Precise magnetic bio transducer

FIELD: physics.

SUBSTANCE: proposed device comprises sensitive surface consisting of binding sections that allow specific attachment to biological objects with magnetic labels, magnetic pickup element, appliance to differentiate between magnetic labels connected to binding sections and magnetic labels, not bound, to act on binding sections. System to determine concentration of target objects consists of measuring device and electronic circuit to detect changes in magneto resistive resonance of magnetic pickup element. Note here that electronic circuit stays in or out of substrate. To determine concentration of target objects in fluid, fluid medium containing magnetic labels is subjected to magnetic field to differentiate between bound and unbound magnetic labels for period when binding process occurs on sensitive surface.

EFFECT: higher accuracy and rate of determining concentration of target molecules in fluid.

24 cl, 9 dwg

 

The present invention relates to a measuring device and system for determining the concentration of at least one type of target objects in a fluid containing at least one type of polarizable or polarized magnetic labels, and the system comprises a measuring device. The present invention additionally relates to a method for determining the concentration of at least one type of polarizable or polarized magnetic labels in the fluid using the measuring device.

In the field of diagnostics, especially in biometric diagnostics, for example medical and food diagnostics both in body and in vitro, and animal diagnostics, diagnostics, health and diseases, or for quality control widespread application of biosensing or biochips. These biopathic or biochips, in General, are used in the form of microarrays biochips, providing the possibility of testing the biological objects, such as, for example, DNA (deoxyribonucleic acid), RNA (ribonucleic acid), proteins, or small molecules, such as hormones or drugs. Today there are many types of tests used for the analysis of small amounts of biological objects or biological molecules or fragments of biological elements stored is, for example linking testing, competitive testing, offset, layer-by-layer testing or testing of diffusion. The challenge of biochemical testing is a small concentration of the target molecules (for example pmol·l-1and less)to be detected in the sample fluid at high concentration changeable background materials (for example, mmol·l-1). Targets can be biological elements, for example peptides, metabolites, hormones, proteins, nucleic acids, steroids, enzymes, antigens, haptens, drugs, components of cells or tissue elements. Background material or matrix may be urine, blood, serum, saliva or other derived from human or derived from human fluids or extracts. Labels attached to the target object, increase the limit of detection of the target object. Examples of tags include optical labels, colored granules, fluorescent chemical groups, enzymes, optical bar coding or magnetic labels.

Biopathic, as a rule, use the sensitive surface 1 with specific binding sites 2, equipped with capture molecules. These capture molecules can specifically bind with other molecules or molecular complexes, present and in the fluid. Other capture molecules 3 and label 4 simplify detection. This is illustrated in figure 1, which shows the sensitive surface 1 of the biosensor to which are attached the capture molecules, providing binding sites with 2 other biological objects, for example, the target molecules 6 or targets 6. In solution 5 contains the target object 6 and the label 4 to which are attached more capture molecules 3.

Targets 6 and the labels 4 are allowed to communicate with the connecting sections 2 sensitive surface 1 of the biosensor in a specific way, which is referred to herein as "specific attachment". However, other configurations of binding, which are referred to herein as "non-attachment", is also possible. On figa, 2b, 2C, 3.1a, 3.1b, a, 3.2b, 3.2, 3.3 illustrates some examples of possible combinations of binding labels 4 with sensitive surface 1 of the biosensor. Figa and 2b represent the so-called combination binding of type 1, which implement the desired biological attachment. On figa shows the desired biological attachment, in which the target molecule 6 is placed between the binding site 2 on the sensitive surface 1 of the biosensor and the capture molecule 3, available on the label 4 (layer-by-layer testing). On fig.2b shown case, the AI biosensor with competitive testing, in which binding sites 2, provided on the sensor surface 1, allow you to bind as a label 4 (by attaching connecting sections 2 to capture molecules 3, labeled 4)and the target object 6. Targets 6 are at least partially and in relation to the connecting sections 2 form and/or behavior similar to the capture molecules 3, so that there was a competition for binding sites between 2 capture molecules 3 (i.e. labels 4) and targets 6. Figure 2 C shows the case of a biosensor testing of inhibition, when the binding sites 2 biologically similar targets 6 and when the labels 4 are associated with the capture molecules 3 (or, in General, to the biological elements)that are associated either with the target object 6, or the binding sites 2. In the ideal case, the target object 6, is associated (via the capture molecules 3) with the mark 4 can no longer be contacted with the connecting surface 2 (the plot).

In the drawings, the bioactive elements (objects) (for example, the capture molecules 3 or binding sites 2) in General represented as directly attached to the solid carrier (for example, sensitive surface 1 or mark 4). As is known in the art, these bioactive layers, in General, are associated with solid novtel the m through intermediate objects, for example, molecules of the buffer layer or the separation of molecules. These intermediate objects are added in order to achieve high density and high biological activity of molecules on the surface. For simplicity and brevity of the intermediate objects are omitted in the drawings.

In contrast, biological attachment to the sensitive surface 1, the label 4 can also bind to the sensing surface 1 non-specific or non-biological way, i.e. the contact surface 1 without the mediation of specific target molecules 6. Figa, 3.1b, a, 3.2b, 3.2s, 3.3 represent this non-attachment, while Figo and 3.1b show the examples of so-called configuration link type 2, when one nonspecific relationship there is between the capture molecules 3, attached to the label 4, and sensitive biosensor surface 1, and/or between the capture molecule 3, attached to the label 4, and a connecting area 2 attached to the sensitive surface 1 of the biosensor. This is usually linking type 2 by only one non-specific communication is weak and can be broken by simple procedures, such as washing or magnetic forces. As shown in figa, 3.2b and 3.2s, the so-called binding configuration type 3 with the sensing surface 1 and/or binding is the missing section 2 Pets with a variety of nonspecific relations in a large area between the marks 4 (or capture molecule 3, attached to labels 4), on the one hand, and sensitive biosensor surface 1 and/or binding sites 2, on the other hand. Configuration type 3 usually provide greater binding force than the connection type 1. Fig illustrates a degenerate version of the type 1, where the label 4 is connected with the sensing surface 1 of the biosensor through specific and nonspecific relations.

When testing is necessary, for example, verification of saliva on the sidelines through the window on drug abuse, for example, for road safety, it is very important to provide equipment testing, reliable enough to use it in everyday work, as well as provide a way to test that provides results that are quite fast and accurate. Such testing can be performed in several formats, for example, in the competitive format of the test or test of inhibition. Figure 4 shows the evolution in time-dependent target signal S1and S2sensors for two different test samples, where the signal S1meets the high concentration of the target object 6, and the signal S2corresponds to the low concentration of the target object 6. Differences S1in comparison with S2due to the fact that the who the who the concentration of target molecules in a test sample, the higher the probability that the label 4, bound to the capture molecules 3, contact with the connecting sections 2 sensitive surface 1.

In international patent publication WO 03/054566 A1 disclosed magnetoresistive measuring device for determining the density of magnetic particles in the fluid. Magnetoresistive measuring device or the chip has a substrate with a layered structure that supports the fluid (the fluid). The layered structure has a first surface area on the first level and the second surface at the second level, and the magnetoresistive sensor element for detecting a magnetic field, at least one magnetic particle in a fluid environment. Magnetoresistive element is placed near the transition between the first and second areas of the surface and addressed to at least one of the areas of the surface. With this device it is possible to determine the concentration of labels 4 in a fluid environment.

The purpose of the present invention is to provide a measuring device, system and method, which allow to determine the concentration of at least one type of the target object in a fluid (the fluid)containing at least one kind of polarizable or polarized magnetic labels in a way that is fast enough and accurate is m, in particular, through the use of a concentration of at least one type of polarizable or polarized magnetic labels in a fluid environment and, in particular, by accurately measuring the speed of impact sensitive surface on the magnetic label, and concentration specifically attached magnetic marks on the sensitive surface.

The above objective is achieved by a measuring device, system and method according to the present invention.

In the first aspect of the present invention provides a measuring device for determining the concentration of at least one target object in a fluid medium containing at least one kind of polarizable or polarized magnetic labels. The measuring device includes at least one sensing surface, the sensing surface contains at least one type of binding sites that allow specific attachment of at least one biological objects associated with magnetic labels. The measuring device further comprises at least one element of the magnetic sensor, the measuring device further comprises a means of discernment to distinguish between the magnetic marks, specifically attached to the binding of the relevant areas, in comparison with the marks, the non-attached way, which is resolved in time.

