Method of immunoassay for analyte detection in sample

FIELD: medicine.

SUBSTANCE: ferric (III) hexacyanferrate (II) hydrosol conjugated with antibodies (antigens) to a required analyte is used as a disperse phase. The presence of the analyte is detected by observing a bright blue strip on an analytical membrane.

EFFECT: faster immunoassay procedure as compared to conventional ones, higher contrast between an analytical zone and a white surface of the membrane.

6 ex, 6 dwg

 

There are many examples of the use of immunochromatographic test-systems for detection of an analyte (ER 0890103 B1; EP 1046913 A2; US 50968375238; US 5238652...). Similar examples are also described in the book "Lateral flow immunoassay" R.C.Wong, H.Y. Tse, eds., Humana Press, N-Y, 2009. In these examples, as the dispersed phase for the conjugation of antibodies (antigens) using nanoparticles (diameter 5-80 nm) gold, colored or fluorescent polymer particles of submicron size (CA 2049919), nanoparticles of a semiconductor material capable of luminescence (US 2006/0008921).

The closest analogue (prototype) of the present invention described in the patent EP 0890103 B1 "Quantitative immunochromatographic assay". In the device, for conducting a lateral flow analysis described in the prototype, it is assumed that the membrane conjugate contains artificially introduced during micro-particles, such as colloidal particles of metals, molecules of organic compounds, liposomes, or organic polymer latex particles. These particles are associated with the detecting antibody capable of binding to the analyte.

In the process of lateral flow analysis of a liquid sample containing the analyte interacts with antibodies microparticles formed complexes of analyte and microparticles capillary fluid flow fond of analytical membrane to narrow and eliticism zone, which contains antibodies capture, stopping microparticles associated with the analyte, thus forming colored or luminescense complex.

The specified method of analysis in our view has at least two drawbacks.

I. Colloidal particles of metals, molecules of organic compounds and, especially, liposomes, or organic polymer latex particles roads in the manufacture, require complex methods of synthesis of the particles, time-consuming chemical methods of introducing functional groups to the surface for subsequent binding to antibodies (antigens).

II. The rate of migration of the analytical membrane complex of the analyte-microparticles and the rate of formation of the analytical signal is limited by convection and diffusion in the membrane [S.Qian, H.H.Bau. A mathematical model of lateral flow bioreactions applied to sandwich assays. / Analytical Biochemistry 2003, v.333, pp.89-98]. The diffusion coefficient in the membrane with identical physical properties depends on the hydrodynamic radius of the complex particles with the analyte and the viscosity of the eluting solvent. The typical time of formation of the analytical zone with the use of particles of colloidal gold average diameter of 35 nm in membranes with different rate of capillary flow (for example, Millipor HF120 and Millipor HF240), at room temperature, between 20% and 45 minutes

In addition, microparticles (who particularly particles of polymer latexes and liposomes) have the ability to aggregation, what causes the reduction in the rate of migration of the latter. About this problem and ways of its overcoming said patent CA 2049919.

When conducting chromatography front eluting solvent on the membrane ahead of the propagation front of the complex particles with the analyte, resulting in uneven movement of painted (luminescent) complexes and their aggregates on the analytical membrane, reducing the contrast between the particles on the membrane and the particles in the analytical area, thereby lowering the sensitivity analysis. We offer the dispersed phase Hydrosol of hexacyanoferrate (II) iron (III) does not have the mentioned disadvantages. Specified dispersed phase has the following advantages:

- The cost of raw materials for its production and the finished product is less expensive than colored or fluorescent polymer latexes, colloids of metals (gold), liposomes.

The process of synthesis of Hydrosol of hexacyanoferrate (II) iron (III) very simple, easy and well replicated.

- Has a well-developed surface and the ability to adsorptive binding of antibodies and antigens, such as protein and lipopolysaccharide character.

Movement of the complex particles with the analyte on the lateral flow membrane occurs uniformly, simultaneously with the front e is wirausaha solvent. Typical transit time of lateral flow process on the membranes Millipor HF120 is from 2 to 15 minutes, It was several times faster than for chromatography using other microparticles. Due to the uniform motion of the dispersed phase immunochromatographic membrane outside the analytic zone almost not colored, in the analytical zone of the formed high-contrast stripes in bright blue, making the analysis more sensitive.

