Nano-biochip used for registration of proteins and albuminous complexes, way of its reception and way of registration of proteins and albuminous complexes with use of catheter microscopy
SUBSTANCE: invention concerns medicine, in particular to medical diagnostics. A nano-biochip for registration of proteins and albuminous complexes and a way of its reception is offered. As a biochip substrate a non-stratified material, for example polycarbonate or polystyrene with a relief grid, is used and then the surface is leveled by means of a die providing equal surface with the size of roughnesses not more than 1 mm; after that updating and immobilisation of the molecules-probes capable to cooperate with investigated proteins and albuminous complexes on a surface of the chip specifically is carried out.
EFFECT: way allows to raise reliability of the received biochip at the expense of exception of its stratification and to raise reliability of diagnostics at the expense of rising of registration sensitivity of specific proteins and albuminous complexes.
12 cl, 2 dwg, 1 ex
The technical field
The invention relates to medicine, in particular to medical diagnosis, and for the diagnosis of cancer by registering specific proteins and their complexes in the study of blood. The invention can be used for diagnosis of other somatic and infectious diseases. In addition, the invention relates to the technology of the production of chips used in scanning probe microscopy.
The main problem of medical proteomics, and early diagnosis is the low level of sensitivity, not more than 10-12M To solve the problem of increasing the sensitivity of the methods in proteomics and diagnostics it is necessary to use molecular detectors, allowing registration of proteins at the level of single molecules. Such detectors include detectors based on scanning probe microscopes, including atomic force microscopes (AFM). Atomic force detector includes a chip with smooth surface and itself atomic force microscope. Chip with modified surface roughness less than 1 nm, coated with a molecule probes capable of forming complexes with macromolecules partners, is nanobiosym (hereinafter - chip).
The main problem in the use of the probe, including atomic force detectors is La proteomics and diagnostics, is
1) in creating the technology of chips with a smooth surface, the size of the rough terrain which is much smaller than the proteins adsorbed on the surface of the chip, i.e. less than 1 nm;
2) in the creation of technology for bioloy biomolecules probes on the chip surface, which has a higher affinity to macromolecules partner that is harvested specifically from a solution of the analyte, resulting in improved sensitivity of the method based on scanning probe microscopy.
The level of technology
Known chips, ways of registration of macromolecules with their use and their manufacturing techniques, in which the macromolecule-probes, immobilizovana in cell hydrogel on a common substrate and form the matrix (U.S. Pat. USA 5552270 and 5552271).
A method of obtaining chips, according to which the cell chip is produced by polymerization of the compositions of the droplets deposited on the substrate using a micropipette (F.N.Rehman, M.Audeh, E.S.Abrams, P.W.Hammond, M.Kenney and T.C.Boles, Nucleic Acids Research, 1999, v.27, 15, p.649-655).
The disadvantages with respect to scanning probe microscopy known methods and used chips are low reproducibility of cell properties. Technically sophisticated manufacturing technology chips that do not provide sufficient homogeneity and reproducibility of the properties of molecules of the analyte, and most importantly, this education is Finance-type materials adsorbed gels, structures with rough terrain, exceeding the size of the analyzed macromolecules.
In addition, the known protein chip, the manufacturing method and the method of registration that uses this chip. The chip is placed on a solid substrate molecule (antibodies, antigens or fragments thereof), with the chip diagnostics by identifying biological macromolecules specific to infectious diseases (hepatitis b, C, syphilis and others (CN 1373365).
The disadvantage of this chip is uncontrolled surface topography and immobilization procedures.
Closest to the proposed chip, method of manufacture and registration of macromolecules with it are the solutions described in patent application No. 2003109969 from 08.04.2003, in particular, the chip on the basis of mica, where the macromolecules adsorbed on the surface of the chip. In this patent document has disclosed a method of registering macromolecules, in particular proteins, using scanning probe microscopy, and the chip on the basis of mica.
Mica is the most suitable substrate in the manufacture of chips for probe microscopy, in particular for atomic force microscope for visual registration of macromolecules with sizes of the order of nanometers (these include proteins, protein complexes, oligonucleotides etc), because when solivan and on its surface are formed long atomically-smooth pad, for analysis using scanning probe microscopy can be mobilitat macromolecules. The disadvantage of the chip based on mica is its ability to delamination during the procedure of incubation mica with immobilized molecules probes in solution, resulting in the destruction of the chip.
