095] The emission light 525 is collected over a sampling time window 250 of a certain length T in order to collect a sufficient amount of emission light 525 to achieve a sufficient signal intensity, preferably such as to enable single-shot sensitivity. The sampling time window 250 and its length T are indicated in Fig. 2 as well. However, the longer the sampling time window 250, the more ambient light is collected in addition to the emission light 525 which might affect the sensitivity of the analysis. Hence, the length T of the sampling time window preferably is limited in order to avoid ambient light effects. For the example of Fig. 2, the length T of the sampling time window 250 is in a range between 1 .0 ps and 3.0 ps, which advantageously may still allow for single-shot sensitivity.

[0096] According to the present invention, the signal intensity and thus the analysis sensitivity is further enhanced by arranging the light-collecting optics 210 such that the optical axis 211 of the light-collecting optics 210 along which emission light 525 is collected has an acute viewing angle <p to the main expansion axis 310. Advantageously, a small viewing angle <p allows the light emitting reference elements within the expanding plasma plume 300 to stay longer within the viewing field / light-collecting area 214 of the light-collecting optics 210, or vice versa enables the light-collecting optics 210 to track the light emitting atoms within the expanding plasma plume 300 over a longer sampling time. This proves particularly beneficial since light emission occurs stochastically which requires the sampling time to be as long as possible in order to collect more emission light 525. A longer sampling time also enhances the reproducibly of the analysis, since fluctuations in light emission (due to its stochastic nature) average out over a longer sampling time. In the embodiment of Fig. 1a, the viewing angle <p is about 13°. The amount of collected emission light 525 and thus the sensitivity increases as the viewing angle <p decreases. It reaches a maximum for a viewing angle <p of 0°. This situation is depicted in Fig. 1 b.

[0097] In general, it is also possible that both the viewing angle <p and the irradiation angle [3 are greater than 0°. This situation is depicted in Fig. 1 c.

[0098] Likewise, it is also possible that the optical axis 211 of the light-collecting optics 210 along which emission light is collected and an optical axis 111 of the laser irradiation arrangement 100 along which the laser beam pulse is provided onto the target surface401 are parallel to each other, at least along a certain common optical path 411 . That is, the optical axis 211 of the light-collecting optics 210, along which emission light is collected, and an optical axis 111 of the laser irradiation arrangement 100, along which the laser beam pulse is provided onto the target surface 401 , may coincide over a common optical path 411 of a certain length and somewhere be separated, e.g. by means of a beam splitter 260, such as beam splitter cube a glass plate or a dichromatic mirror. This configuration is depicted in Fig. 1 d, which proves particularly advantageous in terms of both the irradiation process as well as the detection sensitivity of the light-collecting optics, since both the viewing angle <p and the irradiation angle [3 are equal to 0°.

[0099] Preferably, the light-collecting optics 210, in particular the collector lens 212 of the light-collecting optics 210 has a focal length in a range between 50 mm and 500 mm, in particular between 100 mm and 400 mm or between 100 mm and 300 mm or between 150 mm and 300 mm, more particularly between 180 mm and 250 mm, for example 200 mm. A focal length within these ranges constitutes a good comprises between a sufficiently high collected radiant flux and a sufficiently long light-collecting area 214 or detection volume along the optical axis 211 of the light-collecting optics 210.

[0100] For analysis of the emission light, the light-collecting optics 210 is further configured to transfer the collected emission light 525 to a spectrometer 220, for example by means of an optical fiber (not shown). Alternatively, the light transfer can be accomplished at least partially in free space, in particular by means for directing / deflecting the collected emission light 525, such as deflecting mirrors.

[0101] In the present embodiment, the spectrometer 220 comprises a spectral separator 221 , such as a prism or a diffraction grating, to separate the collected emission light 525 into its spectral components, and a detector array 222 to detect an intensity of the separated spectral components, resulting in a respective emission light spectrum for each laser beam pulse. Preferably, the spectrometer 220 is a low-resolution spectrometer to ensure that a single plasma (laser shot) may provide enough emission light 525 to a respective detector pixel in order to allow for single-shot sensitivity.

