DE102005048807B3 - Apparatus for quantitative or qualitative determination of infrared active contents of liquid based or non-liquid fluids - Google Patents

Abstract

The apparatus has two optical waveguides enclosing a measurement surface which are transparent and have a refractive index greater than that of the fluid. A sample holder (2) has a receptacle (8) for the fluid with an edge in contact with a waveguide. A positioning device brings the fluid surface and the measurement surface of one of the waveguides (4) reversibly into contact. Independent claims also cover use of the device and a method of determining the contents of the fluid.

Description

The The present invention relates to a device for qualitative and / or quantitative determination of IR-active ingredients in liquids and a method for qualitative and / or quantitative determination of IR-active ingredients in liquids.

In medical analysis in particular, constant attempts are being made to reduce the amount of sample required for an analysis. At the same time, as many measurement parameters as possible should be determinable from such a sample in as short a time as possible. From the DE 103 28 998 A1 An IR-ATR based device for the analysis of lowest sample quantities is to be found, in which a metering device for quantities in the nl range and an ATR device in a single instrument are combined. However, components of the blood such as glucose, cholesterol, triglycerides, albumin, total protein and urea can only be determined with this device if the sample is dried on the measurement surface before measuring the ATR-IR spectrum. The drying of the sample to be measured is regularly accompanied by a considerable non-linearity in the determination of the concentration.

ATR immersion probes for the measurement of whole blood are used in the US 5,170,056 described. Measuring solutions can be removed away from the actual spectrometer via a light guide.

A mass flow rate analysis device based on optical measurements is known in the US 6,469,311 disclosed. A large number of liquid samples are determined, for example, in a microtiter plate in a contactless manner by means of optical analysis methods comprising the measurement of the absorption, the photoluminescence and the chemiluminescence.

Also WO 96/04544 A1 deals with an automated method for Determination of blood components. The liquid sample is placed in a receiving device entered to be angle-dependent scattered light and fluorescence signals to detect. To this Way is information about the red and white blood corpuscle available in the blood. The device presented in WO 98/04544 A1 essentially comprises a combination of a conventional hematology analyzer and a fluorescence cytometry analyzer.

The WO 00/66269 A1 has an integrated sample processing system for Subject to prepare samples and make them available for analysis. The device proposed in WO 00/66269 A1 has a liquid source, a connected pump, an output unit and a line from the pump to the dispensing unit, the line open Remains between consecutive contactless fluids a volume of less than 5 μl. For this purpose, preference is given to a dispensing tip made of a hydrophobic Material used. Further information, for example, on specific analytical methods, WO 00/66269 A1 are not removable.

The US 6,187,267 describes a device with which the chemiluminescence of liquid samples is measured. This analyzer is designed to ensure a high sample throughput.

The US 6,808,934 The invention relates to a device and a method by which reservoirs can be filled with quantities of less than 100 .mu.l in order to then be exposed to sound waves which are intended to force the precipitation of dissolved constituents.

The used devices to measure a variety of samples at a high throughput are still very expensive and apparatus associated in their results with relatively large error tolerances, especially if only very small amounts of sample are available.

Of the The present invention was therefore based on the object, a device available too do not deal with the disadvantages of the prior art is afflicted and in particular, even at high sample throughput, from very small fluid sample volumes the simultaneous determination of a multitude of parameters or ingredients as well as easy to use and to load and without further, and without it special methods or materials would need to be cleaned.

Accordingly, an apparatus has been found for the quantitative and / or qualitative determination of IR-active ingredients of aqueous or non-aqueous liquids comprising at least a first ATR body in the form of a light guide comprising a measuring surface in contact with the liquid to be measured which is transparent or partially transparent to the measuring radiation and which has a refractive index which is greater than that of the adjacent to the measuring surface during the measurement liquid, at least one second ATR body, the at least one flat boundary surface, in particular at least two planar, in substantially parallel boundary surface Chen, one of which is the passing of the liquid to be measured in contact measuring surface, and which is transparent or partially transparent to the measuring radiation and which has a refractive index which is greater than that of the measurement adjacent to the measuring surface liquid, at least a sample holder, comprising at least one receiving device for the liquid to be measured with a bearing surface for this liquid and an edge which is dimensioned to contact the measuring surface of the first or second ATR body with the surface of the present in the receiving device to be measured liquid and at least one positioning device for reversibly contacting the surface of the liquid and the measuring surface of the first or second ATR body.

The Measuring surface of Light guide and the preferably flat measuring surface of the second ATR body have a continuous, uniform measuring path, within the one measuring beam preferably at least twice, preferably at least four times, attenuated is totally reflectable, i. with an adjacent, in particular the measuring surface only thoroughly wetted, liquid Medium can interact. In general, 2 to 20 total reflections along a measuring path sufficient to get enough Data for to collect the evaluation. Preferably, the refractive index of the first and second ATR body bigger or equal to 1.5. The in the analysis device according to the invention for Use to be coming ATR body also referred to in the art as ATR crystals, although these Systems are not necessarily in crystal form. Accordingly, presents e.g. also sintered silver chloride a functional ATR body As material for the used ATR body Any material suitable for the radiation used, especially for electromagnetic radiation in the mid-infrared range, transparent is and that about it is also highly refractive or high refractive and over a Refractive index has, the is larger as that of air and / or as the one with the infrared measuring device according to the invention to analyzing or analyzable medium. Suitable materials for the ATR body include diamond, sapphire, cadmium telluride, thallium bromide iodide, Silicon, germanium, zinc selenide, zinc sulphide, magnesium difluoride, cesium iodide, Silver chloride, calcium difluoride, potassium bromide or sodium chloride. For the One skilled in the art will recognize that certain the above-mentioned materials, as long as not with a suitable coating due to their water solubility for the Measurement of aqueous Systems are not suitable, e.g. an ATR body of sodium chloride. In a further embodiment gets on ATR body from one for Infrared radiation transparent material, in particular a polymer material, with a refractive index preferably ≥ 1.5, especially polyethylene, resorted.