The claimed group of inventions is directed to overcoming the above disadvantages of the known solutions.
The technical result consists in increasing the reliability of the obtained chip due to the exclusion of its stratification and increase the reliability of diagnosis by increasing the sensitivity of reception of specific proteins and protein complexes.
The technical result is achieved due to the fact that the chip for the registration of proteins and protein complexes using scanning probe microscopy, made in the form of the substrate, the surface of which is modified and which is applied to the field of immobilized molecules probes capable of specifically interacting with the analyzed proteins and protein complexes, the substrate used nerastraivaisya material, such as polycarbonate or polystyrene, on the surface using a hot stamp relief grid to separate zones when applied markers and is eaten with a stamp, having a working surface of sizescolors mica made a flat surface with the size of the roughness less than 1 nm.
The chip has a surface of the substrate, chemically modified with silane to silanizing surfaces of which are immobilized molecule probes using cross-linkers or
the chip has a surface of the substrate subjected to UV irradiation and subsequent immobilization on the treated surface molecules probes using created photo cross-linkers.
Immobilized molecule probes are antibodies, antigens, aptamers, oligonucleotides, fragments, immobilized alone or in any combination.
A method of producing chip for the identification of proteins and protein complexes using scanning probe microscopy involves the use of a substrate on the modified surfaces of which are immobilized molecule probes that can specifically interact with the studied proteins and protein complexes, and which are covalently bound, a new method is that the blank is cut out from nerasseivayushchee material, such as polycarbonate or polystyrene, using the first stamp is applied to the surface of the workpiece relief the grid using the second stamp, the working surface of which represents the FDS is th svincolo mica, on the workpiece print an atomically smooth surface with the size of the roughness less than 1 nm, cut the finished workpiece on the chips, followed by modification of the surface of the substrate of the chip and the immobilization of the molecules probes.
The surface modification of the substrate of the chip is performed by silane or ultraviolet radiation and directly immobilizing on the surface of the chip molecule probes using created photo cross-linkers.
The method of registration of proteins and protein complexes using generouse probe microscopy is done by placing the chip in a biological medium containing the investigated proteins and protein complexes, the captured proteins and protein complexes due to specific interaction with immobilized to the substrate surface of the chip molecule probes and covalent crosslinking and registration of proteins and protein complexes after laundering field chip from non-specific proteins and protein complexes of the new method is the use of a chip according to claims 1-4.
As a scanning probe microscope using single-channel or multichannel atomic force microscope, scanning tunneling microscope or an optical near-field microscope.
Registration is carried out in a gaseous environment, in vacuum or in a liquid.
Description graphic Mat is rials
Below the claimed invention will be described in more detail with reference to graphic materials, in which figure 1 presents an image of the chip surface with immobilized molecules probes, in particular antibodies to CA-125 (A); figure 2 - image of a chip with complexes (antibody to CA-125/ SA-125) on its surface, the image obtained by AFM.
Disclosure of inventions
The authors concluded that overcoming the above disadvantages of the known technical solutions are possible through the use of the substrate nonlamellar material is specially treated polycarbonate or polystyrene.
In accordance with the assigned task of the invention is associated with the creation of the chip for a scanning probe microscope for registration of proteins and protein complexes on the basis of specially treated polycarbonate. In accordance with the task created chip for the identification of macromolecules using scanning probe microscopy, consists of a field of immobilized molecules located on the substrate, where the substrate used is polycarbonate or polystyrene. Polycarbonate plastic (polystyrene) is a polymer and nonlamellar material. For chip fabrication of polycarbonate (polystyrene) is used for the treatment of 3 basic operations.
1. Virtue the Xia procurement chip polycarbonate or polystyrene. On a polycarbonate substrate using a heated stamp is applied to the grid by contact of the heated metal stamp with a certain relief with polycarbonate surface of the workpiece.