[0102] Once the emission light spectrum is taken, it can be further analyzed by the analysis arrangement 200. For this, the analysis arrangement 200 may comprise computational means that may be configured to evaluate / analyze the spectral data provided by the spectrometer, in particular to carry out the steps of analyzing emissionlight spectra, in particular the steps of pre-processing and applying statistical analysis, as defined in the context of the method according to the present invention. Details of the spectrum analysis are now discussed with respect to Fig. 3

[0103] Fig. 3 shows a flow diagram illustrating the steps of an exemplary embodiment of the method according to the present invention that aims to analyze reference elements in a sample, such as a biological tissue, by laser induced breakdown spectroscopy. The method may preferably be carried out by using one of the exemplary embodiments of the device as described before.

[0104] In the present embodiment, the method comprises the step 1000 of providing one or more focused laser beam pulses at a predefined ablation position on a target surface of the sample in order to ablate material from the target surface such that a plasma plume including one or more reference elements is created for each pulse that expands away from the target surface along a main expansion axis substantially normal to the target surface at the ablation position. The method further comprises the step 2000 of collecting emission light from the plasma plume for each of the laser beam pulses at a distance offset from the target surface with a viewing angle relative to the main expansion axis in a range between 0° and 60°, wherein the emission light is collected over a sampling time window starting with a time delay relative to the provision of the respective focused laser beam pulse onto the target surface.

[0105] The method according to present embodiment further comprises the step 3000 of generating a respective emission light spectrum of the emission light collected for each of the one or more focused laser beam pulses, in particular by directing the collected emission light to a spectrometer, resulting in a set of one or more emission light spectra.

[0106] In addition, the method of present embodiment comprises the step 4000 of analyzing the set of the one or more emission light spectra, in particular the step of analyzing emission lines in the set of one or more emission light spectra.

[0107] According to the present exemplary embodiment, the step 4000 of analyzing the set of the one or more emission light spectra in particular comprises the step of preprocessing 4001 the set of the one or more emission light spectra, which results in a set of one or more pre-processed emission light spectra.

[0108] The step 4100 of pre-processing the set of the one or more emission light spectra may comprise at least one of the following steps:- applying a base line subtraction to each emission light spectrum, resulting in a respective baseline-free emission light spectrum;- normalizing each emission light spectrum, in particular each baseline-free emission light spectrum, preferably by applying a base peak intensity (BPI) normalization to each emission light spectrum, in particular to each baseline-free emission light spectrum;- applying a peak detection and quantification process to each emission light spectrum, in particular to each normalized emission light spectrum, more particularly to each baseline-free emission light spectrum, resulting in a respective emission light spectrum with detected and quantified peaks;- applying a selection process to each emission light spectrum, in particular to each normalized emission light spectrum, more particularly to each baseline-free emission light spectrum, even more particularly to each emission light spectrum with detected and quantified peaks, to exclude spectra:- comprising saturation of one or more peaks; and / or- comprising an intensity below a pre-defined intensity threshold; and / or- comprising hydrogen, oxygen and / or nitrogen emission lines above a predefined level; and / or- resulting from a plasma plume with a temperature outside a pre-defined temperature range.

[0109] In the present exemplary embodiment, the step 4000 of analyzing the set of the one or more emission light spectra further comprises the step 4200 of applying statistical analysis to the set of the one or more pre-processed emission light spectra, in order to differentiate between a desired and an undesired state of the sample, for example to differentiate between non-cancerous and cancerous tissue in a biological tissue sample.

[0110] As such, the shown embodiments of the device and the method may, for example, enable to determine the relative amounts of one or more specific reference elements, e.g. K (potassium), at a specific position within a biological tissue sample of a specific individuum relative to at other positions within the same sample. This in turn may allow to determine a one or two-dimensional spatial profile of the (relative) K amounts across the sample. For this, the sample may, for example, be moved parallel to theablation plane into several positions, wherein for each position a (single) laser beam pulse is provided onto the respective portion of the sample surface that is currently in the ablation position in order to generate a plasma plume to be analyzed that finally allows to determine the relative local K content at each position. In this regard, the device and the method according to the present are particularly advantageous as they allow for a singleshot sensitivity, i.e. to determine the relative local K content at a specific position with one single laser shot. The resulting spatial profile may further allow to identify the spatial expansion or margins of potentially cancerous tissue in that sample, wherein the spatial expansion or margins may be given by the parts of the sample where the (relative) K content for the given sample of the individuum exceeds a certain threshold value.