The advantage of the device according to the invention is that it allows to determine the concentration of target molecules in a test using a magnetic biosensor more accurately and more quickly than previously. Specialists in this field of technology were very surprised and could not expect that you can increase the limit of detection and specificity using a measuring device according to the invention due to the accurate determination of the concentration of molecules directly on the surface of the sensor and the time when the binding process is performed on the sensitive surface.

The authors describe the invention for several different estimates. In the first example of the present invention is illustrated for the case of testing of inhibition. The sample with the target object 6 is subjected to the action of the reagent is labeled 4. The label 4 is equipped with capture molecules 3. In this case, these capture molecules 3 can be considered as a biological capture molecules 3, for example anticleia antibodies that can specifically contact with the target object 6. Due to their rapid kinetics targets 6 contact marks 4 through the capture molecules 3. Depending on the concentration of the target objects and is of waist binding of the capture molecules 3 (for example, constant Association and dissociation) of capture molecules 3 on the surface of the labels 4 are associated to a greater or lesser extent with the target object 6. The extent of coverage of the capture molecules 3 is represented by the parameter ε. In this test, the authors call this parameter fractional inhibition, and it ranges from 0% to 100%. If testing is regulated as a limited targets, i.e. when the test is sensitive mode, the parameter ε can be proportional to the concentration of target in the sample. Now the sensitive surface 1 is covered with binding sites 2, which in this case is similar to the target molecules, for example, the conjugates of drugs. Fluid labeled 4 is in contact with the sensitive surface 1. Magnetic label 4, which can move freely in solution, have the first opportunity to reach the sensitive surface, a second opportunity to enter in the biological contact with the sensitive surface and a third option to contact connecting areas 2 on the sensitive surface 1. The speed at which the labels 4 reach and contact with the sensitive surface, is called the speed of impact. The speed of impact does not depend, or depends to a small extent on the concentration of the target objects in a fluid environment. Atopology different from the speed of the link, which is largely dependent on the concentration of the target object in a fluid environment, for example, by means of the parameter ε. The speed of impact is always higher than the rate of binding and, as a rule, much higher speed link.

The speed of impact and link labels 4 with sensitive surface depends on many parameters. Some of the parameters are easy to control or configure to testing, and other parameters can vary greatly depending on the test conditions and properties of the sample fluid. For example, the area And the sensitive surface very precisely defined in the manufacturing process, for example, due masks and lithographic processing chip. Biological properties of the binding sites 2 and capture molecules 3 (for example, the surface density and biological activity, such as properties of the Association and dissociation) can be controlled and/or configured in advance in the process of biophotometer device or later. However, the speed of impact sensitive surface 1 by means of the labels 4 are difficult to control or configure, because it depends on many parameters such as the number of tags in the reagent, which came into contact with the sample, the rate of solvation of the reagent in the sample (note that the reagent can provide is given in the form of fluid or dry form), the viscosity of the sample, the sample temperature, the efficiency of mixing and/or agitation of the labels in a fluid environment (for example, through thermal diffusion, sedimentation, magnetic forces, acoustic forces, mechanical actuators, shear forces, rotational excitation).

In the above format of inhibiting binding of a target object 6 with labels 4 partially or completely slows associating labels with 4 binding sites 2, such targets. The rate of specific binding dN/dt labels 4 and the binding sites 2 on the sensitive surface 1 is defined by the following equation (unit-1):

where a is the area of the sensitive surface (unit m2), kon- this is a permanent Association of molecular binding (unit m3/[Cap] is the concentration of binding sites on the sensor surface (unit m-2), [L] is the concentration of the label 4 in a fluid environment (unit m-3), especially in solution, is near the sensor, and ε is the percentage of inhibition, which depends on the concentration of targets in the sample. Constant konthe Association depends on biological materials and other kinetic conditions (for example, temperature or forces, which, when estudia to labels in the binding process, for example, magnetic forces)that can be controlled and/or adjusted during or after the process biophotometer device or even to test with calibration fluid (for simplicity and clarity, the authors have neglected the process koffdissociation in the equation).

The purpose of testing is to accurately measure the concentration of the target objects in the original sample, which has a definite relationship with the parameter ε. It is therefore necessary to determine the parameter ε with high precision, i.e. with low Δε/ε. In light of the above equations it is important to determine all other parameters, i.e. dN/dt, A, kon, [Cap] and [L], with high accuracy. This is particularly difficult in the case of a low concentration targets: in this case ε is small, and the uncertainty in all the other parameters strongly affect the accuracy ε.

Therefore, it is necessary to fluid velocity impact sensitive surface on label 4 was accurately known. In this invention it is proposed to determine the rate of exposure by measuring the bulk density of magnetic particles which act as magnetic labels with very high accuracy. According to the present invention, the volume density of magnetic labels or granules ideally determined directly above the sensor and is measured at the time when the imp is under test. However, bulk density can also be measured in any other position or at another time, as long as the measurement is specific to the actual bulk density of about over binding sites. Therefore, the sensing surface 1 should be understood in the context of the present invention as the place where the measurement is specifically attached labels 4, i.e. binding sites 2 and running the measurement of bulk density magnetic labels 4 (i.e. non-specific labels attached 4) to determine the speed of the impact. Moreover, in measuring the speed of the impact, i.e. bulk density nonspecific attached magnetic labels 4, the term dimension "time resolution" should be understood as not requiring multiple measurements of bulk density labels during the time interval of sampling of the measurement signal (i.e. a signal indicating the magnetic label, specifically attached to sensitive surfaces).

The following is a second example of biological testing, namely competitive testing. Components of competitive testing are described on fig.2b. The rate of specific binding dN/dt labels 4 and the binding sites 2 on the sensitive surface 1 is defined through the your of equation (1), but now ε is the longitudinal occupancy of the binding sites 2 targets 6. As in the first example, quick and accurate data on the concentration of the target 6 in the fluid can be extracted by measuring the speed of impact sensitive surface on the magnetic label, and concentration specifically related magnetic marks on the sensitive surface.

In the third example shows the layer-by-layer testing. As in figure 1, the label 4 with capture molecules 3 are in contact with a sample containing the target object 6, and these materials come into contact with the connecting sections 2. The desired type of specific binding shown in figa. Note that the capture molecules 3 and binding sites 2, as a rule, are antibodies; as a rule, it is not the same molecules, but they are associated with different parts of the target object 6. The type of binding on figa can be performed in various sequences, for example, the target object 6 can first contact the labels 4, and then the binding sites 2, or Vice versa. For clarity of the present description it is assumed that the target object 6 first contact with labels 4. Depending on the concentration of the target objects and the properties of the binding of the capture molecules 3 (for example, the constants of Association and dissociation), mole capture the uly 3 on the surface of the labels 4 are associated to a greater or lesser extent with the target object 6. The extent of coverage of the capture molecules 3 target 6 is represented by the parameter ε. In this type of testing, the authors call this parameter fractional coverage. The rate dN/dt associating labels with 4 binding sites 2 on the sensitive surface 1 is defined by the following (unit-1):

where a is the area of the sensitive surface (unit m2), kon- this is a permanent Association of molecular binding (unit m3/[Cap] is the concentration of binding sites on the sensor surface (unit m-2), [L] is the concentration of the label 4 in a fluid environment (unit m-3), especially in solution, is near the sensor, and ε is the share of the coverage, which depends on the concentration of targets in the sample [note the difference with equation (1), which is a member of (1-ε)]. Constant konthe Association depends on biological materials and other kinetic conditions during testing (for example, temperature, magnetic forces)that can be controlled and/or configured to process biophotometer or directly before testing.

In the above example, the parameter ε is the share of the coating on the label 4. In the case when the sequential test, and they are the NGOs first make the target object 6 in contact with the connecting sections 2 and subsequent conversion of a sensitive surface 1 into contact with labels 4, the parameter ε corresponds to the longitudinal occupancy of the binding sites 2 targets 6.