Under the proviso that the second ATR body over at least one, in particular at least two flat, substantially parallel boundary surfaces has, can otherwise this ATR body be present in any geometry, as long as this allows an incident Beam can be adjusted so that this before exiting the ATR body Totally reflected at least twice attenuated is. The same applies to the first ATR body. In one embodiment The invention will be with six or seven such total reflections already optimal analysis results in terms of sensitivity, Accuracy and speed also for aqueous multicomponent systems achieved. Under test section In the present case, that section of the ATR body which is in contact is understood occurs with the medium to be analyzed and the total for attenuated total reflections to disposal stands.

With the device according to the invention a sample holder is preferably used with a receiving device, which is designed to be at least one, in particular exactly one, liquid drops take. In particular, this receiving device is designed with an amount of liquid a volume in the range of 0.1 to 400 .mu.l, in particular from 1 to 50 ul record.

amounts of liquid in the nl and μl range are about suitable pipetting readily in the receiving devices to apply. For example, see WO 00/66269 A1 described output unit. The expert is also available Automatic pipetting robots are available, which allow a high mass flow rate is easily accessible. Nor is it special Difficulty, a precisely defined quantity or a given number of drops, for example, 1, 2, 3, 4, 5, 6 or 7 drops in the receiving device to apply.

A receiving device is preferably provided in the sample holder, in which the side of the liquid to be measured lying opposite the support surface lies at least in regions above the edge of the receiving device when the sample holder is substantially vertical. This constellation can be set for the skilled person readily depending on eg the capacity and the diameter of the receiving device and the polarity of liquid and inner wall material of the receiving device. In this case, the surface tension of the liquid quantity to be measured present in the receiving device causes liquid to flow over the edge of the receptacle protrudes direction and thus even for such a measuring surface of an ATR body is accessible, which is larger than the size of the receiving device. The embodiment described above finds particular application in very small-sized receiving devices, for example with a volume in the range of 1 to 100 ul. Of course, the amount of liquid present in the receiving device can also be dimensioned such that it does not protrude beyond the edge of this receiving device. In this case, the ATR body is brought to the surface of the liquid present in the receiving device so long, or conversely, the receiving device to the ATR body until the measuring surface of the ATR body is wetted by the liquid. The edge of the receiving device is thus dimensioned in height and / or extent or extent that liquid protrudes beyond the edge due to surface tension phenomena and is available for contact with the measuring surface of an ATR body or that the ATR body has a smaller size Expansion has as the edge and thus can be introduced into the receiving device.

Also particularly advantageous is a measuring surface of the ATR body in the range of 1 to 10 mm ² , in particular in the range of 3 to 20 mm ² . The device according to the invention accordingly allows the use of a smallest possible dimensioned ATR body with which an IR measurement is still possible. On the one hand, this considerably reduces the material costs and, on the other hand, allows the design of very small-dimensioned systems which can be used without difficulty for, for example, point-of-care measurements. By only very small amounts of liquid are required when using the device according to the invention, for example, in the blood test, the required amount of blood can be drastically reduced. So far, for usual blood tests about 5 ml of blood required, it now comes out for the same investigation with only about 5 ul. Moreover, the amount of waste products that regularly accumulates when larger amounts of blood are withdrawn can be significantly reduced. For example, it is not uncommon for conventional clinics to take about 1,200 samples per day.

A very appropriate development draws characterized by the fact that the Measuring surface at least partially has a, in particular hydrophobic, coating, the for the measuring radiation is transparent.

Especially if the coating is a starch which is less than the wavelength of the measuring radiation used, can on any for the measuring radiation transparent coating material can be used. The fat or strength the coating, however, is not critical when it comes to the coating material for such a for an ATR body is, i. e. has approximately a refractive index, that of the respective used ATR body equivalent. For example, an ATR material such as zinc sulfide or zinc selenide, which is particularly preferred for the invention used ATR body is used, be provided with a layer of diamond. you gets up this way one with an extremely resistant and inert coating provided ATR body. Particularly preferred the diamond layer after one of H.J. Neubert in Optics, February 2002, page 11, described method. After that will with the help of a carbon dioxide laser with a power of about 6 up to 7 kW nearby a surface a temperature in the range of 15,000 to 20,000 ° C generated. By initiating Argon in this area, a plasma is generated. Are gaseous hydrocarbons, For example, methane, in this plasma, so are free carbon atoms formed on a substrate, in this case, for example on a flat boundary surface the second ATR body, precipitate to form a very thin layer of diamond. Of advantage is the previously described coating in particular for materials for ATR body, the toxic, soluble, z. B. in the sample medium, and / or sensitive to mechanical stress are. Especially suitable for the measuring device according to the invention are ATR body of zinc sulfide or zinc selenide coated with a coating, in particular a diamond coating, are provided.

In a preferred embodiment, the coating of the ATR body of the device according to the invention has a thickness which is less than the, preferably half, wavelength of the infrared measuring radiation used, in particular over a thickness in the range of about 2 nm to about 250 nm Thickness of the coating is particularly preferably in a range of about one quarter of the measuring wavelength. Furthermore, it is recommended to use a coating which is homogeneous in terms of thickness and composition and has the smoothest possible surface. It is advantageous if the unevenness in the coating does not exceed the extent of about one quarter of the measuring wavelength on average. Suitably, recourse is made to those coatings which do not reflect the measurement radiation, even partially. The coating performs a dual function by protecting the test medium from contamination with, possibly toxic, ATR body material as well as the ATR body against mechanical damage on the one hand. In the investigation of aqueous systems, in particular coatings have proven that a diffusion from water molecules through this layer to the ATR body.

In a further preferred embodiment may instead of a diamond coating, a layer of a transparent or translucent plastic, in particular polyethylene be used.

In a further advantageous embodiment, the with a Coating providable ATR body materials selected of sapphire, cadmium telluride, thallium bromide iodide, silicon, germanium, Zinc sulphide, zinc selenide magnesium difluoride, cesium iodide, silver chloride, calcium difluoride, Potassium bromide and sodium chloride, with zinc selenide and zinc sulfide are preferred.

In As a rule, the receiving devices are in the form of hollows before and are preferably in a field or array of recording devices summarized.