2. To obtain an atomically smooth surface is spaced alignment according to claim 1 surface using another stamp, the working surface of which is svincolo (just before stamping) mica. Instead of mica may use silicon, diamond or sapphire, i.e. materials which during processing it is possible to obtain a roughness less than 1 nm. So get ready billet, which is then cut into chips (e.g., size 7×10 mm).
3. For immobilization of the molecular probes on the surface of polycarbonate chip it is modified.
Modification of such a chip to obtain analicia can be performed in two ways:
a) modification of the silane and the further use of cross-linkers for immobilization of probes;
b) processing by ultraviolet radiation and direct immobilization on the chip areas molecules probes using created photo cross-linker.
This approach provides a chip for a scanning probe microscope with a relief of about 1 nm, which is sufficient for the registration of proteins in proteomics and diagnostics.
BC is now used technology for bioloy biomolecules probes on the chip surface by modification of molecules partners Biotin for education communication Biotin-streptavidin (avidin), high Kd~10-15 M [http://meeting.biophysj.org/cgi/reprint/82/1/337/d.pdf]. This technology unsuitable for chips used in proteomics and diagnostics, where the solution of the analyte is not just one model molecule, and many types of molecules. This is because when immobilization of molecules of streptavidin (avidin) on the chip surface can be specific to only register biotinylated molecules partners, and not all proteins from a multicomponent mixture of the analyte, which form complexes with the immobilized molecules probes.
To overcome this drawback associated with low Kd is possible through the use of modifications of the cross-linkers molecules as probes, such as antibodies, antigens, aptamers, oligonucleotides, or fragments thereof using methods of chemical, electrochemical, photochemical, radiochemical processing of these molecules directly on the chip and their immobilization on the chip surface in order to conduct the reaction of covalent binding immobilized on the surface of the chip for a scanning probe microscope molecules probes with macromolecules partners. This leads to irreversible binding molecules probes with macromolecules partner complexes on the surface of the chip to the probe microscope.
The proposed technology for the analysis of proteins and their complexes with scanning probe mi is roscopy is
1) the creation of technology nerasseivayushchee chip for a scanning probe microscope with a surface that allows you to achieve the size of the rough terrain which is less than or, at least, does not exceed the amount of proteins adsorbed on the surface of the chip;
2) development of technologies for bioloy molecules probes on the chip surface, which has a higher affinity to macromolecules partners, which is achieved by modification of the molecules as probes directly on the chip and their immobilization on the chip surface cross-linkers through which is conducted the reaction of covalent binding immobilized on the surface of the chip for a scanning probe microscope molecules probes with macromolecules partners from a solution of the analyte, and
3) registration of the formed complexes on the surface of the chip using scanning probe microscopy.
Example 1. As an example accomplish the task was created chip, consisting of zones of molecules immobilized on a substrate of polycarbonate chip to a scanning atomic force microscope.
Figure 1 as an embodiment of the present invention obtained by using atomic force microscope image of the chip for antibodies to CA-125 : cancer marker (A), which shows a macromolecule antic is l, covalently immobilized on a substrate of polycarbonate, modified silane; figure 2 shows the position (IN) - SA macromolecules, concentrated from solution of the analyte on the surface of the chip and covalently grafted to covalently immobilized antibody CA-125 using the cross-linker. The images were taken in tapping mode of measurement. As can be seen, the sizes of the complexes (antibody to S/ S) exceeds the size of antibodies SA, which means that the chip on the polycarbonate allows you to register marker proteins of cancer.
Similar results were obtained on substrates made of polystyrene with playgrounds silicon, graphite and mica with fixed ends to reinforce its structure modified in the field of electric discharge.
With the chip from polycarbonate and mica with fixed ends were recorded proteins HCVcoreAg markers of the disease hepatitis, located in the analyte at a concentration of From 10-17M, which indicates an increase in sensitivity by more than 2 orders of magnitude compared to existing proteomic methods.
Similar results were obtained when the immobilized to the substrate macromolecules used antigens, aptamers, oligonucleotides, or fragments thereof in the form of a separate chip, and any combinations of these molecules is the form field.
When using nanopore - substrate for scanning probe microscopes with immobilized molecules of different types with different types of immobilized probes the proposed method allows the registration of various types of diseases Desk complexes probe/target simultaneously.