The fourth example of a test that can be used is anticomplex test. This test uses the components of figure 1 with the specific property that the binding site 2 is chosen to bind to the capture molecules 3 in the presence of the target object 6, and does not capture molecules 3 one. This format is suitable for detection of small molecules, characterized in that the number of associated labels 4 increases with the number of targets 6. In sensitive test mode can be applied to equation (2).

The fifth example to demonstrate the use of this invention in testing, is testing using selective blocking agent. This format of testing in the future, also referred to as checked with a blocking agent. This type of testing, in addition to the components of figure 1, uses a blocking agent. A blocking agent, for example, may be similar to the target molecule attached to a larger object (element). In the absence of the target object 6 blocking agents are attached to the capture molecules 3, thereby blocking the binding of labels with 4 binding sites 2. When the target objects prisutstvie the Ute, they are partially or fully cover the capture molecules 3. Now the label 4 can contact connecting areas 2. This binding may include binding targets 6 (as figa), but this is optional. Linking can also be accomplished by the parts of the capture molecules 3, provided that this binding is not performed, when the blocking agent is associated with a capture molecule 3.

The number of associated labels 4 increases with the concentration of the target object 6 in the sample fluid. In sensitive test mode can be applied to equation (2). This format is suitable for large and small molecules. Small molecules can be, for example, zloupotreblenie drugs.

In biological testing reagents can be connected simultaneously (for example, in the well plate microtitration) or may come into contact sequentially in time (for example, in the hole, using the stages of sequential pipetting or device spreading of liquid drops in the radial direction). For example, you can first lead targets 6 and the capture molecules 3 in the contact, and then to bring the material into contact with the blocking agents. To speed up the materials can be brought in contact at the same time. The disadvantage of the first may be that the locking agents associated with capture molecules 3 before as the target object 6 can be contacted with the capture molecules 3, thereby reducing the fraction ε of the coating. This reduces the speed of the link labels with 4 binding sites 2. However, in the case of rapid molecular kinetics of the target object 6 can be shifted blocking agents associated with capture molecules 3, so that the proportion ε of the cover was addressed to a small extent.

The test examples described above demonstrate that the measured speed link labels 4 with sensitive surface 1 depends on the concentration of the target object 6 in a fluid environment. The present invention claims that the concentration of the at least one target object 6 can more accurately and therefore more quickly be deduced from the measured rate of specific binding of labels with 4 binding sites 2 through additional measurements of velocity of impact marks 4 binding sites 2.

This is illustrated by the kinetic equations involving fraction of the parameter ε, which is related to the concentration of target objects 6.

In a preferred embodiment, the speed of impact is determined by measuring the concentration of labels 4 near the surface 1.

In a preferred method, the concentration of the target object 6 is determined by calculating the relationship of the speed of binding with the speed of impact. More preferably, the concentration of the target object 6 is determined by calculating the ratio of the measured rate of specific binding with the measured concentration of labels 4 near the binding sites 2.

In General, the measuring device must be sensitive to labels, specifically attached to a sensitive surface (binding of type 1, see above), as well as to labels, which are nonspecific attached, but are located near sensitive surface. This second alternative can be implemented either by associating labels with sensitive surface by way of type 2, or by labels, not tied to sensitive surfaces, but placed near the surface.

According to the present invention, these various concentrations labels are measured independently. According to one variant of the invention, for example, can be distinguished specifically attached magnetic labels from other labels by the difference of rotational and/or translational mobility specifically attached labels and nonspecific attached labels and labels that are not attached. For example, you can apply a magnetic field to determine dependent mobility signals. These magnetic fields can also be modulated, for example, means that onezumi wires or magnets, to attract the magnetic label to sensitive surfaces or repel magnetic labels from sensitive surfaces, or to move the magnetic label on sensitive surfaces. The comparison of the signal of the magnetic sensor element for various positions of the magnetic labels provides the possibility of determining the number of mobile magnetic labels near sensitive surface, which are present in the measured solution.

In a preferred embodiment of the present invention the tool of discernment contains the means of forming a magnetic field for forming a magnetic field. Means forming a magnetic field can be placed on the measuring device, or may be, for example, the current-carrying wire or a two-dimensional conductive structure. Means forming a magnetic field can generate a rotating magnetic field. In another embodiment of the invention, means forming a magnetic field can generate a unidirectional or one-dimensional magnetic field, for example a pulsed unidirectional magnetic field or sinusoidal modulated magnetic field. In this case, different translational or rotational mobility of magnetic labels, different associated with sensitive surface, may be correlated with the different MSE of the awn migration of the first group of magnetic labels in a certain direction through the fluid, for example, liquid or gas, or may be correlated with the different speeds of rotation of the group of magnetic labels. Thus, different groups of magnetic labels may vary or be detected by the measuring device according to the invention.

In an additional preferred embodiment of the invention a means of distinguishing the measuring device includes means forming two magnetic fields, placed on each side of one element of the magnetic sensor, i.e. left and right or above and below. Alternatively, the sensor element is placed between two current-carrying lines, for example parallel current-carrying layers. The advantage of this type of embodiments of the present invention is that the magnetic sensor element is partially or completely insensitive to the magnetic field forming two magnetic fields, provided that the two magnetic fields cancel each other out to some extent in the position of the sensor element. Therefore, the magnetic sensor element, in fact, feels the magnetic field due to the presence of magnetic marks on the sensitive surface or near sensitive surface. By placing the magnetic sensor element in the volume, where the net magnetic field to which the sensor is compensated by means of the formation of two magnetic fields, the possible saturation of the sensor element is prevented. This is particularly important in the sensitive direction of the sensor, i.e. in-plane components of the magnetic field.

In an additional preferred embodiment of the present invention, the means forming the magnetic field is a two - dimensional wire structure placed on the measuring device.

As described above, the measuring device must be sensitive to labels, specifically attached to a sensitive surface (binding type 1), as well as to labels, which are nonspecific attached, but are located near the surface, for example the linking of labels of type 2, or labels that are not bound to sensitive surfaces, but located near sensitive surface. According to the present invention, these various concentrations labels are measured independently. According to an additional variant of the invention, the measuring device comprises a means of discernment, having a first surface area on the first level and the second surface at the second level, the element of the magnetic sensor is placed near the transition between the first and second region and facing the at least one area of the surface. In this embodiment, the measuring device prepact the tion, to the element of the magnetic sensor was centered around the transition between the first and second level when viewing almost perpendicular to the projection.

The advantage of the measuring device according to the second variant embodiment of the invention is that the concentration of magnetic labels near sensitive surface is available only through the change of the geometrical shape of the sensitive surface, respectively, without requiring the use of continuous or modulated magnetic fields. Through this, additional parameters associating labels with sensitive surface more easily available or temporal resolution of these measurements is increased.

In an additional preferred embodiment of the present invention a means of distinguishing the measuring device includes a tool capacitive sensor. One preferred method of measurement [L] according to the invention is a capacitive detection, i.e. the measurement range of impedance through the signal is removed, which is sensitive to the concentration of label in solution. For this purpose, the tool of discernment contains the capacitive means of perception. The capacitive means of perception may be provided by means of two electrodes, for example plates or wires of the capacitor, is whether over the sensitive surface or near sensitive surface. The capacitor plates can be provided as metallized areas on or near sensitive surface. Alternatively, the capacitor plates may be provided in the form of regions of semiconductor material such as silicon, polysilicon or any other appropriate material. The capacitor plates can be placed almost parallel to the substrate plane of the measuring device. The capacitor plates can be placed almost opposite each other in the direction perpendicular to the substrate plane. This gives the advantage that a significant volume of the sample is covered or accounted for capacitive measurement [L]. Alternatively, the capacitor plates can be placed almost opposite each other in a direction parallel to the substrate plane. This gives the advantage that the capacitor plates can be made on virtually the same plane as the sensing surface 1, which reduces the complexity of the manufacturing process of the measuring device.

In yet another additional embodiment of the present invention the above-described embodiments of the present invention can also be combined in that the means of discernment contains the means of forming a magnetic field for forming magnetodipole, in the same way as the first surface area on the first level and the second surface at the second level, the element of the magnetic sensor is placed near the transition between the first and second areas of the surface and addressed to at least one of the first and second areas of the surface.

The advantage of the measuring device according to the third variant embodiment of the invention is that the concentration of magnetic labels near sensitive surface even more accessible because you can combine different principles of measurement of the first and second variant implementation of the present invention to improve the temporal resolution and/or accuracy of the measuring device.