The Recording device, for example, also part of a Device for taking blood, containing at least one capillary gap, be or represent such. Such blood collection devices are for example as so-called test strips for diabetics known.

there can be provided that the Sample holder a carrier, for example in the form of a plate, e.g. similar to a microtiter plate, with at least two, preferably at least eight and in particular a plurality of recording devices.

When practicable has e.g. proven when at least eight picking devices arranged in a row.

advantageously, lie in or on the sample holder several rows of recording devices successive and form, for example, an array of receiving wells, as known for example from microtiter plates. alternative can the rows of receptacles also star-shaped, circular, spiral or be arranged linearly on the sample plate. Preferably are at a star-shaped Arrangement in each row the corresponding recording devices arranged approximately equidistant from the center and are therefore on the same circumference. In this way, by rotation of the sample holder, the ATR body can rapidly, unproblematic and reliable over one new recording device are positioned.

For a linear Arrangement of the recording devices already enough only two such Recording devices. These can, for example, alternately with a sample liquid and subsequently with a rinsing liquid filled become. In combination with a suitable pipetting robot allowed already such a sample holder a continuous sample measurement at high throughput.

Of the Sample holder features preferably via a planar geometry, but may e.g. also be constructed stepwise. The Recording devices can be placed on the sample holder or attached or can be integral Be part of the same. According to one In another embodiment, the sample holder has openings or inlets, in each case a receiving device, in particular reversible, can be inserted or recessed. For example, in such a sample holder recorded such recording devices containing a blood collection device containing at least a capillary gap, represent or are part of such a device.

According to one Another aspect of the invention is provided that the positioning device designed is to the measuring surface of the ATR body towards a in a receiving device of the sample holder amount of liquid present or away from and / or around the sample holder in the direction of the measuring surface or move away from it, in particular to make contact or cancel.

satisfactory Results occur especially when the infrared light source one or more quantum cascade lasers.

Quantum cascade lasers that are suitable for the device according to the invention, for example, from EP 0 676 839A as well as from the US 5,509,025 in which their basic operation and their structure are described. Preferably, recourse is made to quantum cascade lasers which emit electromagnetic radiation in the mid-infrared range. For example, such quantum cascade lasers come into consideration for the device according to the invention, which emit only a defined frequency, in particular from the mid-infrared range, as well as those which emit two, three, four, five or more frequencies, in particular from the mid-infrared range can. Furthermore, it is also possible to make use of such quantum cascade lasers which emit defined frequency bands. Of course, these Infrarotmeßvorrichtungen can not be equipped with only one, but with two or more of the previously described quantum cascade laser. If in a device according to the invention a quantum cascade laser is used, which is capable of at least electromagnetic radiation mindes at least two different frequencies, in particular from the mid-infrared range, or if several quantum cascade lasers are used side by side in such a measuring device, the electromagnetic radiation, in particular if it has different frequencies, at the same time or almost simultaneously or in chronological order be emitted. In this way it is possible to comprehensively characterize the spectrometric behavior of a substance in a sample. Furthermore, it is possible to examine several ingredients present in a sample in the shortest possible time, ie simultaneously or almost simultaneously. Almost simultaneously or simultaneously within the meaning of the present invention means that signals are radiated so slightly offset in time, that no significant differences from the absorption signals detected at the same time emitted radiation can be seen from the respective detected absorption signals.

In a preferred embodiment becomes from the used quantum cascade laser electromagnetic Radiation in the form of pulses with a defined period of time and / or intensity emitted. This pulse duration and / or intensity is free within wide ranges selectable and can be used to test for each ingredient to be tested to generate optimized spectrometric examination conditions. So, if several ingredients in a sample are to be measured, different depending on the frequency of the emitted electromagnetic radiation long pulse durations and / or different intensities are selected. For example, you can Ingredients with weakly absorbing chromophores Pulses exposed to longer pulse duration be while For strongly absorbing substances very short pulse durations are sufficient to to be able to detect a satisfactory signal. Already this different Absorption behavior can for the analysis will be used and in operative association with a detector standing for a detector for the purpose of immediate adjustment stored or incorporated into suitable analysis programs. Accordingly, it is with the device according to the invention possible, electromagnetic radiation of different frequency, either starting from only a Quan tenkaskadenlaser or by use several quantum cascade lasers, in any order, for example to broadcast in sequential order. This is especially true also to the previously described pulsed radiation.

About the free choice of the pulse sequence of the radiated frequencies of the electromagnetic radiation can According to the invention pulse pattern and / or intensity pattern be used, which are tailored to the particular analysis problem. For example, you can with knowledge or justified Guess in a liquid present or possibly present ingredients, the frequencies, pulse durations and / or intensities such be given that about the Art and the scope of the detected signals by means of an evaluation unit, in particular a computer-aided Evaluation unit, can easily determine which ingredients in which concentrations are present in the examined sample. These Pattern of pulse sequence, pulse length and / or pulse intensity of two or more frequencies can in turn be used to generate certain patterns of response signals to produce that for certain compositions are characteristic. In this way Is it possible, within the shortest possible time Time to determine if or in which concentration certain ingredients present in a sample. Accordingly, electromagnetic is particularly preferred Radiation of different frequency and / or intensity after one Multiplex pattern in particular pulsed, radiated. This can be on one known multiplex spectrometer or electrical multiplexers, which are primarily used on the detector side come, resorted become. On the other hand, such multiplexers are preferably used, which not only influence the detector or wavelength-specific but also the wavelengths to be emitted and / or intensities choose and control the broadcasting sequence or the broadcasting pattern. The present invention used Multiplexer controls according to another embodiment additionally also the intensity the respective measuring radiation, especially depending from the respective measurement problem. These so-called optical multiplexers thus coordinate the radiation source and the detector and match them. Switching such multiplexers among other things, measuring radiation after one on the measuring problem tailored pulse sequence on and off, in particular under wavelength-dependent modulation the intensity, and thus control the light source. The detected signals become preferably by known methods such as factor analysis, Multiple Least Square Algorithms or Neural Network Analysis evaluated. Therefor gets up regularly computerized Evaluation units used.