Use instead of 1-channel multi-channel, for example 24-50 channel atomic force microscope (AFM)allows you to raise the performance of the method probe microscopy in the corresponding number of times, for example 24-50 times, and when using AFM mode simultaneous measurements at many points nanopoly to increase sensitivity to zeptomole level in the measuring mode, in which at these points immobilized the same type of molecules.
As scanning probe microscopes in this way the registration of macromolecules and chips coated with a conductive coating can be used, in addition to atomic force and scanning tunneling microscopes.
When using immobilized fluorescent molecules and/or macromolecules, or the introduction of a special fluorescent labels in the immobilized molecules macromolecules to obtain fluorescence in the selected spectral range instead of the atomic force microscope can be used in near-field scanning is Opticheskie microscopes.
1. Nanobiosym for registration of proteins and protein complexes using scanning probe microscopy, made in the form of the substrate, the surface of which is modified and which is applied to the field of immobilized molecules probes capable of specifically interacting with the analyzed proteins and protein complexes, and which are covalently bound, wherein the substrate used nerastraivaisya material, such as polycarbonate or polystyrene, on the surface using a hot stamp relief grid, and then using a punch having a working surface of sizescolors mica, formed a flat surface with the size of the roughness less than 1 nm.
2. Nanobiosym according to claim 1, characterized in that surface of the substrate, chemically modified with silane to which using crosslinkers immobilized molecule probes.
3. Nanobiosym according to claim 1, characterized in that it has a surface of a substrate subjected to UV irradiation and subsequent immobilization on the treated surface molecules probes using photocrosslinker.
4. Nanobiosym according to claims 1, 2 or 3, characterized in that the immobilized molecule probes are antibodies, antigens, aptamers, oligonucleotides, fragments, immobi isawanya alone or in any combination.
5. The method of producing analicia for the identification of proteins and protein complexes using scanning probe microscopy, including the use of a substrate on the modified surfaces of which are immobilized molecule probes that can specifically interact with the studied proteins and protein complexes that can covalently knit, characterized in that the cut blank from nerasseivayushchee material, such as polycarbonate or polystyrene, using the first stamp is applied to the surface of the workpiece relief the grid using the second stamp, the working surface of which is svincolo mica, form a surface with the size of the roughness less than 1 nm, followed by modification of the surface of the substrate of the chip and the immobilization of the molecules probes.
6. The method according to claim 5, characterized in that the surface modification of the substrate of the chip is performed by the silane.
7. The method according to claim 5, characterized in that the surface modification of the substrate of the chip is conducted by ultraviolet radiation and directly immobilizing on the surface of the chip molecule probes using created photo cross-linkers.
8. The method of registration of proteins and protein complexes using scanning probe microscopy, which consists in the room nonbiocidal biological environment, containing the investigated proteins and protein complexes, specific fish proteins and protein complexes, covalent linking them with immobilizerturning on the surface of the substrate of the chip molecule probes and registration of proteins and protein complexes after laundering fields analicia from non-specific proteins and protein complexes, characterized in that as analicia use the chip according to claims 1-4.
9. The method according to claim 8, characterized in that as a scanning probe microscope using single-channel or multichannel atomic force microscope, scanning tunneling microscope or an optical near-field microscope.
10. The method according to claim 8 or 9, characterized in that the registration is carried out in a gaseous environment.
11. The method according to claim 8 or 9, characterized in that the registration is carried out in a vacuum.
12. The method according to claim 8 or 9, characterized in that the registration is carried out in a liquid.
FIELD: veterinary science.
SUBSTANCE: invention refers to veterinary microbiology and biotechnology can be used for development of specific diagnostic aids. According to the invention the method covers producing antigen erythrocytic colibacillosis diagnosticum by extracting an adhesive antigen from Escherichia cultures, centrifuging the extract, and separating the supernatant. Thereafter formalinised tanninised animal erythrocytes are sensitised with the produced antigen. Extraction is performed in culture fluid containing Escherichia cell culture with phosphate-carbamide buffer 1.8-2.0 M of pH 7.2-7.4 at temperature 40-45°C during 25-30 min. The culture fluid containing Escherichia cell culture and phosphate-carbamide buffer are taken in mass ratio 1:0.4-0.6 respectively. After extraction the supernatant is heated up at 65-68°C within 25-30 min.