For all embodiments of the measuring device element of the magnetic sensor can be one of the element AMR, GMR or TMR sensor. Of course, the elements of the magnetic sensor on the basis of other principles, such as items Hall sensors or SQUID, is also valid according to the present invention.

Further, the present invention is mainly described with reference to a magnetic label, also called magnetic granules or pellets. Magnetic labels are not required to have a spherical shape and can have any appropriate shape, for example the form of spheres, cylinders or erased is it cubes, ovals, etc. or may be uncertain or volatile form. The term "magnetic labels should be understood that the label include any suitable form one magnetic particle or more magnetic particles, for example magnetic, diamagnetic, paramagnetic, superparamagnetic, ferromagnets, i.e. any form of magnetism, which generates a magnetic dipole in a magnetic field, permanently or temporarily. For the implementation of the present invention there is no limitation on the shape of the magnetic labels, but spherical labels at the present time the most simple and inexpensive to manufacture reliable way. The size of the magnetic label in itself is not the limiting factor of the present invention. However, for the detection of interactions on the magnetic biosensor label small size are predominant. When using magnetic granules micron size as magnetic labels, they limit the decrease in the scale, because each label occupies an area of at least 1 μm2. Moreover, a small magnetic labels have the best properties of diffusion and, in General, show less tendency to sedimentation (deposition)than large magnetic granules. According to the present invention, the magnetic labels are used in a range of sizes from 1 to 3000 nm, more pre is respectfully, from 5 to 500 nm.

In the present description and the claims, the term "biological objects" should be interpreted in a broad sense. It includes bioactive molecules, such as proteins, peptides, RNA, DNA, lipids, phospholipids, carbohydrates, such as sugar, etc. the Term "biological objects" also includes fragments of cells, such as parts of cell membranes, in particular parts of the cell membranes, which may contain the receptor. The term "biological objects" also applies to small compounds that can potentially bind to the biological object. Examples are hormones, drugs, ligands, antagonists, inhibitors, and modulators. Biological objects can be isolated or synthesized molecules. Synthesized molecules can include unnaturally occurring compounds such as modified amino acids or nucleotides. Biological objects can also occur in nutritional or fluid, such as blood or serum, or saliva, or other fluids or secretions in the body, or extracts or tissue samples from cellular structures, or any other sample containing the biological objects, such as food, feed, and water samples, etc.

The present invention also includes a system for determining the concentration, what about the least one type of the target object in a fluid medium containing at least one type of polarizable or polarized magnetic labels, and the system comprises a magnetoresistive measuring device according to any of the above embodiments. The system includes a measuring device along with appropriate mechanical environment, such as packaging, chambers, channels, pipes, etc. for sampling, pre-treatment of samples, moisture sensitive surface, etc. the System further comprises a measuring device along with suitable electrical and/or electronic medium, such as a power source, a means of data collection and analysis tool output.

The present invention also includes a method of determining the concentration of at least one type of the target object in a fluid medium containing at least one type of polarizable or polarized magnetic labels, using the measuring device according to any of the above embodiments, the method includes the steps:

- providing a fluid containing magnetic labels, over the sensitive surface,

- application of a magnetic field,

- differentiate with a time resolution between the magnetic marks, specifically attached to the binding in which asdam, and labels that are not bound and/or nonspecific attached.

According to the invention, in particular, it is preferable to determine the concentration of the target object by calculating the ratio of the concentration of labels, specifically attached to concentrations not bound labels, i.e., the ratio of the speed of binding (represented by the concentration of magnetic marks on the sensitive surface) to the speed of impact (represented by the concentration of the magnetic marks in the bulk liquid). The concentration of the target objects according to the invention is proportional to the parameter ε, the parameter that represents the fraction of occupancy of the binding components, which are available on the label 4 or sensitive surfaces 1, depending on the type of test. This parameter is associated with the concentration of target objects in a fluid environment in a way that depends on the test.

The Central idea of the present invention is to measure the concentration of the targets of the two dimensions, namely (i) the rate of specific binding of labels with binding sites and (ii) the speed of impact marks on the binding sites. The speed of impact is preferably measured by the concentration of unbound labels near sensitive surface, ie [L]. Provided by the various the s measurements [L]. One way to measure [L] is the measurement signal, which is specific for labels with high mobility. Another way to define [L] is the comparison of the sensor signals for two different situations, namely the situation when unbound labels are in the sensitive area of the sensor, and the situation when the labels are removed from the sensitive zone of the sensor, for example, are removed by magnetic forces, thermal diffusion, fluid flow, or other transport mechanisms.

These and other features, characteristics and advantages of the present invention should become apparent from the subsequent detailed description, taken together with the accompanying drawings, which illustrate as an example the principles of the invention. The description is only an example, without limiting the scope of the invention. Reference position below refers to the accompanying drawings.

Figure 1 illustrates the biosensor to which are attached the first capture molecules in the solution containing the target objects and labels, joined by the second capture molecule.

Figa, 2b, 2C, 3.1a, 3.1b, 3.2a, 3.2b, 3.2s, 3.3 illustrate some examples of possible configurations of binding labels 4 with sensitive biosensor surface.

Figure 4 illustrates the development over time of signaldata for two different test samples with a high concentration of the target objects and the low concentration of target objects.

Figure 5 illustrates a schematic representation of the system and the measuring device according to the present invention.

6 illustrates a schematic representation of the device according to the first variant implementation of the present invention.

7 illustrates a schematic representation of the device according to the second variant of implementation of the present invention.

Fig illustrates a schematic representation of the device according to the third variant of implementation of the present invention.

The present invention is described in relation to particular embodiments and with reference to certain drawings, but the present invention is not limited, but only by the claims. Described drawings are only schematic and non-limiting. In the drawings the size of some elements can be increased and is not illustrated to scale for illustrative purposes.

Moreover, the terms "first", "second", "third", etc. in the description and the claims are used for distinguishing between similar elements and not necessarily for describing a sequential or chronological order. It should be understood that in this way the terms used are interchangeable under appropriate circumstances and embodiments of the invention described in this d is the document, allow operation in sequences other than those described or illustrated in this document.

In addition, the terms "upper", "lower", "above", "below", etc. in the description and the claims are used for descriptive purposes and not necessarily for describing relative positions. It should be understood that in this way the terms used are interchangeable under appropriate circumstances and embodiments of the invention described in this document allow the orientations than described or illustrated in this document.

It should be noted that the term "comprising"used in the present description and the claims should not be interpreted as limited to the means listed thereafter; it does not exclude other elements or steps. Thus, the scope of the expression "a device containing means a and b"should not be limited to devices consisting only of components a and B. This means that in relation to the present invention, the only relevant components of the device are a and B.

Figure 1-3, already described in the introductory part of the description.

Figure 4 shows the evolution in time-dependent target signal S1and S2sensors for two different test samples. The intensity of the signal is La depends on the concentration of the target objects by the way, depending on the type of test. For example, when layer-by-layer test signal S1corresponds to the low concentration of target objects, and the signal S2corresponds to high concentration of target objects. The reverse applies to the example test inhibition or competitive test (i.e., the signal S1corresponds to a higher concentration of the target objects than the signal S2). For the interval tmtime, which corresponds to the time dimension, it is possible to measure the concentration of targets with sufficient accuracy. A few small circles in figure 4 represent the performance actually measured by the measuring device. The interval tmtime corresponds to the time until the result of the measuring device. At the beginning of the time interval measurement is time twadmission of fluid, particularly liquid, over the sensitive surface. Note that the drawing gives an example of a more or less linear behavior of the signal in time. In some cases, the signal may be more complex, for example a polynomial of higher order, due to, for example, time of activation of the biological layer or the time of diffusion or the drift time of the granules in the direction of the sensitive surface.

Magnetic biopathic, in General, made the so, to be sensitive to the granules, specifically related to the sensitive surface. However, the concentration measurement is specifically attached granules (or labels) to the surface may be disturbed due to the presence of unbound or non-specific associated granules (or tags). Therefore, reliable data point for measuring the concentration of target objects by concentrating labels preferably taken when unbound and/or non-specific bound granules are removed from the surface.