The purpose of the invention is the detection of analytes in the sample for a shorter time and with higher sensitivity.

The technical result is to accelerate the procedure immunochromatographic analysis, in comparison with traditionally used as the dispersed phase of a colloidal solution of gold colored polymer latexes or quantum dots, are enhanced contrast between the analytical area and the white surface of the membrane.

The claimed technical result is achieved due to the fact that the method of conducting a lateral flow assay for detection of analytes in the sample, including the creation of multimembrane composite, consisting of membranes for application of the sample, membranes for conjugate, analytical membrane and membrane for absorption of the sample containing dry conjugate of the dispersed phase with the antibody (anti-Christ. Yong) to one of the antigenic epitopes of the analyte, applying immunoreagent (second antibodies to other antigenic epitope of the analyte or antigen) on the analytical membrane in the form of a narrow zone, conducting the immunochemical reaction and separation immunochemical complex at the same time during the chromatographic migration on the surface of the analytical membrane, characterized in that the dispersed phase to the first conjugation of the antibody (antigen) use the Hydrosol of hexacyanoferrate (II) iron (III).

The invention consists in that the lateral flow assay is used as the dispersed phase of the Hydrosol of hexacyanoferrate (II) iron (III), conjugate with antibodies (antigens) to the desired analyte. Spend immunochromatographic analysis, the presence of the analyte is judged by the appearance of the analytical membrane painted in bright blue stripes.

The method is as follows.

When implementing the method are conjugate particles Hydrosol of hexacyanoferrate (II) iron (III) (GCFI; another name for this connection - "Prussian blue") with antibodies to the analyte (antigen)or with antigen to the investigated analyte (antibodies), are conjugate in conjugate membrane with subsequent drying, put the second antibody to the antigenic epitope of the analyte or antigen on the analytical membrane, in the form in which some bands in the analytical area, form multimembrane composite, spend immunochemical reaction on the surface of the analytical membrane, with subsequent determination of the investigated analyte (antigen or antibody) by the appearance of a colored band in the analytical zone. The content of the analyte in the sample is proportional to the amount of the formed complexes conjugate particles Hydrosol GCFI with the analyte captured in the analytical zone. The Hydrosol particles GCFI painted in bright blue color, the intensity of the color stripes of the analytical zone is proportional to the amount of analyte in the sample.

EXAMPLES

Example 1. Obtaining conjugates Hydrosol GCFI with antibodies or antigens

Hydrosaline drugs GCFI get when mixing solutions of ferric chloride and ferric Sinanodonta potassium in the water.

For sensitization of the surface of the Hydrosol particles GCFI antibodies or antigens to the drug Hydrosol add an aqueous solution containing immune component, the mixture is incubated on a magnetic stirrer for 30 min at room temperature and add bovine serum albumin (BSA) to a final concentration of 0.1%. After 2 min incubation under the same conditions, the mixture is centrifuged at 50000-60000 xg for 30 minutes the Precipitate is suspended in a 0.1% solution of BSA. The suspension is again centrifuged under the same conditions. Sediment suspended the 0.1% solution of BSA. Determine the wavelength of the absorption maximum and the optical density (OD) at the maximum of the spectrophotometer. Using a solution consisting of 5% sucrose and 0.25% BSA prepared suspension Hydrosol with density (OD)equal to 3.0 at the maximum absorption, and put it on the membrane to conjugate. The specified membrane is dried for at least one day in a room with relative humidity from 20% to 30% and a temperature of + 20°C to plus 25°C.

Example 2. Creating multimembrane composite

The Assembly of the composite (figure 1) is carried out at relative humidity ranging from 20% to 30% at a temperature of from 20 to 25°C.

On a nitrocellulose membrane (3) in the area of analytical zone in the form of a narrow strip (4) is applied solutions of antibodies to antigenic epitopes of the analyte or antigen (as analyte antibody). In the area of the control zone (5) is applied individule polyclonal or monoclonal antibodies.

On the laminate substrate nitrocellulose membrane (6) paste dried conjugate membrane (2) with printed conjugate OP=3.0 and the membrane (1) for application of the sample. On the opposite side, on the laminate substrate nitrocellulose membrane thickness membrane for absorption of the sample (7). Collected multimembrane composite is cut into strips with a width of 5 mm, and a length of 60 mm, which are placed in plastic about what rivki (figure 2), the upper surface of which has an aperture for application of the sample (S) and the hole to read the results of the analysis are indicated by the letters T and C.