[0111] Fig, 4 illustrates an example of a LIBS spectrum obtained from performing the method according to the present invention using the device according to the present invention and as described herein. The diagram of Fig. 4 shows the measured spectral intensity (ordinate) versus the wavelength of the emission light collected from the respective plasma plume (abscissa). The spectrum covers the spectral wavelength range between 190 nm and 890 nm. The emission spectrum was taken with a time delay At of 7 ps delay time and a detection distance of 9 mm. That is, the focal point of the lightcollection optics was located above the ablation plane at a detection distance of 9 mm to the ablation position at the sample surface. Most important, the emission spectrum was taken for a single ablation laser beam pulse.

[0112] For comparison, Fig. 5 shows a LIBS spectrum recorded with the same sample over substantially the same spectral wavelength range (190 nm to 890 nm) using the same setup and the same method. Yet, in contrast to the measurement associated with Fig. 4, the detection distance was reduced to essentially zero, and the time delay At was adjusted accordingly, more specifically reduced to 1 ps. That is, while the focal point of the light-collection optics for the measurement associated with Fig. 4 was located significantly spaced apart from the ablation plane and thus spaced apart from the sample surface, the focal point of the light-collection optics for the measurement associated with Fig. 5 was located essentially in the ablation plane, i.e. on the sample surface.

[0113] Fig. 6 shows the signal scatter (ordinate) over a total number of 30 subsequent ablation laser beam pulses (abscissa), taken on the one hand under the conditions of the measurement associated with Fig. 4 (lower line) and on the other hand under the conditions of the measurement associated with Fig. 5 (upper line).

[0114] As follows from a direct comparison of Fig. 4 and Fig. 5 as well as from a comparison of the lower and upper lines in Fig. 6, the LIBS spectra taken under the conditions used for the measurement associated with Fig. 4 show stable signals, low scatter, and a flat base line, which demonstrates that the device and the method according to the present invention allow for an accurate and reliable (semi-)quantitative sample analysis on a single shot basis. In contrast, the LIBS spectra taken under the conditions used for the measurement associated with Fig. 5 show no stable signals, high scatter, and a complex base line. Due to these limitations, the latter spectra are not useable for a quantitative sample analysis.

[0115] In this respect, Figs.4, 5 and 6 clearly illustrate the significant difference in quality between the spectra and highlight the outstanding achievements of the device and method according to the present invention. These achievements are based in particular on the fact that the focal point of the light-collection optics is located considerably spaced apart from the ablation plane and thus spaced apart from the sample surface.

[0116] This description and the accompanying drawings that illustrate aspects and embodiments of the present invention should not be taken as limiting the claims defining the protected invention. In other words, while the invention has been illustrated and described in detail in the drawings and foregoing description, such illustration and description are to be considered illustrative or exemplary and not restrictive. Various mechanical, compositional, structural, electrical, and operational changes may be made without departing from the spirit and scope of this description and the claims. In some instances, well-known circuits, structures and techniques have not been shown in detail in order not to obscure the invention. Thus, it will be understood that changes and modifications may be made by those of ordinary skill within the scope and spirit of the following claims. In particular, the present invention covers further embodiments with any combination of features from different embodiments described above and below.

[0117] The disclosure also covers all further features shown in the figures individually although they may not have been described in the afore or following description. Also, single alternatives of the embodiments described in the figures and the description and single alternatives of features thereof can be disclaimed from the subject matter of the invention or from disclosed subject matter. The disclosure comprises subject matter consisting of the features defined in the claims or the exemplary embodiments as well as subject matter comprising said features. Also, the present disclosure covers intermediategeneralizations of features or groups of features of the embodiments described and shown in the figures. I.e., specific features or groups of features as disclosed in the figures and the associated sections of the description may be combined with the more general embodiments of the invention disclosed in connection with the description of the invention. In particular, such specific features or groups of features may be provided in the more general embodiments of the invention in isolation from further specific features shown in the figure. It is understood that those skilled in the art are able to incorporate specific features from the description of the figures into the embodiments of the description of the invention.