Accordingly, the following sequence or cycle can be applied one or more times:

- popping granules in the surface direction, whereby the binding can be made;

then the granules can push or move away from the sensitive surface, to make a distinction between specific binding to the surface and non-specific binding or unbound granules;

after this phase shift can be measured actual signal specifically associated granules.

Fig.9 illustrates the sensor, which is sensitive to labels associated with the surface, and to a certain extent sensitive to an unrelated granules near sensitive surface of the sensor. The signal illustrated ka is a function of time, and how can be extracted slope of the curve of binding to the surface. In the context of the present invention is surface-sensitive signal is also called raw signal-dependent target signal S of the sensor (or S1, S2illustrated in figure 4). The above sequence or cycle is used to measure the tilt of the surface-sensitive signal, represented by the dashed line. The signal represented by the dotted line, identical to the dependent target signal S of the sensor. Therefore, the measured slope of this signal leads to the determination of the concentration of target in the sample fluid. The above sequence or cycle is also presented in Fig.9, where the reference position 210 denotes a step of providing a label near the surface or popping labels to the surface, and the reference position 220 indicates the phase marks removal or ejection marks on the surface. The reference position 230 denotes an interval of one sample or event "unit of measurement" in the form of small circles in figure 4. For time tmmeasurement (see figure 4) is required to collect a certain number of these sampling intervals.

Signal for phase denoted by the reference position 210, arises from Serrano, communicating with the sensing surface, as well as by unrelated granules near sensitive surface. Using signals in the process steps indicated by reference positions 210 and 220, the signal binding surface, and the signal due to unbound granules can be extracted. As a result, the concentration of label in the solution, and the concentration of labels associated with the sensing surface can be extracted. According to the present invention, these two dimensions lead to a very accurate determination of the concentration of the target object in a fluid environment.

The slope of the curve is proportional to the speed of associating labels with sensitive surface. The average slope dS/dt signal during time tmmeasurement is set by the signal S (at the end of tm)divided by the time tmthe measurements. The concentration of the target objects associated with the rate of binding in a way that depends on the test. The concentration of the target objects can be very accurately determined when the signal is registered high signal-to-noise ratio. In case of detection by the magnetoresistive biosensor high signal-to-noise ratio can be achieved through the use of high currents. High currents can cause heating or permanently modify biomaterials. However, to the Yes signals measured at the end of the test, heating and modification of biomaterials is not important. In other words, the signal at the end point (i.e. specifically associated labels and/or unbound label in the solution near the binding sites) can be measured with very high signal-to-noise ratio, which increases the accuracy of determining the concentration of target objects.

Figure 5 shows the system 35 and the measuring device 10. The present invention provides the measuring device 10, such as, for example, the biosensor or biochip, especially suitable for use as matrix biosensing, i.e. many biosensing hosted on the same substrate material. The measuring device 10 is part of a system 35 according to the present invention. In a preferred embodiment, application of the measuring device 10 according to the present invention is used in a test kit for testing on the sidelines through the window, for the presence of drugs in the saliva, in order to road safety. As an example, this device is equipped for competitive testing (see fig.2b). The measuring device 10 includes a sensitive surface 1, which contains binding sites 2. Connecting the sections 2 are provided to specifically bind to the capture molecules 3 targets 6. Targets 6 is biological about jetty (for example, used drugs), and capture molecules 3 is a similar target molecules that are attached to labels 4. Objects 3 and 6 can contact areas 2, respectively, it is called the competitive format of the test. The device may also be equipped to test inhibition (see figs), but for simplicity, this section explains only the case of a competitive test. The measuring device 10 includes a substrate 20. Preferably, but not necessarily, the measuring device 10 includes means 13 forming the magnetic field. At least, if the tool 13 forming the magnetic field is not provided in the substrate 20 of the measuring device 10, the means 40 forming a magnetic field external to the measuring device 10, typically presents with system 35 according to the invention. System 35 further comprises a housing 21 forming at least a channel or chamber 22 and the like, to provide sufficient space for the fluid 5, mainly liquid containing such a target capture molecules 3 bound to the labels 4. Moreover, the fluid medium 5 contains the target object 6.

In another preferred embodiment, the device 5 is equipped for the format of the test inhibition (see figs). In this case, connecting the sections 2 are similar to the target molecule, attached to the sensitive surface 1. Targets 6 is a biological objects, such as zloupotreblenie drugs, etc. and capture molecules 3 is biological objects (for example, anticleia antibodies that can specifically bind to a target object 6 and similar targets binding sites 2. This is called format test inhibition, since binding targets 6 marks 4 partially or completely inhibits the binding of the label 4 with similar targets binding sites 2.

Of the two examples above it is obvious that the device may be equipped for a number of different formats of the test, for example a competitive test, test, inhibition, test bias, layer-by-layer testing. As is known in the art, biochemical, and chemical species (for example, target object, such as a target molecule, a label, binding sites) can be connected simultaneously or sequentially. To increase the speed mainly to combine the reagents at the same time. In the second case, the kinetics of the processes and the actual sequence of the binding process depends, for example, on the speed of diffusion and binding.

Note that the substrate of the sensor or chip may be any appropriate the mechanical bearing element made of organic or inorganic material, for example glass, plastic, silicon, or a combination. In a preferred embodiment, the measuring device 10 is an electronic circuit 30 is provided in the substrate 20. The electronic circuit 30 is provided in order to collect the signals or data collected or measured through item 11 of the magnetic sensor placed in the substrate 20. In an alternative embodiment of the present invention an electronic circuit 30 can also be placed outside of the substrate 20.

Figure 6 shows a schematic representation of a first variant implementation of the measuring element 10. On the substrate 20 is sensitive surface 1 and the element 11 of the magnetic sensor. Moreover, the means 13 forming a magnetic field is placed on the substrate 20 of the measuring element 10. The means 13 forming a magnetic field creates a magnetic field 130. If the external tool 40 forming a magnetic field (see figure 5) is present, the above-mentioned magnetic field 130 is a component of the resulting magnetic field created by the tool 30 of the formation of the magnetic field together with external means 40 forming a magnetic field.

Means 13 forming the magnetic field can be, for example, magnetic materials (rotating or non-rotating) and/or conductors, such as, for example, current-carrying wiring is Yes 13. In the described embodiment, the means 13 forming the magnetic field is preferably generated by current-carrying wires. Detection of rotational and/or translational motion of the labels 4 may preferably be performed by a magnetic field. In the first, and in subsequent versions of the implementation of the present invention, the magnetic detection may preferably be performed by using an integrated element 11 of the magnetic sensor. Can be used various types of items 11 sensors, such as, for example, a Hall sensor, a magneto-impedance, SQUID or any other appropriate magnetic sensor. Item 11 of the magnetic sensor preferably is provided as a magnetoresistive element, such as element 11 GMR - or TMR or AMR sensor. Means forming a rotating magnetic field can be provided by current-carrying wires, and means forming a current, integrated in the substrate 20 of the measuring device 10. Item 11 of the magnetic sensor can be, for example, the geometry of the oblong (long and narrow) strips. A rotating magnetic field is thereby applied to the magnetic marks 4 through current flowing in the integrated current-carrying wires. Preferably, the current-carrying wire can accommodate t is thus, they form a magnetic field in the volume where the magnetic label 4.

7 shows a schematic representation of a second variant implementation of the measuring element 10. On the substrate 20 is sensitive surface 1 and the element 11 of the magnetic sensor. Sensitive surface 1 contains as a means of distinguishing the first and second surface, marked together with the reference position 14. Separately, the first and second surface denoted by reference positions 141 and 142, respectively, and are located on the first and second levels, with the transition 143 between them.

On Fig shows a schematic view of the third variant of the implementation of the measuring element 10. On the substrate 20 is sensitive surface 1 and the element 11 of the magnetic sensor. Moreover, the first tool 131 forming the magnetic field and the second tool 132 forming a magnetic field are placed on the substrate 20 of the measuring element 10, together creating a magnetic field 130. In addition, the sensing surface 1 contains as an additional part of a means of differentiating the first and second surface, marked together with the reference position 14. On Fig can be seen that in the position of the element 11 of the magnetic sensor components of the magnetic fields generated by first and second means 131, 132 formed the project of the magnetic field, compensated, at least in part the result of the magnetic field for which the sensitive element 11 of the magnetic sensor.