In addition, using quantum cascade lasers, it is possible to transmit not only radiation of different frequencies and / or intensities in a particular time sequence through the ATR body. Rather, one can also radiate different frequencies at the same time, in particular, the number and frequency of the emitted radiation can be varied continuously. In this way, it is possible to assemble frequency patterns which generate characteristic absorption signals for compositions to be investigated, as a result of which multicomponent mixtures, in particular especially in aqueous systems, can be analyzed in a short time. The analysis method described above may also be referred to as matrix coding / matrix decoding. A characteristic feature associated with the device according to the invention is that in multicomponent mixtures, several ingredients can be determined simultaneously and qualitatively next to each other.

alternative of course also be resorted to infrared light sources, which is a continuous Emit spectrum.

such Light sources are those skilled in the art, for example, as Nernst pins, which consist essentially of zirconium oxide and rare earth additives, and as so-called globars, essentially consisting of silicon carbide, known. Further comes as a light source, an electrically conductive ceramic in question. in principle can Light sources are used, which are over the entire spectral Infrared range or just about emit certain areas of this spectrum. For the determination of Ingredients with the FT-IR measuring device according to the invention preferably such light sources are used, in the middle Infrared range emit electromagnetic radiation, ie in Range of about 1 micron to about 25 microns.

Ideally The positioning device is designed to keep a distance between the measuring surface and the edge of the receiving device, which is smaller than 1 mm, preferably less than 15 microns, in particular without the Measuring surface the Edge touched. The ATR body and liquid sample in the recording device are usually only as long as relative to each other moved towards each other until wetting by capillary effect the measuring surface of the ATR body with the liquid he follows. The distance between measuring surface and edge can be generally in wide limits are varied. However, it has proven to be beneficial proved the distance in dependence from the volatility the liquid to be measured, the measuring temperature and / or the duration of the measurement not too big, so while the measuring phase not so much fluid that can evaporate the concentration of the ingredients changes noticeably. positioning devices, which are capable of receiving device, pipetting device and / or the first or second ATR body to bring to the desired position with very high accuracy are known in the art. Accuracies in the μm range are easy possible.

The inventive device is regularly equipped with an infrared measuring device comprising the ATR body, at least one infrared light source, at least one detector and at least one evaluation unit. Integrated in the evaluation unit or separately, there may be a display unit.

When Detectors for the registration of the measuring radiation can be used on all common, in infrared measuring devices to be used for coming systems.

So far the detected signals further processed or evaluated should be, come for this the skilled person known evaluation units, in particular computer-aided evaluation units into consideration. An evaluation unit in the sense of the present invention may also comprise a data memory and / or a display unit, e.g. a pen or a screen. These elements can Of course also available separately.

A embodiment the invention provides that the Evaluation unit is suitable to those recorded by the detector Evaluate signals by Fourier transform. That in the detector of FT-IR spectrometry device recorded interferogram, the an overlay of all the wavelengths occurring in the spectrum is recorded in the Evaluation unit computer-aided by Fourier transformation into the frequencies of the individual oscillations disassembled. Details of the Fourier transform are e.g. at N.B. Colthup, L.H. Daly, S.E. Wiberley, Introduction to Infrared and Raman Spectroscopy, Academic Press, San Diego, 1990, to which reference is hereby made. With the FT-IR measuring device can be several ingredients at the same time with substantially high sensitivity, speed and wavenumber precision.

Of further can the device of the invention at least one filling device for filling and / or rinsing at least one, in particular all Receiving devices, and / or at least one drying device for drying at least one emptied and / or rinsed receiving device include.

in this connection it is easily possible the positioning so interpreted to a variety of on the sample holder present receiving devices or the therein present, to be measured liquid successively, in particular automatically, to the measuring surface of the ATR body zoom and after the measurement again lead away.

The inventive device is suitable for, in particular simultaneous, qualitative and / or Quantitative determination of IR-active ingredients in aqueous or non-aqueous liquids.

When come to be measured aqueous liquids e.g. in question body fluids such as blood, serum, saliva, sweat, semen, urine, lymph, spinal fluid and interstitial body fluid as well beverages like beer, wine, milk, dairy, fruit juice, spirits or Soft drinks.

In body fluids such as blood, serum, urine, saliva, sweat, sperm or interstitial body fluid can with the device according to the invention e.g. medically relevant components such as glucose, albumin, total protein, Urea, cholesterol and / or tricglycerides and their concentrations simultaneously side by side with high accuracy and high throughput be determined.

Of others can also non-aqueous liquids, For example, systems containing at least one in an organic solvent dissolved IR-active substance can be measured with the device according to the invention. In this context, the device according to the invention, for example be used in the investigation of color or lacquer patterns.

• a) Einführen einer Flüssigkeitsprobe, insbesondere mindestens eines Tropfens, der zu vermessenden Flüssigkeitsprobe in eine erste Aufnahmevorrichtung des Probenhalters,
• b) Inkontaktbringen der Meßfläche des ersten oder zweiten ATR-Körpers mit der Oberfläche der in einer ersten Aufnahmevorrichtung vorliegenden, zu vermessenden Flüssigkeitsprobe, so daß eine Fläche bedeckt ist, die so groß ist wie oder größer ist als die optisch aktive ATR-Fläche,
• c) Vermessen der Flüssigkeitsprobe, enthaltend mindestens einen IR-aktiven Inhaltsstoff unter Verwendung einer Infrarotmeßvorrichtung, umfassend das Durchleiten von von einer Infrarotlichtquelle ausgehenden IR-Strahlung durch den ATR-Körper und Detektieren der von dem ATR-Körper ausgehenden Signale in mindestens einer Detektiereinheit, und
• d) Entfernen der Meßfläche des ATR-Körpers von der Oberfläche der Flüssigkeit, wobei nach Schritt c) und/oder d) die detektierten Signale in einer Auswerteeinheit zwecks qualitativer und/oder quantitativer Bestimmung mindestens eines Inhaltsstoffs der Flüssigkeitsprobe ausgewertet werden.