Example 3. The definition of the cholera exotoxin (XT)

Use multimembrane composite (example 2), in the analytical zone of which is coated with polyclonal rabbit antibodies to XT in the control zone caused goat anti-rabbit antibody conjugate pad contains a conjugate of Hydrosol particles GCFI with antibodies to XT. In the hole for application of the sample contribute 120 μl of a solution containing 1% tween-20 and 1% BSA, or XT, in the same solution.

Introduction in the hole for applying a sample of 120 μl of 1% solution of tween-20 and 1% BSA leads to the appearance of an intense band of blue color in the control area, and its absence in the area of analytical strip (figa), indicating the absence of non-specific interaction of the conjugates of Hydrosol particles GCFI containing antibodies to XT, with anticholergenic antibodies and components of the blood serum applied in the analytical area.

Introduction to the study sample XT (source - cholera vaccine) causes the membrane of two stripes of blue: analytical (8) and control (9) zones (figb). The intensity of the signal in the analytical zone increases with increasing concentration of the analyte in the sample and the time of analysis. The range of the measured concentrations of XT from 2.0 to 500 μg/ml source solution for protein. A plot of the intensity of staining of the control area on the concentration of XT in the sample is shown in figure 4. The analysis 2-15 minutes Evaluation of signal intensity in the analytical area visually or register using the analyzer videochefosama "Relcom".

If the introduction of the hole for the application of the sample solutions is not formed colored lines (pigv), this test system is considered unsuitable, and its results are ignored.

Example 4. Determining in a sample of blood serum titer of antibodies to the causative agent of salmonellosis

Use multimembrane composite (obtained according to example 2), in the analytical zone which caused the preparation of lipopolysaccharide (LPS) of Salmonella typhimurium, in the control zone marked monoclonal antibodies to the O-antigen of Salmonella group B, conjugate pad contains a conjugate of Hydrosol particles GCFI with sorbed to LPS of S. typhimurium with OD=3.0mm. To obtain the conjugate use FSC solutions with a concentration of 50 ng/ml and 200 ng/ml.

The solution containing 1% tween-20 and 1% BSA, prepared a number of consecutive working dilutions of antisera kS.typhimurium from 1:20 to 1:10 240.

Introduction in the hole for the sample of 120 μl of a solution consisting of 1% tween-20 and 1% BSA, leads to the appearance in the area of the control area of the blue color in the absence of a colored band in the range of the Academy of Sciences of the lytic zone.

Introduction to the above mixture AT the causative agent of salmonellosis causes in the analytical zone of the blue color, the intensity of which depends on the level of specific AT and reaction time. Measurement range of titles AT the causative agent of salmonellosis from 1:20 to 1:5120. Figure 5 shows a plot of the intensity of staining analytical areas from cultivation of antibodies to the causative agent of salmonellosis. The analysis 1-15 minutes Evaluation of signal intensity in the analytical area visually or register using the analyzer videochefosama "Relcom".

Example 5. Determining in a sample of serum or plasma blood of antibodies to Mycobacterium tuberculosis

Use multimembrane composite (obtained according to example 2), in the analytical zone of which is coated with recombinant antigen - glycoprotein complex of Mycobacterium tuberculosis H37Rv, in the control zone caused goat rabbit antibody conjugate pad contains a conjugate of Hydrosol particles GCFI OP=3,0 adsorbed rabbit antibodies to the sum of human immunoglobulins classes (IgG+IgM+IgA). The solution containing 1% tween-20 and 1% BSA, prepared a number of consecutive working dilutions of serum (plasma) blood of the person from 1:2 to 1:10, suspected of containing antibodies to the causative agent of tuberculosis

Introduction in the hole DL the sample 120 ál solution consisting of 1% tween-20 and 1% BSA, leads to the appearance in the area of the control area of the blue color in the absence of a colored band in the analytical zone (figb), indicating that the absence in the sample of antibodies to Mycobacterium tuberculosis. Introduction in the hole for the sample of 120 μl of a solution consisting of 1% tween-20 and 1% BSA and containing serum (plasma) blood of a person infected with TB, leads to the appearance in the analytical zone of the two stripes of blue (tiga), the intensity of which depends on the level AT specific and reaction time. The analysis 1-15 minutes to Check the results visually or by instrument "Relcom". If the introduction of the hole for the application of the sample solutions is not formed colored lines (pigv), this test system is considered unsuitable, and its results are ignored.