The applied magnetic field 130 is such that it generates torque for labels 4. Thus, the labels 4 are rotated relative to another body (for example, different label, 4, sensitive surface 1 and so on) using the magnetic field 130. As stated above, the marks 4 contain magnetic material known in the art. The label 4 can be, for example, magnetic bead, magnetic particle, magnetic rod, a garland of magnetic particles or magnetic material within a nonmagnetic matrix. The parameter associated with rotational or movement freedom marks 4 can be detected by measuring device 10. The method according to the invention provides high-frequency measuring motor or rotational freedom. By measuring this type of distinction between specifically attached and nonspecific attached biological objects 3 possible, and through this discovery of different concentrations labels 4 connected in another way with the sensing surface 1.

Another possibility of defining different concentrations specifically attached in comparison with nonspecific attached bio is practical objects 3, is to provide such gradiententry configuration sensitive surface 1 working, at least with the first and second surfaces 141, 142. By providing this structure in a sensitive surface 1 can retrieve additional information associated with the concentration of magnetic labels 4. This is more explained in international patent publication WO 03/054566 contained in this description as a reference in respect of the following matters:

structure sensitive surface 1, at least with the first surface and the second surface in order to determine the volume density of magnetic labels 4 and/or the density of magnetic labels 4 according to the first, second, and third variant implementation,

- method of measuring bulk density and density of magnetic labels 4.

This invention is intended for use in immunological tests. Specialists in the art should be obvious that the testing of other targets and other binding objects can be used, for example, a test nucleic acids and objects hybridization.

Note that the above invention can be combined with the multiplexing of the sensors and/or multiplexing m is current. When the multiplexing of sensors sensors are used with different types of binding sites 2. In addition, the capture molecules 3 marks 4 can be of different types. When multiplexing labels using different types of labels 4, for example labels with different size or different magnetic properties.

1. The measuring device (10) for determining the concentration of at least one type of target object (6) in fluid (5)containing at least one kind of polarizable or polarized magnetic labels (4),
when this measuring device (10) comprises at least one sensitive surface (1),
while the sensing surface (1) contains, at least partially, at least one type of binding sites (2), allowing specific attachment of at least one kind of biological objects (3)associated with magnetic labels (4),
when this measuring device (10) further comprises at least one element (11) of the magnetic sensor,
when this measuring device (10) further comprises means (12) for distinguishing between the magnetic marks, specifically attached to the binding sites (2), and unbound magnetic labels (4), affecting the binding sites (2), during the time when the process of binding the tion is performed on sensitive surfaces.

2. The measuring device (10) according to claim 1, in which the impact of unbound labels on the binding sites (2) is set by the concentration of unbound magnetic labels (4) in a fluid environment in the immediate vicinity of the binding sites (2).

3. The measuring device (10) according to claim 1, in which the tool (12) distinguish contains a means (13) forming a magnetic field for forming a magnetic field (130).

4. The measuring device (10) according to claim 1, in which the tool (12) distinguish contains two means (131, 132) forming a magnetic field, placed on each side of one element (11) of the magnetic sensor.

5. The measuring device (10) according to claim 3 in which the means forming the magnetic field is a two - dimensional wire structure placed on the measuring device (10).

6. The measuring device (10) according to claim 3, in which the tool (13) forming a magnetic field generates a rotating magnetic field (130).

7. The measuring device (10) according to claim 3, in which the tool (13) forming a magnetic field generates a unidirectional magnetic field (130).

8. The measuring device (10) according to claim 1, in which the tool (12) distinguish contains the first region (141) surface on the first level and the second area (142) surface at the second level, while the element (11) of the magnetic sensor is placed adjacent to the area (143) is prehoda between the first and second regions (141, 142) surface and facing the at least one area (141, 142) of the surface.

9. The measuring device (10) of claim 8, in which the element (11) of the magnetic sensor is centered around the area (143) transition when viewed in a perpendicular projection.

10. The measuring device (10) according to claim 3, in which the tool (12) distinguish contains the means of the capacitive sensor.

11. The measuring device (10) according to claim 1, in which the element (11) of the magnetic sensor is a magnetoresistive element of the sensor element preferably AMR, GMR or TMR sensor.

12. The measuring device (10) according to claim 1, in which the magnetic labels (4) are provided as the magnetic granules.

13. The system (35) for determining the concentration of at least one type of target object (6) in fluid (5)containing at least one kind of polarizable or polarized magnetic labels (4), the system includes a measuring device (10) according to claim 1 and further comprises an electronic circuit (30) for detecting the change of the magnetoresistive effect element (11) of the magnetic sensor, and an electronic circuit (30) is present in the substrate (20) or outside of the substrate (20).

14. The system (35) in item 13, additionally containing an external tool (40) forming a magnetic field for forming a magnetic field.

15. The method of determining the concentration of, at the very measures which, one type of targets (6) in fluid (5)containing at least one kind of polarizable or polarized magnetic labels (4), using the measuring device (10) according to claim 1, the method contains the steps that
provide the fluid (5)containing at least one kind of magnetic labels (4)on top of the sensitive surface (1), applied magnetic field (130),
perform distinction between magnetic labels (4), specifically attached to the binding sites (2), and unbound magnetic labels (4) during the time when the binding process is performed on the sensitive surface.

16. The method according to item 15, in which the concentration of the target objects (6) is determined by calculating the ratio of the concentration of magnetic labels (4), specifically attached to the concentration of unbound magnetic labels (4).

17. The method according to item 15, in which the concentration of the target objects (6) is determined by calculating the ratio of the measured rate of specific binding of at least one type of magnetic labels (4) and the binding sites (2) to the measured speed of the magnetic labels (4) binding sites (2), preferably determine the rate of exposure by measuring the concentration of unbound magnetic labels (4) in the fluid near St. the binding sites (2).

18. The method according to item 15, in which the distinction between magnetic labels (4), specifically attached to the binding sites (2), and unbound magnetic labels (4) is performed using the difference between the rotational and/or translational mobility specifically attached magnetic labels (4) and unbound magnetic labels (4).

19. The method according to item 15, in which the distinction between magnetic labels (4), specifically attached to the binding sites (2), and unbound magnetic labels (4) is performed using at least one modulated magnetic field (130).

20. The method according to item 15, in which the specific attachment of the target object (6) to the binding sites (2) are obtained by testing in the format of inhibition.

21. The method according to item 15, in which the specific attachment of the target object (6) to the binding sites (2) get through the test in a competitive format.

22. The method according to item 15, in which the specific attachment of the target object (6) to the binding sites (2) are obtained by testing in layered format.

23. The method according to item 15, in which the specific attachment of the target object (6) to the binding sites (2) are obtained by testing in anticomplex format.

24. The method according to item 15, in which the specific attachment of targets (6) binding to the m sections (2) are obtained by testing in the format of a blocking agent.



 

Same patents:

FIELD: medicine.

SUBSTANCE: method of obtaining erythrocyte diagnosticum for reaction of indirect hemagglutination (RIHG) in case of cattle coccielosis consists of fractional formalinisation of sheep erythrocytes and their sensibilisation with coccielosis antigen at 70°C for 30 minutes. For sensibilisation used are erythrocytes, loaded with sensitin obtained from vaccine strain C.burnetii M-44, heated on water bath at 70°C for 30 minutes, with further triple washing of erythrocytic diagnosticum with buffer solution with pH -7.2.

EFFECT: invention ensures increase of specificity and activity of diagnosticum in RIHG in case of cattle coccielosis.

2 tbl

FIELD: medicine.

SUBSTANCE: claimed is method of obtaining reactive sugar which includes the following stages: i) providing sample, containing reducing sugar; ii) providing solid substrate covalently bound to linker, which includes catch group containing -NH2 group, in which said linker is optionally bound to said solid substrate via spacer; iii) reaction of said reducing sugar with said -NH2 group resulting in obtaining immobilised sugar; iv) reaction of free -NH2 groups with camping-agent, with camping-agent containing reactive group able to react with -NH2 group and v) reduction of C=N bonds with reducer resulting in obtaining reactive sugar of SugarCHn-NH- structure, bound with solid substrate via linker and, optionally, spacer, where n is 1 or 2 manipulation with immobilised carbohydrates by derivation.