The object underlying the invention is further achieved by a method for the quantitative and / or qualitative determination of IR-active ingredients in aqueous or non-aqueous liquids using a device according to the invention, comprising the steps

• a) introducing a liquid sample, in particular at least one drop, of the liquid sample to be measured into a first receiving device of the sample holder,
• b) contacting the measuring surface of the first or second ATR body with the surface of the liquid sample to be measured in a first receiving device so as to cover an area as large as or greater than the optically active ATR surface,
• c) measuring the liquid sample containing at least one IR-active ingredient using an infrared measuring device comprising passing infrared radiation emanating from an infrared light source through the ATR body and detecting the signals emanating from the ATR body in at least one detecting unit, and
• d) removing the measuring surface of the ATR body from the surface of the liquid, wherein after step c) and / or d) the detected signals are evaluated in an evaluation unit for the purpose of qualitative and / or quantitative determination of at least one ingredient of the liquid sample.

there Preferably, a plurality of present in the liquid sample Measure IR-active ingredients simultaneously and substantially simultaneously determined qualitatively and / or quantitatively. For example, let easily five, ten or even 20 ingredients essentially simultaneously qualitative and / or determine quantitatively.

The Steps a) to d) can Of course As often as required, they can be switched or repeated in succession at each new step a) preferably a certain amount of liquid a new sample to be examined is given up.

• e) Reinigen der Meßfläche des ATR-Körpers,
• f) Inkontaktbringen der gereinigten Meßfläche des ATR-Körpers mit der Oberfläche einer in einer zweiten Aufnahmevorrichtung vorliegenden Flüssigkeit,
• g) Vermessen der Flüssigkeitsprobe, enthaltend mindestens einen IR-aktiven Inhaltsstoff unter Verwendung einer Infrarotmeßvorrichtung, umfassend das Durchleiten von von einer Infrarotlichtquelle ausgehenden IR-Strahlung durch den ATR-Körper und Detektieren der von dem ATR-Körper ausgehenden Signale in mindestens einer Detektiereinheit, und
• h) Entfernen der Meßfläche des ATR-Körpers von der Oberfläche der Flüssigkeit, wobei nach Schritt g) und/oder h) die detektierten Signale in einer Auswerteeinheit zwecks qualitativer und/oder quantitativer Bestimmung mindestens eines Inhaltsstoffs der Flüssigkeitsprobe ausgewertet werden.

Accordingly, a further development is characterized by the following additional steps:

• e) cleaning the measuring surface of the ATR body,
• f) contacting the cleaned measuring surface of the ATR body with the surface of a liquid present in a second receiving device,
• g) measuring the liquid sample containing at least one IR-active ingredient using an infrared measuring device comprising passing IR radiation emanating from an infrared light source through the ATR body and detecting the signals emanating from the ATR body in at least one detection unit, and
• h) removing the measuring surface of the ATR body from the surface of the liquid, wherein after step g) and / or h) the detected signals in an evaluation unit for the purpose of qualitative and / or quantitative determination of at least one ingredient of the liquid sample are evaluated.

conveniently, the steps e) to h) become at least twice and in series of measurements repeatedly through with a high sample throughput, wherein in the receiving device in each case a new liquid sample to be measured is present.

The cleaning step e) can take place, for example, in such a way that in a first step e ₁ ) a receiving device on a sample holder with a rinsing or

Cleaning liquid is filled, which is then brought reversibly in a second step e ₂ ) with the measuring surface of the ATR body in contact.

• i) Einführen einer Referenzflüssigkeit, insbesondere mindestens eines Tropfens der Referenzflüssigkeit, in eine dritte Aufnahmevorrichtung des Probenhalters,
• j) Inkontaktbringen der Meßfläche des ersten oder zweiten ATR-Körpers mit der Oberfläche der in der dritten Aufnahmevorrichtung vorliegenden, zu vermessenden Referenzflüssigkeit, so daß vorzugsweise eine Fläche bedeckt ist, die so groß ist wie oder größer ist als die optisch aktive ATR-Fläche,
• k) Vermessen der Referenzflüssigkeit, enthaltend mindestens einen IR-aktiven Referenzinhaltsstoff unter Verwendung einer Infrarotmeßvorrichtung, umfassend das Durchleiten von von einer Infrarotlichtquelle ausgehenden IR-Strahlung durch den ATR-Körper und Detektieren der von dem ATR-Körper ausgehenden Signale in mindestens einer Detektiereinheit, und
• l) Entfernen der Meßfläche des ATR-Körpers von der Oberfläche der Referenzflüssigkeit, wobei nach Schritt k) und/oder 1) die detektierten Signale in einer Auswerteeinheit zwecks qualitativer und/oder quantitativer Bestimmung mindestens eines Referenzinhaltsstoffs der Flüssigkeitsprobe ausgewertet werden.

In accordance with a further aspect of the method according to the invention, additional method steps are provided which can be followed, for example, by steps d), e) and / or h):

• i) introducing a reference liquid, in particular at least one drop of the reference liquid, into a third receiving device of the sample holder,
• j) contacting the measuring surface of the first or second ATR body with the surface of the reference liquid to be measured in the third recording device, so that preferably an area as large as or greater than the optically active ATR surface is covered,
• k) measuring the reference liquid containing at least one IR-active reference ingredient using an infrared measuring device comprising passing IR radiation emanating from an infrared light source through the ATR body and detecting the signals emanating from the ATR body in at least one detecting unit, and
• l) removing the measuring surface of the ATR body from the surface of the reference liquid, wherein after step k) and / or 1) the detected signals in an evaluation unit for the purpose of qualitative and / or quantitative determination of at least one reference ingredient of the liquid sample are evaluated.

Of the Detection process in the context of the present invention comprises in general also saving the recorded signals or the recorded Raw data set in a suitable memory, so that this Data will be available for further evaluation. Suitable detection units and devices for storing larger amounts of data are those skilled in the art adequately known.

Of course it is it also possible, in particular in a sample holder comprising a plurality of receiving devices, these recording devices first with measuring liquid and optionally cleaning and / or reference liquid to fill. Subsequently can the measuring surface of ATR body with the surface the liquid samples to be measured be brought into contact one after the other. Preferably, the measuring surface of the ATR body first cleaned after each contact with a liquid to be measured, preferably by contacting with that in a receiving device present cleaning or rinsing liquid.