[0118] Furthermore, in the claims the word "comprising" does not exclude other elements or steps, and the indefinite article "a" or "an" does not exclude a plurality. A single unit or step may fulfil the functions of several features recited in the claims. The mere fact that certain measures are recited in mutually different dependent claims does not indicate that a combination of these measures cannot be used to advantage. The terms “essentially”, “about”, “approximately” and the like in connection with an attribute or a value particularly also define exactly the attribute or exactly the value, respectively. The term “about” in the context of a given numerate value or range refers to a value or range that is, e.g., within 20%, within 10%, within 5%, or within 2% of the given value or range. Components described as coupled or connected may be electrically or mechanically directly coupled, or they may be indirectly coupled via one or more intermediate components. Any reference signs in the claims should not be construed as limiting the scope.

Claims

New International Patent Application Raylution AGAttorney ref.: P6844PC00CLAIMS1 . A device (1 ) for use in analyzing reference elements in a sample (400), in particular in a biological tissue, by laser induced breakdown spectroscopy, the device (1 ) comprising: a laser irradiation arrangement (100) configured to provide one or more focused laser beam pulses (140) at a predefined ablation position (130) in a predefined ablation plane (131 ) of the laser irradiation arrangement (100), at which a target surface (401 ) of the sample (400) is to be positioned, wherein the one or more laser beam pulses (140) are configured to ablate material from the target surface (401 ) when positioned at the ablation position (130), thereby creating a respective plasma plume (300) including one or more reference elements, the plasma plume (300) expanding away from the target surface (401 ) along a main expansion axis (311 ) substantially corresponding to a normal to the ablation plane (131 ) at the ablation position (130); an analysis arrangement (200) including light-collecting optics (210) configured to collect emission light (525) from the respective plasma plume (300) for each of the one or more laser beam pulses (140), wherein the light-collecting optics (210) is configured and arranged such that an optical axis (211 ) of the light-collecting optics (210) along which emission light (525) is collected has a viewing angle (cp) to the main expansion axis (311 ) in a range between 0° and 60° and such that a focal point (213) of the light-collection optics (210) is located above the ablation plane (131 ) at a detection distance (215) to the ablation position (130) in order to collect emission light (525) from the plasma plume (300) at a distance above the target surface (401 ), and wherein the analysis arrangement (200) is configured to collect emission light (525) from the plasma plume (300) over a sampling time window (250) starting with a time delay (At) relative to the provision of the laser beam pulse (140) to the ablation position (130).

2. The device (1 ) according to claim 1 , wherein the time delay (At) is chosen, in particular adjustable, based on at least one of the following parameters: the detection distance (215), an energy of the laser beam pulse (140), a spot diameter of the laser beam pulse (140) at the ablation position (130), a material composition of the sample (400) at the target surface (401 ), a density of the sample (400) at the target surface (401 ), a water content of the sample (400) at the target surface (401 ), and a maximum temperature within the plasma plume (300).

3. The device (1 ) according to any one of the preceding claims, wherein the time delay (At) is at least 0.8 ps, in particular at least 1 .0 ps, more particularly at least 1 .3 ps, preferably at least 1 .5 ps or 2.0 ps or at least 2.5 ps or at least 3.0 ps or at least 4.0 ps or at least 5.0 ps or at least 7.5 ps or at least 10.0 ps; or wherein the time delay (At) is in a range between 0.8 ps and 30.0 ps, in particular between 1.0 ps and 25.0 ps, more particularly between 1 .3 ps and 20.0 ps or betweenl .5 ps and 20.0 ps or between 2.0 ps and 20.0 ps or between 2.5 ps and 20.0 ps or between 1 .3 ps and 15.0 ps or between 1 .5 ps and 15.0 ps or between 3.0 ps and 15.0 ps.

4. The device (1 ) according to any one of the preceding claims, wherein the detection distance (215) is chosen, in particular adjustable, based on at least one of the following parameters: the time delay (At), an energy of the laser beam pulse (140), a spot diameter of the laser beam pulse (140) at the ablation position (130), a material composition of the sample (400) at the target surface (401 ), a density of the sample (400) at the target surface (401 ), a water content of the sample (400) at the target surface (401 ), and a maximum temperature within the plasma plume (300).