EFFECT: improvement of method

45 cl, 2 tbl, 7 ex, 25 dwg

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention concerns development of a diagnostic test system in immunochip format and a method of simultaneous and differential detection of reaginic antibodies and antibody spectrum to diagnostically significant mmunologically relevant proteins Treponema pallidum of G (IgG) and M (IgM) classes. The diagnostic test system in immunochip format for differential serum diagnostics of syphilis consists of an immunosorbent with separately immobilised antigens Treponema pallidum Tp15, Tp17, TmpA, Tp47, conjugate and reactants required to detect an antigen-antibody complex. Cardiolipin is additionally immobilised on the immunosorbent; antigens Treponema pallidum and cardiolipin are immobilised at least, in two repetitions, and conjugate consists of mixed antispecies human IgG antibodies and human IgM antibodies modified by two phosphors with different spectral characteristics. On the immunosorbent, there can be additionally immobilised at least, one antigen and/or peptide Treponema pallidum specified e.g. of Tp39, Tp41, Tp42, Tp44.5, Tp92, Tp0453 at least in two repetitions. Cardiolipin can represent an oxidised cardiolipin derivative bounded with protein. The differential serum syphilis diagnostic technique with using the diagnostic test system in immunochip format involves application of human IgM modified by two phosphors with different spectral characteristics. On the immunosorbent, there can be additionally immobilised at least one antigen and/or peptide Treponema pallidum, specified e.g. of Tp39, Tp41, Tp42, Tp44.5, Tp92, Tp0453 at least in two repetitions. Cardiolipin can represent an oxidised cardiolipin derivative bounded with protein. The differential serum syphilis diagnostic technique with using the diagnostic test system in immunochip format implying that a cultivation solution for check and test samples is introduced on the immunosorbent with separately immobilised antigens Treponema pallidum and cardiolipin; the check and test samples are introduced; the prepared mixture is incubated at temperature 20-42°C for 15-60 min to prepare the antigen-antibody complex; the immunosorbent is rinsed; then a conjugate solution of mixed human IgG antibodies and human IgM antibodies modified by two phosphors with different spectral characteristics is introduced on the immunosorbent; it is followed with incubation at temperature 20-42°C for 15-60 min; the immunosorbent is rinsed, dried to detect the prepared antigen-antibody complex.

EFFECT: improved diagnostic accuracy.

20 cl, 5 dwg, 2 ex

FIELD: chemistry; biochemistry.

SUBSTANCE: method for combined immunobiological analysis of cells using a biochip involves incubation of the biochip which contains immobilised antibodies, with suspension of cells, washing the biochip from non-bonded cells, determination of coexpression of antigens on the bonded cells. The obtained result is assessed by determining presence of bonded cells in the region of the stain of the biochip and bonding density of cells and interpretation of the obtained result. Coexpression of antigens on cells bonded to the biochip is determined by carrying out one or more immunocytochemical reactions. When reading out the result, morphological analysis of cells bonded to the biochip is also carried out and presence and character of colouring of cells and their components with the reaction product are determined.

EFFECT: use of the disclosed method provides high reliability and information content of analysis.

9 cl, 6 dwg, 2 ex

FIELD: medicine.

SUBSTANCE: after antigen-antibody complex is prepared in a reaction compartment wherein said antigen to be modified and antibody are bound, said reaction compartment is washed with using a sample solution. The invention allows detecting a signal reflecting quantity of said antigen to be modified at precision comparable when using a washing solution to this without using said solution. Herewith, the washing solution is not required to be supplied from the outside of a chip or to be ensured therein beforehand.

EFFECT: immunoassay is easily realised on the chip.

6 cl, 6 dwg, 1 tbl, 2 ex

FIELD: medicine.

SUBSTANCE: invention can be used for evaluating the antibody level in blood serum by enzyme-linked immunosorbent assay (ELISA) in diagnostics of the diseases caused by capsular forms of such agents, as Meningococcus, Pneumococcus, Streptococcus, Haemophilus, Neisseria, Salmonella, Klebsiella, Pseudomonas, and also for evaluating the postvaccinal immunity level.

EFFECT: covalent binding of polysaccharide to the protein pre-immobilised on the tray surface ensures reliable and simple antigen fixation that does not involve effect of foreign chemical groups on the analysis results that leads to simplification of the method for tray sensitisation in the ELISA reliability control.

1 dwg, 1 tbl, 3 ex

FIELD: instrument making.

SUBSTANCE: set of invention relates to devices to determine the level of analysed substances in biological fluids. Proposed system comprises (a) measuring device with housing, logical circuit arranged in said housing, visual display arranged on said housing and measuring system arranged therein. It comprises also (b) cartridge incorporating at least one test strip to analyse lateral flow. Note here that said test strip incorporates (i) lateral flow transport matrix, (ii) zone of analysing specific binding on transport matrix to take up fluid specimen to produce required reaction, and (iii) zone of general chemical analysis on transport matrix to take up fluid specimen to produce required reaction. Note also that sizes of cartridges are selected to allow arranging analysed biological fluid substances in measuring device so that measuring system in above described zones on test strip. Mind that transport matrix first and second segments are made from different materials and bound so that said segments does not allow the product formed on first segment to contribute into reaction on second segment. Proposed invention covers other versions of aforesaid device.

EFFECT: higher efficiency, accuracy and reliability.

162 cl, 27 dwg

FIELD: veterinary.

SUBSTANCE: claimed is test-system of immuno-enzyme analysis, which allows to determine antibodies to viruses of infectious rhinotracheitis (IRT), viral diarrhea-disease of mucous membranes (VD-DMM), parainfluenza viruses -3 (PIV-3), respiratory syncytial (RS) and adenoviral (AVI) infections of livestock. Serological examination of animals allows to detect zones of infection spreading and estimate post-vaccination immunity.

EFFECT: application of claimed test-system IEA will allow to carry out simultaneously epizootological monitoring of five important infections of livestock, retrospective diagnostics of respiratory infections, and estimation of immunity stress in animals resulting from application of vaccines, determination of level of colostral antibodies in young animals in the first weeks or days of life, estimation of therapeutic medicine quality.

10 tbl

FIELD: medicine.

SUBSTANCE: group of inventions refer to medical diagnostics and covers diagnostics of early myocardial infarction by direct determination of myoglobin from human myocardium. Said biosensor represents a disposable graphite electrode modified by colloidal gold and monoclonal antibodies to myoglobin from human myocardium. It is produced the way as follows. A three-contact graphite main electrode is covered with colloidal gold solution stabilised with didodecyl methylammonium bromide in chloroform. It is dried at room temperature. The electrode structure is modified by antibodies to myoglobin from human myocardium by covering the main graphite electrode to myoglobin from human myocardium modified with didodecyl methylammonium bromide and colloidal gold. Then it is dried, settled at +2°÷+6°C, rinsed with water, processed with a blocking buffer, dried, settled at room temperature and rinsed with water.

EFFECT: possibility to determine myoglobin in aqueous solutions.

3 cl, 2 dwg, 5 ex

FIELD: medicine.

SUBSTANCE: invention refers to medicine, immunology and biotechnology. Substance of the invention consists in developing the method to lower the content of thyroid hormone antibodies by blood passage through prepared original granulated magnetic agent with immobilised forms of thyroid hormones on the basis of polyacrylamide granules with magnetic properties.

EFFECT: improved effectiveness and reduced consumption of the agent as compared to a prototype, owing to higher specific activity and effective multiple regenerability, simplified manipulations with the agent and maintained suspension of granules thereof in sorption process ensured by constant magnetic field that improves the active sorption area, and reduced destruction of perfused blood corpuscles.

5 ex, 2 tbl

FIELD: physics.

SUBSTANCE: invention relates to a method of making a piece (2, 4) of a film (1) of magnetoelastic material having initial flexural rigidity when flexural rigidity is increased in the first direction. In accordance with this method, a piece (2, 4) is provided with at least one linear depression (5) in the first direction of the piece (2, 4). Also the piece (2, 4) is curved along at least one linear depression from one or more linear depressions (5) in order to obtain a piece (2, 4) with stable flexure in a direction across the said first direction, through which high flexural rigidity is ensured in the first direction of the piece (2, 4). The invention also discloses an absorbent product made using said method, a sensor, an absorbent structure and an absorbing article having said sensor.