According to one another embodiment the measuring surface of the ATR body before taking the survey of present in the recording devices fluid samples and / or in the course of measuring a plurality of such liquid samples at least once with a reference liquid brought into contact. In this way, a calibration or recalibration can be made become. In a preferred embodiment of the method according to the invention serves the reference liquid at the same time as rinsing liquid, whereby both the material and the process cost again can be reduced.

In a preferred embodiment of the method according to the invention, the receiving devices present on a sample holder are first filled with the liquid samples to be measured and optionally the rinsing and / or reference liquid, ie the steps a) and optionally e ₁ ) and / or i) are preceded. Subsequently, the sequences of steps b) -c) -d) and optionally f) -g) -h) and / or j) -k) -l) are then used. These sequences can be repeated as desired or combined with each other.

In a sample holder can For example, recording devices alternately according to the following pattern filled be: measuring liquid, rinse, ..., rinse, etc. Alternatively, the sequence may provide: reference liquid, rinse, measuring liquid, Rinsing liquid, ..., Reference liquid measuring liquid, rinse, Etc.

Basically, the respective liquids be combined in any order in their sequence.

The measurement result let yourself optimize again by the fact that the Measuring surface of ATR body after the contacting with the present in a receiving device, to be measured liquid with a cleaning liquid, that of a further, in particular adjacent, receiving device is present on the sample holder, is brought into contact.

From Of particular importance is that the measuring liquid in general only for a duration of 1 to 60 seconds is to be measured. Reach regularly for one Sample also measurement times from 2 to 30 seconds already completely out, with even measuring times in the range of 1 to 3 seconds are readily accessible. Due to the small sample volumes and the very short measuring time per Sample can without further sample holder are used, which has more as 100, for example 500 to 1000 or 1000 to 2000 recording devices feature.

Especially reliable measurement results Imagine when a variety of on a sample holder in Receiving devices present liquid drop successively, especially automatically, be measured. Of particular advantage is further that at least a liquid to be measured represents a reference sample. moreover can be provided that the Measuring surface of ATR body after a measuring step and / or a cleaning step is subjected to a drying step.

Of the present invention was the surprising Understanding that even with a non-contactless optical measuring method for Determination of ingredients in liquids a very high mass flow rate achieve, namely even when using sample volumes in μl range, without the accuracy suffer from. Another advantage is that a sample preparation or Evaporation or drying of the sample is eliminated and you also not on relies on the use of analytical reagents. Through the miniaturization both the receiving device for the liquids to be measured as well as the actual measuring body itself The material and cost can be significantly reduced. Furthermore, one is no longer dependent on a regular recalibration, to be reliable measurement results to ensure. moreover allows the method according to the invention a linear calibration over a very wide measuring range. Also eliminates the generally necessary dilution series. The otherwise common encountered problems with the turbidity of solutions in photometric Determinations, for example as from the measurement of solutions with high cholesterol known, occur with the device according to the invention not up. In particular, the presence of suspended particles, i.e. particulate Ingredients, in the measuring liquid for the Goodness of Measurement result at Use of the device according to the invention not of relevance and upset the measurement accordingly not. For example, you can do not deposit any suspended particles on the measuring surface and then not or difficult to remove. It is also of particular advantage that in a single measurement also from a very small sample, the only 1 to 5 μl comprises several parameters or ingredients at the same time qualitatively and can be determined quantitatively. About that in addition, the miniaturized version the ATR unit and the associated small measurement volumes the Use of the method according to the invention in neonatal and paediatrics, Where so far mainly test strip methods due to the low blood levels available be used. However, these methods are much too inaccurate and also not very reliable. While for example, in the measurement of blood after conventional Methods relatively large Amounts required are sufficient according to the inventive method already amounts in the lower μl Area for reliable Measurements off. Another advantage here is that even only still very small volumes of reference solution is required, which across from usual Methods brings a significant cost savings. In any case can with the method according to the invention be assured that not more at the expense of accuracy, the number of reference measurements over the required level is reduced.

From Another advantage is that with the device according to the invention the liquid samples to be measured very promptly can be measured for sampling. This makes itself for example in the determination of the glucose content in the blood is significantly noticeable. For example, if a measurement is delayed, due to the in the blood sample continues to degrade glucose too low a glucose level certainly. As a rule, it is sufficient for the preparation of the to be measured Blood samples from these with an anticoagulant agent. If necessary, you can also be added to agents that inhibit glucose degradation, for example Sodium fluoride.

Further embodiments The invention will be described in detail with reference to the following figures described without the Invention be limited to these particular embodiments should. Show it

1 a cross-sectional view of a sample holder and an ATR body of the device according to the invention;

2 a cross-sectional view of a sample holder and an ATR body of an alternative embodiment of the device according to the invention;

3 a plan view of a sample holder of an embodiment of the device according to the invention;

4 a plan view of an alternative sample holder for a device according to the invention; and

5 a plan view of an alternative sample holder for a device according to the invention.

The 1 shows a sample holder 2 a device according to the invention 1 as well as an ATR body 4 in cross section. The sample holder 2 has essentially a flat base plate 6 on the trough-shaped receiving devices 8th attached or integrated. The recording devices 8th lift up with their edge 10 from the plane spanned by the base plate level and preferably form the vertical point in the highest point of the receiving device. In the trough-shaped receiving device 8th there is a liquid drop to be measured 12 , An ATR body 4 becomes over the cradle in the way 8th positioned that its measuring surface 14 in contact with the liquid droplet 12 can occur. The drop of liquid 12 lies on the shelf 16 the cradle 8th on and is surrounded by the perimeter 10 limited. Due to surface tension phenomena, the liquid drop protrudes 12 in vertical alignment over the edge 10 out and so stands for the contact with the measuring surface 14 of the ATR body 4 to disposal. Preferably, the sample holder 2 in the direction of the arrow to a fixed ATR body 4 with the help of a positioning device 18 brought on and off again. The positioning device 18 is both able to align the sample holder in the vertical direction, as well as after completion of the measurement of the drop 12 ie after shutting down the sample holder 2 move it horizontally until the next, preferably adjacent receiving device containing a liquid drop 20 directly below the measuring surface 14 of the ATR body 4 to come to rest. Then, the positioning device 18 bring the sample holder back to the measuring surface, but without it to a contact of the measuring surface 14 with the edge 10 the cradle 8th comes.