EFFECT: method allows for high-quality end product due to improved sensitivity and specificity.
FIELD: veterinary science.
SUBSTANCE: invention refers to veterinary microbiology and biotechnology can be used for development of specific diagnostic aids. According to the invention the method covers producing antigen erythrocytic pasteurellosis diagnosticum by extracting pasteurellosis capsular antigen from Pasteurella cultures with sodium chloride brine, with centrifuging the extract and separating the supernatant exposed to sterilisation filtration and used for sensitisation of formalinised anionised animal erythrocytes. Pasteurellosis capsular antigen is extracted with 2.0-2.5% sodium chloride brine at 40-42°C during 30-40 min. Prior to sterilisation filtration, the supernatant is heated up at 65-70°C within 15-30 minutes.
EFFECT: application of the method allows for high-quality end product due to improved sensitivity and specificity.
SUBSTANCE: invention refers to medicine, namely to medical diagnostics. Offered is method of specific macromolecules record in biological sample by means of the device equipped with optical disk (laser CD) used as sensitive element with applied macromolecular probes specific to disease associated macromolecular markers. Detected macromolecules and their complexes are recorded on CD surface using laser CD reader.
EFFECT: simplification of record process, as well as in higher sensitivity of test system.
4 cl, 3 dwg
SUBSTANCE: invention concerns molecular biology and physics and can be applied in detection of analysed object in a medium. Photosensitive layer is formed on a substrate reflecting or diffusing light. This sensitive material is specific to the object to be detected, and can bind it. When the sensitive material covering the substrate is irradiated in the presence of a stencil, a biosensor with a pattern of active and inactive sensitive material areas is obtained.
EFFECT: evokes visible diffraction at the contact and light irradiation of a medium containing the analysed object.
23 cl, 5 dwg, 1 ex
FIELD: medicine, immunology, allergology.
SUBSTANCE: method involves determination of the level of allergen-specific antibodies - immunoglobulins of subclasses IgG1 and IgG4 in serum blood using the specific allergen Coefficient of specific allergizing (SAC) is calculated by the following formula:
SAC = (parent (IgG4 + IgG1) - control sample (G4C + G1C))/(parent (IgG4 + IgG1) - sample with allergen (G4A + G1A)) wherein IgG1 and IgG4 are the level of the parent indices of corresponding subclasses of immunoglobulins G; G1A and G4A are the levels of corresponding subclasses of immunoglobulins G in addition of allergen; GrC + G1C are the level of corresponding subclasses of immunoglobulins G without addition of allergen. The presence of IgG-dependent sensitizing to a specific industrial allergen is provide at SAC value less 2.5. Method provides enhancing diagnosis for the presence of sensitizing to allergen and to reduce time of analysis. Invention can be used for diagnosis of professional diseases.
EFFECT: improved method of diagnosis.
SUBSTANCE: method involves applying biochip test system for analyzing samples. The system has at least one biochip having at least one reactor having probe molecules immobilized inside it. The molecules have specificity to structure or type of ligands following given order set as probes matrix and/or probes arrangement circuit. The ligands show specific activity in antigen-antibody reaction or specific affinity to antigen and/or antibody and does not enter into reaction with antibody-probe and/or antigen-probe that results in reduced cross-reaction probability. Method involves analyzing biological samples with said test system.
EFFECT: high accuracy of test results.
10 cl, 4 tbl
FIELD: medicine; methods of diagnostics.
SUBSTANCE: device for revealing antigen-target in liquid sample has reaction chamber; immobilized antibody fixed inside reaction chamber; complex-reporter provided with probe and antigen of complex-reporter, where probe is connected with antigen of complex-reporter. Antigen of complex-reporter is connected with immobilized body. Antigen of complex-reporter is connected with antigen-target less tight than antigen-target. Device has revealing chamber, input for entering reaction chamber and passage for sample between reaction chamber and revealing chamber. When part of antigen of complex-reporter dissociates from immobilized body to liquid sample, antigen-target binds with immobilized body. When liquid sample is transferred to revealing chamber, amount of complex-reporter is found, which amount has to be indicator of quantity of antigen-target that quantity was presented in liquid sample initially. Described device has to be single-use immune sensor, which doesn't need to be washed through. There are no requirements to user concerning stages of measurement. Sensor can be easily adapted to antigen-antibody interactions within wide kinetic limits. Availability or absence of antigen in sample can found precisely.