EFFECT: invention enables formation of a piece of film having initial flexural rigidity when flexural rigidity is increased in the first direction.

15 cl, 4 dwg

FIELD: physics.

SUBSTANCE: essence of the method lies in evaluating stress-strain state of narrow-profile articles made from ferromagnetic steel, which involves generating a magnetising magnetic field in the material of the article and measuring coercive force on the excited section of the article. The monitored section of the article is mechanically loaded, a magnetic field is then generated in its loaded state and the coercive force is measured. The stress-strain state focus of the material is determined by comparing the coercive force values with and without the load. The area of the spot of the magnetising field is stretched along the narrow-profile surface of the article. The longitudinal-stretched magnetic field and the coercive force sensor are scanned along the narrow-profile surface of the article, and the coercive force in both cases is measured in like points of the article.

EFFECT: higher resolving power, accurate detection of the behaviour of the defect and its dimensions due to narrowing of the exciting magnetic field which causes concentration of field lines and using the said field to scan along the narrow-profile surface of the article.

1 dwg

FIELD: chemistry.

SUBSTANCE: disclosed method of analysing ferromagnetic particles in oil involves successive exposure of the analysed sample to a magnetic field in different directions. The said magnetic field is constant. An alternating magnetic field whose parametres depend on differential magnetic permeability averaged in the volume of the oil is additionally applied in a fixed direction. Concentration of particles is determined from the difference between these parametres measured for different directions of the constant magnetic field.

EFFECT: selective analysis of irregularly shaped particles formed only as a result of friction.

3 cl, 2 dwg

FIELD: instrument making.

SUBSTANCE: invention relates to oil-and-gas industry and can be used to control efficiency of cathode protection of underground pipeline against corrosion. Proposed device comprises ferroprobe with first compensating winding, analog converter, phase detector and low-frequency filter connected in series, first current-to-voltage converter, analog-to-digital converter and excitation generator. Note that it additionally comprises second compensating winding. Analog converter comprises additionally output device, time interval shaper, indicator, keyboard, second selective amplifier and current-to-voltage converter, digital-to-analog converter and controlled switch.

EFFECT: additional measurement channel to measure cathode protection current continuously.

1 dwg

FIELD: electrical engineering.

SUBSTANCE: invention relates to inspection technology, more specifically to non-destructive testing using electromagnetic methods, and can be used to determine steel grades of longitudinal-extended objects, for example bars, rods, pipes etc. The non-destructive method of testing longitudinal-extended objects involves stimulating an electromagnetic field in the test model, measurement of parametres which characterise electromagnetic properties of the material of the test model and processing measuring results by comparing them with limits of similar parametres of known steel grades. Parametres used, which characterise electromagnetic properties of material of the test model, are a value, inversely proportional to the speed of propagation of electromagnetic field, and degree of damping of electric magnetic field on a given section of the model. Measurement results can be processed by a programmable microprocessor.

EFFECT: invention allows for determining steel grades of longitudinal extended objects using a non-destructive method by selecting more informative parametres.

2 cl, 1 dwg

FIELD: physics, semiconductors.

SUBSTANCE: invention is related to the field of semiconductor equipment and electronics. Method for measurement of photoferromagnetic effect in magnetic semiconductors consists in measurement of electromotive force that occurs in the secondary winding of transformer, which is wound on adjacent section of core from magnetic semiconductor in the form of doubled ring. The primary winding of transformer represents two coils wound on nonadjacent sides of doubled ring symmetrically relative to plane of symmetry that separates core in two rings. As a result of unbalance, in sinusoidal signals of magnetic flows directed and same in value that penetrate the secondary winding that are serially and antiphase-connected to outlet of generator, due to illumination of strictly half of core on one side from mentioned plane of symmetry, in this winding electromotive force occurs that is proportional to variation of magnetic permeability under light effect.

EFFECT: provision of possibility to perform measurements of photoferromagnetic effect amplitude in more sensitive scales of metering instruments.

FIELD: magnetic measurements.

SUBSTANCE: method of thermal treatment of specimen for calibration and adjustment of magnetic control devices, in which steel is annealed at temperature of 580±10°C or 700±10°C, depending on required gradient of normal component of remanent magnetisation field, for 10 hours with time delay, intermediate cooling for 5 hours together with furnace down to temperature of 250±10°C and ageing with final cooling. Final cooling is also carried out together with furnace and is completed at the temperature of 20...50°C. Specimen for calibration and adjustment of magnetic control devices is made in the form of plate manufactured from thin-sheet low-alloyed steel of preset size. Specimen was subject to thermal treatment at preset gradient of normal component of remanent magnetisation field, and is made as square one with side "a" from steel of grades 08"ю", 08"кп" or 08"пс" with thickness that is equal to (0.002...0.008)·a, mm. Gradient of normal component of remanent magnetisation field of this specimen lies within the limits of (30...200)·102 A/m2, where a = 125...200 mm.

EFFECT: method of thermal treatment of specimen for calibration and adjustment of magnetic control devices and optimisation of parameters of this specimen that has been produced with the help of this method, allows to increase reliability of rolling monitoring and accuracy of magnetic control facilities, and also to reduce production costs.

3 cl, 1 dwg

FIELD: oil-producing industry; natural gas industry; production of the devices of the non-tactless magnetometric control over the state of the pipeline metal.

SUBSTANCE: the invention is pertaining to the oil-producing industry and the natural gas industry and may be used for control over the state of the pipeline metal. The technical result of the invention is the increased efficiency, operability and accuracy of the measurements. The device contains two blocks of the flux-gate magnetometers with three flux-gate sensors rigidly connected among themselves, the block of electronic switches, the indicating device, the control unit, the block of determination of the position, on which the first and second blocks of the flux-gate magnetometers are rigidly set, the block of the control and the data processing, the inductive sensors of the magnetic field. The inductive sensors are rigidly connected among themselves and are installed along a straight line being the prolongation of the radius of the pipeline duct. The shafts of sensitivity of the inductive sensors are in parallel to each other and are oppositely directed, arranged in the plane perpendicular to the pipeline, and are collinear to the rigidly connected to each other shafts of sensitivity of the second flux-gate sensors of each block of the magnetometers.

EFFECT: the invention ensures the increased efficiency, operability and accuracy of the measurements.

1 dwg

FIELD: measuring technique.

SUBSTANCE: method comprises recording current value in the coil of the crane electromagnet when it lifts a portion of scrap, measuring the mass of the scrap portion, and calculating the volumetric density of the scrap. The carbon concentration is determined by the procedure proposed.

EFFECT: enhanced efficiency.

1 dwg

FIELD: the invention refers to applied magneto-optics and may be used for controlling authenticity of denominations, valuable papers, falsifications etc if they have magnetic means of protection from falsifications.

SUBSTANCE: the arrangement has a body, a source of constant magnetic field, a source of light, a polarizer, a magneto-optical film, an analyzer, an ocular. The feature of the arrangement is that it is provided with a holder in the shape of a plate with a hollow; the magneto-optical film is fastened motionlessly on the holder and partly covers the hollow in it, and the source of the magnetic field is fulfilled in the shape of two magnets located in the body above the magneto-optical film and ensures the magnetic field with a flat component of the field for saturation of the field of the controlled article and its orthogonal component does not prevail on value the field of touching the domen walls in the magneto-optical film. Due to this possibility is provided for visual control of matching of the part of the design printed with using a magnetic paint on the surface of the article with the corresponding surface of the magneto-optical the arrangement for controlling authenticity along the whole surface of the controlled article in any direction without dependence of its linear dimensions.

EFFECT: provides possibility to control authenticity of an article.

2 dwg

FIELD: physics.

SUBSTANCE: apparatus has a laser which is connected to an optical fibre carrying the laser radiation. A waveguide installation placed in the analysed medium is freed from the shell in order to provide contact between radiation and nanoparticles suspended in the liquid. Radiation scattered on nanoparticles is carried by a non-homogeneous wave which exponentially decays from the lateral surface of the optical waveguide. The radiation is collected on a photodetector which has a signal pre-processing unit. A version of the apparatus has a mirror with an opening for fibre, inclined at an angle to the fibre passing through the opening.

EFFECT: high scattering efficiency.

7 cl, 6 dwg

Up!