The embodiment of the analysis device according to the invention 1 according to 2 differs from the according to 1 solely by not being an ATR body 4 but a variety of ATR bodies 4 . 4 ' . 4 '' . 4 ''' available. In this way, the mass flow rate can be increased considerably again.

3 gives a view of a sample holder 2 a device according to the invention 1 again. This sample holder 2 has a rectangular base with a total of 96 receptacles 8th which are present in an 8 × 12 field (indicated). Here are each eight recording device 8th arranged side by side in a row objected to on a line. In a preferred embodiment, the field which is moved up first to the measuring surface of the ATR body may contain a drop of a reference liquid KA, with which the device is first calibrated. The following recording device in the series then contains a cleaning liquid R, to which a first liquid M to be measured follows in the following receiving device. Thereafter, in turn, cleaning and liquid samples to be measured close to the end of the first row. Finally, the positioning device moves the sample holder to the adjacent first receiving device of the second row, which, in turn, may for example be first filled with a reference liquid KA for the purpose of calibration. Subsequently, recording devices alternate with cleaning and liquid to be measured. Following this pattern, the entire sample holder can be fed.

4 shows an alternative sample holder 2 on which the recording devices 8th are arranged on a circumference. For the movement of the recording devices in the desired position, an angle motor is sufficient. In this variant, it is of particular advantage that it can easily be used fully or semi-automatically for a continuous operation mode. Thus, for example, in areas that are spaced from the fixed ATR body, already measured measuring liquid or once used cleaning or reference liquid disposed of their receiving devices and optionally cleaned and then filled again with liquid to be measured or cleaning or reference liquid become.

In 5 is an alternative embodiment of a sample holder 2 played. On the sample holder are rows of recording devices 8th arranged substantially star-shaped. In this embodiment, positioning of a receiving device can be effected by rotation of the sample holder under an ATR body.

Of course they are Numerous other variants of sample holders in any geometry and Arrangement of the individual receiving devices for the device according to the invention used.

The in the above description, in the claims and in the drawings disclosed features of the invention can both individually and also in any combination for the realization of the invention in their various embodiments be essential.

Claims (37)