5. The device (1 ) according to any one of the preceding claims, wherein the detection distance (215) is at least 2 mm, in particular at least 5 mm, more particularly at least 10 mm or at least 15 mm; or wherein the detection distance (215) is in a range between 2 mm and 30mm, in particular between 5 mm and 20 mm, more particularly in a range between 7.5 mm and 15 mm, for example 10 mm.

6. The device (1 ) according to any one of the preceding claims, wherein a length (T) of the sampling time window (250) is in a range between 0.5 ps and 10.0 ps, in particular between 0.75 ps and 5.0 ps, more particularly between 1.0 ps and 3.0 ps; or wherein a length (T) of the sampling time window (250) is at least 0.5 ps, inparticular at least 0.75 ps, more particularly at least 1.0 ps or at least 1.5 ps or at least 2.0 ps.

7. The device (1 ) according to any one of the preceding claims, wherein the viewing angle (cp) is in a range between 0° and 45°, in particular in a range between 0° and 30° or between 2° and 30° or between 0° and 20° or between 2° and 20° or between 0° and 15° or between 2° and 15°, more particularly in a range between 0° and 10° or between 5° and 15° or between 10° and 20°.

8. The device (1 ) according to any one of the preceding claims, wherein the lightcollecting optics (210), in particular a collector lens (212) of the light-collecting optics (210) facing the sample (400), has a focal length in a range between 50 mm and 500 mm, in particular between 100 mm and 400 mm or between 100 mm and 300 mm or between 150 mm and 300 mm, more particularly between 180 mm and 250 mm, for example 200 mm.

9. The device (1 ) according to any one of the preceding claims, wherein the laser irradiation arrangement (100) is configured to provide the one or more focused laser beam pulses (140) to have a spot diameter at the ablation position (130) in the ablation plane (131 ) of at least 30 pm, in particular at least 50 pm, more particularly at least 100 pm, preferably at least 125 pm; or wherein the laser irradiation arrangement (100) is configured to provide the one or more focused laser beam pulses (140) to have a spot diameter at the ablation position (130) in the ablation plane (131 ) in a range between 30 pm and 400 pm, in particular between 50 pm and 200 pm or between 80 pm and 200 pm, more particularly between 100 pm and 200 pm, for example 125 pm or 150 pm.

10. The device (1 ) according to any one of the preceding claims, wherein the laser irradiation arrangement (100) is configured to focus the laser beam pulses (140) to have a focal point (113) at the ablation position (130) in the ablation plane (131 ).11 . The device (1 ) according to any one of the preceding claims, wherein an optical axis (111 ) of the laser irradiation arrangement (100) along which the laser beam pulse (140) is provided onto the target surface (401 ) forms an irradiation angle ([3) with a normal to the ablation plane (131 ) at the ablation position (130) in a range between0° and 60°, in particular between 0° and 45°, more particularly in a range between 2° and 30° or between 5° and 20° or between 5° and 15° or between 10° and 20°; and / or wherein the optical axis (211 ) of the light-collecting optics (210) along which emission light (525) is collected and an optical axis (111 ) of the laser irradiation arrangement (100) along which the laser beam pulse (140) is provided onto the target surface (401 ) form an acute angle to each other, or are parallel to each other.

12. The device (1 ) according to any one of the preceding claims, wherein the laser irradiation arrangement (100) comprises at least one pulsed laser source (120), in particular at least one pulsed laser source with Q-switching, more particularly at least one pulsed laser source with passive Q-switching.

13. The device (1 ) according to any one of the preceding claims, wherein the analysis arrangement (200) is configured to analyze emission lines in the emission light (525) collected for each of the one or more focused laser beam pulses (140).

14. The device (1 ) according to claim 13, wherein the analysis arrangement (200) comprises a spectrometer (220) for generating a respective emission light spectrum of the emission light (525) collected for each of the one or more focused laser beam pulses (140).

15. The device (1 ) according to claim 14, wherein the light-collecting optics (210) is configured to transfer the emission light (525) collected for each of the one or more focused laser beam pulses (140) to the spectrometer (220).