Example 6. The timing of maximum analytical signal for use as the dispersed phase Hydrosol GCFI and colloidal gold on the sample registration XT.

The example illustrates the reduction of the time analysis of the same analyte, ceteris paribus.

Received multimembrane composite with GCFI (as described in example 2) and multimembrane composite with colloidal gold.

To obtain conjugates is ASTIC colloidal gold with antibodies to XT to the preparation of colloidal gold is added an aqueous solution, containing anticholergenic antibody mixture is incubated on a magnetic stirrer for 10 min at room temperature and add BSA to a final concentration of 0.1%. After 1 min incubation in the same conditions, the mixture is centrifuged at 50000 × g for 30 minutes, the Suspension is washed twice with a solution containing 10% sucrose and 0.25% BSA, each time depositing the particles by centrifugation in the same conditions. The precipitate is suspended in 0.02 M phosphate buffer pH of 7.9, containing 10% sucrose and 0.25% BSA. Determine the wavelength of the absorption maximum and the value of OP in the maximum on the spectrophotometer. Using the same buffer solution to prepare a suspension of gold Sol with OP, is 3.0 at the maximum absorption, and put it on the membrane to conjugate. The membrane is dried as described in example 1. The Assembly multimembrane of the composite is carried out, as described in example 2.

In the hole for applying a sample frame containing the conjugate particles of colloidal gold with anticholergenic antibodies contribute 120 μl of the solution XT, prepared in 0.1 M phosphate buffer solution pH of 7.9, containing 0.5% BSA and 0.4% tween-20.

XT determined as described in example 3. The intensity of the signal in the analytical area is recorded by videochefosama analyzer Replicom". A plot of signal intensity in the analytical area from the time of the analysis are the and 6. When used as the dispersed phase Hydrosol GCFI the maximum signal in the zone of the analytical strip appears in 5 min after introduction of the sample. When using conjugates of gold Sol for maximum analytical response requires a longer time (>40 min).

For all the above-mentioned examples is characterized by fast moving particles conjugate Hydrosol GCFI on nitrocellulose membrane and the appearance of colored stripes in analytical and control zones, which distinguishes immunochromatographic tests using hydrosaline drugs GCFI from the immunochromatographic test based sols of gold, for which the analysis is 15-40 minutes At high concentrations of analyte visible to the eye signal appeared after 1-2 min after injection of the sample. The Hydrosol particles GCFI sensitized with an antibody or antigen, passed through the analytical membrane smooth front, almost giving "stains" and "flows". Background analytical membrane was from white to light blue, formed line in analytical and control zones differed with good contrast.

Increases the contrast between the analytical area and the white surface of the membrane is ensured due to more complete removal of coloring Hydrosol of hexacyanoferrate (II) iron (III) and as a consequence, improvement in visual or instrument of registration of the results of the analysis.

The method of conducting lateral flow assay for detection of analytes in the sample, including the creation of multimembrane composite, consisting of membranes for application of the sample, membranes for conjugate, analytical membrane and membrane for absorption of the sample containing dry conjugate of the dispersed phase with the antibody (antigen) to one of the antigenic epitopes of the analyte, causing immunoreagent (second antibodies to other antigenic epitope of the analyte or antigen) on the analytical membrane in the form of a narrow zone, conducting the immunochemical reaction and separation immunochemical complex at the same time during the chromatographic migration on the surface of the analytical membrane, characterized in that the dispersed phase to the first conjugation antibodies (antigen) use the Hydrosol of hexacyanoferrate (II) iron (III).



 

Same patents:

FIELD: medicine.

SUBSTANCE: device contains precipitation plate with applied on it layer of gel, which contains antibodies, illuminator of precipitation plate in form of source of diffusive light of toroidal form and reading videodevice, connected with computer. Precipitation plate is made in form of disk and placed on base, provided with light adsorbing coating from precipitation plate side, in such way, that geometrical centers of reading videodevice, illuminator and precipitation plate are placed on one vertical axis.

EFFECT: device application allows to provide even contrast range of immuno-diffusive rings image on precipitation plate, to read them with required accuracy, and increase productivity of measurement process.

3 cl, 4 dwg

FIELD: medicine.