Contraption ( 1 for the quantitative and / or qualitative determination of IR-active ingredients of aqueous or non-aqueous liquids, comprising a) at least one first ATR body in the form of a light guide, which comprises a contact surface coming into contact with the liquid to be measured, the is transparent or partially transparent to the measuring radiation and has a refractive index which is greater than that of the liquid adjacent to the measuring surface during the measurement, at least one second ATR body ( 4 ) having at least one flat boundary surface, which communicates with the liquid to be measured in contact surface ( 14 ) which is transparent or partially transparent to the measuring radiation and which has a refractive index which is greater than that of the liquid adjacent to the measuring surface during the measurement, b) at least one sample holder ( 2 ), containing at least one receiving device ( 8th ) for the liquid to be measured with a bearing surface ( 16 ) for this fluid and a rim ( 10 ) dimensioned to allow contact of the measuring surface ( 14 ) of the first or second ATR body ( 4 ) with the surface of the in the receiving device ( 8th ) to ensure the liquid to be measured, and c) at least one positioning device ( 18 ), the surface of the liquid and the measuring surface ( 14 ) of the first or second ATR body ( 4 ) reversibly in contact. Contraption ( 1 ) according to claim 1, characterized in that the receiving device ( 8th ) is adapted to receive at least one drop. Contraption ( 1 ) according to claim 1 or 2, characterized in that the receiving device ( 8th ) is adapted to receive a quantity of liquid having a volume in the range of 0.1 to 400 .mu.l, in particular from 1 to 50 ul. Contraption ( 1 ) according to one of the preceding claims, characterized in that in the receiving device ( 8th ) of the bearing surface ( 16 ) opposite side of the liquid to be measured in a substantially vertical orientation of the sample holder ( 2 ) above the edge ( 10 ) of the receiving device ( 8th ) lies. Contraption ( 1 ) according to one of the preceding claims, characterized in that the measuring surface ( 14 ) of the ATR body ( 4 ) has an area in the range of 1 to 100 mm ² , in particular in the range of 2 to 20 mm ² . Contraption ( 1 ) according to one of the preceding claims, characterized in that the measuring surface ( 14 ) at least partially has a, in particular hydrophobic, coating. Contraption ( 1 ) according to one of the preceding claims, characterized in that the receiving device ( 8th ) is designed trough-shaped. Contraption ( 1 ) according to one of the preceding claims, characterized in that the receiving device is a device for collecting blood, containing at least one capillary gap, or is part of the same. Contraption ( 1 ) according to one of the preceding claims, characterized in that the sample holder ( 2 ) a carrier with a plurality of receiving devices ( 8th ). Contraption ( 1 ) according to claim 9, characterized in that the receiving devices ( 8th ) on the sample holder ( 2 ) are arranged linearly or spirally. Contraption ( 1 ) according to claim 9 or 10, characterized in that the sample holder at least two, in particular a plurality of successive linear rows of receiving devices ( 8th ). Contraption ( 1 ) according to claim 9, characterized in that on the sample holder a plurality of rows of receiving devices are aligned with each other in a star shape. Contraption ( 1 ) according to one of the preceding claims, characterized in that the sample holder contains a multiplicity of openings or inlets, in each of which there is a device for collecting blood, containing at least one capillary gap, or a component thereof. Contraption ( 1 ) according to one of the preceding claims, characterized in that the positioning device ( 18 ) is designed around the measuring surface ( 14 ) of the ATR body ( 4 ) in the direction of one in a receiving device ( 8th ) of the sample holder ( 2 ) or away from and / or around the sample holder ( 2 ) in the direction of the measuring surface ( 14 ) or to move away from it, in particular to establish or cancel a contact. Contraption ( 1 ) according to one of the preceding claims, characterized in that the positioning device ( 18 ) is adapted to a distance between the measuring surface ( 14 ) and the edge ( 10 ) of the receiving device ( 8th ), which is smaller than 1 mm, preferably smaller than 15 μm, without the measuring surface ( 14 ) the edge ( 10 ) touched. Contraption ( 1 ) according to one of the preceding claims, further characterized by an infrared measuring device comprising the ATR body ( 4 ), at least one infrared light source, at least one detector and at least one evaluation unit. Contraption ( 1 ) according to claim 16, characterized in that the infrared light source comprises one or more quantum cascade lasers. Contraption ( 1 ) according to claim 16, characterized in that the infrared light source emits a continuous spectrum. Contraption ( 1 ) according to claim 16, characterized in that the evaluation unit is suitable for evaluating the signals recorded by the detector by means of Fourier transformation. Contraption ( 1 ) after one of the preceding The claims, further comprising at least one filling device for filling and / or rinsing at least one, in particular all receiving devices, and / or at least one drying device for drying at least one emptied and / or rinsed receiving device. Contraption ( 1 ) according to one of the preceding claims, characterized in that the positioning device ( 18 ) is designed to hold a plurality of on the sample holder ( 2 ) receiving devices ( 8th ) or the liquid to be measured therein successively, in particular automatically, to the measuring surface ( 14 ) of the ATR body ( 4 ) and back off after the measurement. Use of the device according to one of the preceding claims for, in particular simultaneous, qualitative and / or quantitative Determination of inactive ingredients in aqueous or non-aqueous liquids. Use according to claim 22, characterized that the to be measured aqueous liquids Blood, serum, saliva, sweat, Sperm, urine, lymph, spinal fluid, interstitial body fluid, Beer, milk, milk products, wine, fruit juice, spirits or soft drinks include and that the non-aqueous liquids organic solvents include. Use of the device according to one of claims 1 to 21 for the quantitative determination of glucose, albumin, total protein, Urea, cholesterol and / or triglycerides in blood, serum, urine, Saliva, sweat, Sperm or interstitial body fluid. Method of quantitative and / or qualitative Determination of IR-active ingredients in aqueous or non-aqueous liquids using a device according to claims 1 to 21, comprising steps a) Introduce a fluid sample, in particular at least one drop of the liquid sample to be measured in a first receiving device of the sample holder, b) bring into contact the measuring surface of first or second ATR body with the surface the present in a first recording device, to measure the liquid, so that one area that's so big like or bigger as the optically active ATR surface, c) Measuring the liquid sample, containing at least one IR-active ingredient using an infrared measuring device, comprising the passage of emanating from an infrared light source IR radiation through the ATR body and detecting the signals originating from the ATR body in at least one detection unit, and d) removing the measuring surface of ATR body from the surface the liquid, wherein after step c) and / or d) the detected signals in a Evaluation unit for the purpose of qualitative and / or quantitative determination evaluated at least one ingredient of the liquid sample become. The method of claim 25, further comprising steps e) cleaning the measuring surface of the ATR body, f) Contacting the cleaned measuring surface of the ATR body with the surface a present in a second receiving device to be measured Fluid sample G) Measuring the liquid sample, containing at least one IR-active ingredient using an infrared measuring device, comprising the passage of emanating from an infrared light source IR radiation through the ATR body and detecting the signals originating from the ATR body in at least one detection unit, and h) removing the measuring surface of ATR body from the surface the liquid; wherein after step g) and / or h) the detected signals in a Evaluation unit for the purpose of qualitative and / or quantitative determination evaluated at least one ingredient of the liquid sample become Method according to claim 26, characterized in that that the Steps e) to h) at least twice, in particular several times through become. Method according to one of Claims 25 to 27, characterized that the Measuring surface of the ATR body after the contacting with the present in a receiving device, to be measured liquid with a cleaning fluid in another, in particular adjacent, receiving device is present on the sample holder, is brought into contact. Method according to one of claims 25 to 28, characterized that the liquid in the cradle for a duration of 1 to 60 seconds is measured. Method according to one of claims 25 to 29, characterized in that a plurality of on the sample holder in the receiving devices ( 8th ) present liquid quantities successively, in particular automatically, is measured. Method according to one of claims 25 to 30, characterized that at least a present in a receiving device of the sample holder, liquid to be measured represents a reference sample. Method according to one of claims 25 to 31, characterized that the Measuring surface of the ATR body after a measuring step and / or a cleaning step to a drying step becomes. Method according to one of claims 25 to 32, characterized that the Receiving devices on or in the sample holder circumferentially are arranged at least one circumference. Method according to one of claims 25 to 32, characterized that the in the receiving device present measuring liquid has a volume in the range from 0.1 to 400 μl, in particular 1 to 50 μl, having. Method according to one of Claims 25 to 33, characterized that the Measuring surface at least having in sections one, in particular hydrophobic, coating, in particular with a thickness in the range of 2 nm to 250 nm. Method according to one of claims 25 to 35, characterized that one Variety of in the liquid sample present IR-active ingredients measured simultaneously and in the substantially simultaneously qualitatively and / or quantitatively determined. The method of any of claims 25 to 36, further comprising the steps i) Introduce a reference liquid, in particular at least one drop of the reference liquid, in a third receiving device of the sample holder, j) contact the measuring surface of first or second ATR body with the surface the present in the third recording device, to be measured Reference liquid so that preferably an area that's so big like or bigger as the optically active ATR surface, k) Measuring the reference liquid, containing at least one IR-active reference ingredient below Use of an infrared measuring device, comprising the passage of emanating from an infrared light source IR radiation through the ATR body and Detecting the from the ATR body outgoing signals in at least one detection unit, and l) Remove the measuring surface of the ATR body from the surface the reference liquid, in which after step k) and / or 1) the detected signals in an evaluation unit for the purpose of qualitative and / or quantitative determination at least evaluated a reference ingredient of the liquid sample become.