EFFECT: improved precision of measurement.
42 cl, 4 dwg
FIELD: medicine, biology, agriculture.
SUBSTANCE: the present innovation deals with fulfilling sorption immunoenzymatic assay to detect small quantities of biomolecules, such as an antigen, an antibody, etc. The innovation, in particular, refers to heating-mediated immobilization of antigen or antibody onto activated surface followed by carrying out further ELISA stages at controlled temperature. The innovation shortens total time required for implementing ELISA by approximately up to 3 h. It is quick, economic, reproducible and simple technique.
EFFECT: higher efficiency.
24 cl, 14 ex, 14 tbl
FIELD: chemistry of polymers, biochemistry.
SUBSTANCE: invention relates to a rapid method for preparing photoreactive polymers and immobilization of biomolecules on indicated polymers. Invention describes a method for preparing photoreactive polymers followed by immobilization of molecules on them that involves the following steps: (a) effect of microwave radiation on mixture of photolinker and solvent; (b) washing out polymer with a corresponding solvent followed by drying at the environment temperature to obtain photoreactive polymer; (c) addition of biomolecule dissolved in corresponding buffer on photoreactive polymer; (d) treatment of mixture by the photoenergy source for period from 2 min to 2 h for immobilization of molecule on indicated polymer, and (e) washing out indicated polymer having immobilized molecule in corresponding buffer and the following drying indicated polymer at the environment temperature to obtain photoreactive polymer with molecules immobilized on its.
EFFECT: improved preparing method.
22 cl, 2 tbl, 18 ex
FIELD: medical engineering.
SUBSTANCE: device has protein chip as component of diagnosing system for determining multiple parameters. Method involves obtaining diagnostic system. Standard solutions sample set has sodium azide, fetal bull blood serum and antigens available in high concentrations. Reaction solution has two or more proteins, marked with luminescent label.
EFFECT: increased system stability and sensitivity.
3 cl, 2 dwg, 4 tbl
FIELD: medicine, immunology.
SUBSTANCE: method involves synthesis of magnetic latex in the presence of colloidal-dispersed ferromagnetic particles of iron oxide wherein iron oxide is added into microspheres of polyacroleinic sorbents after their synthesis. Method provides simplifying the technology in preparing the serological test-system.
EFFECT: improved preparing method.
SUBSTANCE: method involves applying ELISA. When doing it, antigen or antibody is immobilized on activated substrate. Microwave radiation is applied at frequency taken within the limits of 2300 and 2500 MHz at each stage from 5 to 200 s. Device has loading chamber, reaction chamber having magnetron, air-exhauster and unit for focusing light, chamber for washing and drying base plate or unit, detection chamber, platform for transferring base plates and computer means.
EFFECT: accelerated analysis method.
14 cl, 8 tbl
SUBSTANCE: method involves making interaction of biological sample with solid phase reagent covered with catching antibodies showing affinity to component A binding to produce the first reacted particle. Making the particle to come in contact with labeled antibodies showing specific binding affinity with respect to A and B components results in the second reacted particle production. Labels are measured on the second reacted particle by applying flowing cystometry method.
EFFECT: wide range of functional applications in early stage detection of infections like AIDS or hepatitis C.
19 cl, 2 dwg
FIELD: pharmaceutical chemistry.
SUBSTANCE: invention relates, in particular, to a composition for interaction of ligands wherein the composition comprises a noncovalent associate of multiple separate conjugates being each of that comprises a head group and a tail group wherein tail groups of conjugates form hydrophobic aggregate. Conjugates are mobile within the associate and in the presence of ligand at least two head groups are places by a method corresponding to the epitope formation that is able to interact with ligand stronger as compared with each separate head group. Invention provides applying conjugates in combinatory approach for detecting effective combinations to induce the desirable interaction in binding in receptor-specific treatment of patients.