16. The device (1 ) according to any one of claim 14 or 15, wherein the spectrometer (220) is a low-resolution spectrometer, in particular a low-resolution spectrometer having a resolving power of at most 6’000 and / or having a wavelength range between 180 nm and 1000 nm.

17. A method of analyzing reference elements in a sample (400), in particular in a biological tissue, by laser induced breakdown spectroscopy, preferably by using a device (1 ) according to any one of the preceding claims, the method comprising: providing one or more focused laser beam pulses (140) at a predefined ablation position (130) on a target surface (401 ) of the sample (400), wherein theone or more focused laser beam pulses (140) are configured to ablate material from the target surface (401 ), thereby creating a respective plasma plume (300) including one or more reference elements, the plasma plume (300) expanding away from the target surface (401 ) along a main expansion axis (311 ) substantially corresponding to a normal to the target surface (401 ) at the ablation position (130); collecting emission light (525) from the plasma plume (300) for each of the one or more focused laser beam pulses (140) for spectral analysis; wherein collecting emission light (525) for each of the one or more focused laser beam pulses (140) comprises collecting emission light (525) from the plasma plume (300) at a viewing angle (cp) relative to the main expansion axis (311 ) in a range between 0° and 60° at a distance, in particular a detection distance (215), offset from the target surface (401 ) and over a sampling time window (250) starting with a time delay (At) relative to the provision of the respective focused laser beam pulse (140) onto the target surface (401 ).

18. The method according to claim 17, wherein collecting emission light (525) for each of the one or more focused laser beam pulses (140) comprises using light-collecting optics (210), wherein the emission light (525) is collected along an optical axis (211 ) of the light-collecting optics (210) orientated at the viewing angle (cp) relative to the main expansion axis (311 ), and wherein a focal point of the light-collection optics (210) is located above the target surface (401 ) at the detection distance (215) to the ablation position (130).

19. The method according to any one of claim 17 or 18 comprising generating a respective emission light spectrum of the emission light (525) collected for each of the one or more focused laser beam pulses (140), in particular by directing the collected emission light (525) to a spectrometer, resulting in a set of one or more emission light spectra.

20. The method according to claim 19, further comprising analyzing the set of the one or more emission light spectra, in particular analyzing emission lines in the set of one or more emission light spectra.21 . The method according to 20, wherein analyzing the set of the one or more emission light spectra comprises pre-processing the set of the one or more emission light spectra, resulting in a set of one or more pre-processed emission light spectra.

22. The method according to 21 , wherein pre-processing the set of the one or more emission light spectra comprises:- applying a base line subtraction to each emission light spectrum, resulting in a respective baseline-free emission light spectrum;- normalizing each emission light spectrum, in particular each baseline-free emission light spectrum, preferably by applying a base peak intensity (BPI) normalization to each emission light spectrum, in particular to each baseline- free emission light spectrum;- applying a peak detection and quantification process to each emission light spectrum, in particular to each normalized emission light spectrum, more particularly to each baseline-free emission light spectrum, resulting in a respective emission light spectrum with detected and quantified peaks;- applying a selection process to each emission light spectrum, in particular to each normalized emission light spectrum, more particularly to each baseline- free emission light spectrum, even more particularly to each emission light spectrum with detected and quantified peaks, to exclude spectra:- comprising saturation of one or more peaks; and / or- comprising an intensity below a pre-defined intensity threshold; and / or- comprising hydrogen, oxygen and / or nitrogen emission lines above a pre-defined level; and / or- resulting from a plasma plume (300) with a temperature outside a predefined temperature range.

23. The method according to any one of claims 20 to 22, wherein analyzing the set of the one or more emission light spectra comprises applying statistical analysis to the set of the one or more emission light spectra, in particular to the set of the one or more pre-processed emission light spectra, to differentiate between a desired and an undesired state of the sample (400).

24. The method according to any one of claims 17 to 23, wherein the sample (400) is an extracted biological tissue such that the method is an ex-vivo method.

25. The method according to any one of claims 17 to 24 not being a method for treatment of the human or animal body by surgery or therapy, nor a diagnostic method practiced on the human or animal body.

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