EFFECT: valuable properties of epitopes.
31 cl, 2 dwg, 4 ex
FIELD: medicine, clinical laboratory diagnosis.
SUBSTANCE: method involves using carboxylated polystyrene latex spherical particles suspension with immobilized specific polyclonal antibodies raised against antithrombin III on their surface and wherein milk α-casein (antithrombin III-latest) is added as an inert component. Method for assay involves successive fractional plasma dilutions and the concentration of antithrombin III in the parent sample is calculated by the formula. Method provides rapid determination of antithrombin III concentration in blood plasma. Method can be used for diagnosis of pathological states in the blood coagulation system.
EFFECT: improved assay method.
2 tbl, 2 ex
FIELD: immunochemistry, sorbents.
SUBSTANCE: method for preparing immunosorbent involves modifying aerosil with chitosan solution in acetic acid in common with iron formate followed by oxidation and immobilization of the protein ligand. Invention can be used in diagnosis for detection of specific response antigen-antibody with using immunoenzyme analysis and immunofluorescence reaction.
EFFECT: improved preparing method.
1 tbl, 4 ex
FIELD: biotechnology, immunology, microbiology.
SUBSTANCE: freeze-dried 10-40 % cell slurries are treated singly or twice with 25-35 % iron sulfate solution. After repeat freeze drying treatment with 20-25 % ammonium aqueous solution is carried out. Then obtained magnetic cells are activated with 5 % glutaric aldehyde for 4 h. Further cells are treated with species-specific competent immunoglobulins from lemic, tularemia, or brucellosis serum containing 10 mg/ml of antibodies for 24±2 h. Method of present invention makes it possible to detect lemic bacteria, tularemia, and brucellosis in concentration of 103-104 m.c./ml. Obtained biomagnetic immunosorbents may be stored for 2 years at 4-8° without losses of specific activity.
EFFECT: biomagnetic immunosorbents of improved sensitivity and prolonged storage-time.
5 cl, 9 ex
SUBSTANCE: method involves determining complex formed between HCV anticore antigen and NS3/4a-antibody capable of recognizing both HCV- antigens and antibodies available in sample when using common solid base and solid substrates usable in immunoassay.
EFFECT: high accuracy and sensitivity of hepatitis C diagnosis.
47 cl, 8 dwg, 10 tbl
FIELD: drawing liquid samples for fast and simple qualitative and quantitative analytic analysis of liquid sample components.
SUBSTANCE: proposed device 1 is used for carrying the sample over capillary active passage 2 from point 3 of sampling to point 4 of receiving the sample; capillary active passage 2 is formed by backing 5, coat 6 and intermediate layer 7 located between backing and coat; intermediate layer 7 defines geometry of capillary active passage; backing 5 at sampling point 3 projects beyond coat 6 and intermediate layer 7 is shifted backward at this point in way of receiving point 4 and consequently backing 5 and coat 6 project beyond intermediate layer 7.
EFFECT: ease of drawing samples by means of auxiliary facilities and proportioning of liquid.
9 cl, 3 dwg
FIELD: medicinal microbiology.
SUBSTANCE: gist of invention resides in using polymer carrier prepared by a method of anionic polymerization of acryl aldehyde monomer in presence of sodium hydroxide as polymerization initiator, said polymer carrier being in the form of colored spherically-shaped equigranular particles 2±0.5 μm in diameter and having surface reactive aldehyde groups in quantity 1.3-1.6 μmol/g carrier, which carrier is washed with 0.1 M carbonate buffer solution having pH 9.0-9.2, centrifuged for 10 min at 3000 rpm, immobilized by suspending washed out carrier in 0.1 M carbonate buffer solution, pH 9.0-9.2, to which diphtheria antitoxic immunoglobuline is then added. Components are held in contact for 2 h at 20°C, cooled to 7°C, kept at this temperature for 16-18 h, and stabilized. Stabilization is effected by adding 1% gelatose solution in 0.9% sodium chloride solution, stirring for 2 h at 20°C, and lyophilization of diagnosticum. Diphtheria toxin is revealed using space agglomeration reaction.
EFFECT: increased specificity of diagnosticum.
3 cl, 3 ex