SUBSTANCE: invention concerns medicine area, namely to diagnostics of infectious diseases and concerns a method of revealing antibodies to antigens of hepatitis viruses. The essence of the way consists in use of an immunochromatographic strip which consists from fiber glass non-sorbing matrix (6) on which the strips containing conjugates of painted particles with monoclonal antibodies to human immunoglobulins G and M (7,8) plastic substrates are placed and immobilised by means of drying, covered with an adhesin layer with low penetrating ability (1) on which a microporous nitrate cellulose membrane with evaporated on a side close to the substrate with a protective polymeric basis (2) is mounted, and admixtures of antigens of viruses of a hepatitis A, B, C (3, 4, 5) are placed perpendicularly to the long side of the strip on the membrane from the non-protected side. The strip is moistened with investigated blood serum and at detection of the painted strip around the strip of antigens of hepatitis A, antibodies to an antigen of a virus of hepatitis A are detected. At detection of the painted strip around the strip of antigens of hepatitis B, antibodies to an antigen of a virus of hepatitis B are detected. At detection of the painted strip around the strip of antigens of hepatitis C, antibodies to an antigen of a virus of hepatitis C are detected, which are formed at linkage of antibodies of blood serum with conjugates of antibodies to immunoglobulins G and M with the painted particles.

EFFECT: method allows to tap simultaneously antibodies to antigens of viruses of a hepatitis A, B, C and to define to what type they are specific to.

2 ex, 1 tbl, 1 dwg

FIELD: medicine.

SUBSTANCE: invention concerns research and analysis of biological materials. Invention includes membrane with input zone for liquid sample application at least two indicator zones which can interact with the analyte(s), and at least one absorption zone absorbing liquid after they pass indicator zones. Indicator zones are positioned between input zone and absorption zone. Flow directions (tracks) from input zone through respective indicator zones to absorption zone are almost parallel. At least two different tracks are present. Invention claims method of defining several analytes or their derivatives in liquid sample, involving sample application to input zone of membrane. Indicated amount of the sample is sufficient to maintain sample fluid passage in direction of absorption zone through indicator zones and formation of complexes by analytes or their derivatives in indicator zones.

EFFECT: possible simultaneous definition of cell and plasmatic sample parametres.

25 cl, 14 dwg, 6 ex, 7 tbl

FIELD: medicine.

SUBSTANCE: invention concerns medicine area, namely, urology, and can be used for predicition of a persistent infection of a prostate formation. Determine presence of precipitate factors to a spermine and/or a spermidine in semenal plasma of the patient.

EFFECT: increase of sensitivity and accuracy of forecasting of formation of chronic prostatitis.

4 ex

FIELD: biotechnology, microbiology.

SUBSTANCE: invention proposes a novel method for immunochromatography detection of Escherichia coli in water using a test-band. Test-band consists of A, B and C assembled on backing D wherein part of A overlaps part B by 1-2 mm. Part A represents inert porous carrier made of fiber glass (PED) and comprises reaction zone 1 applied on it (with applied conjugate of rabbit monospecific antibodies raised to E. coli with horse radish peroxidase) and reaction zone 2 (with applied conjugate of murine monoclonal antibodies labeled with colloidal gold against horse radish peroxidase). Conjugates are applied on reaction zone by parallel bands in the central region of PED and perpendicularly to liquid flow in distance 2 mm between conjugates. Part B in test-band represents nitrocellulose immobilized on lavsan base with applied test-zone 6 and control zone 7. Test-zone 6 represents zone with applied band parallel to long side of test-band for 2 mm from middle of nitrocellulose with rabbit monospecific antibodies against E. coli. Control zone 7 represents zone with applied by parallel band for 3 mm from middle and nearer to suction filter C by rabbit monospecific antibodies against murine immunoglobulins. Part B shows a capillary linkage with reaction zone of carrier A. C-suction filter is used for absorption of unreacted reagents. Presence of Escherichia coli is estimated by the presence of color of test-zone 6. Color of control zone shows working capacity of test-band. Using invention provides universality of band in detection of any type of strains Escherichia coli with sensitivity 1-10-8 g/ml.

EFFECT: improved method for detection.

3 dwg, 1 ex

FIELD: medicine.

SUBSTANCE: method involves determining precipitating factors to lyzozyme and/or lactoferrin in patient blood serum as diagnostic test. The factors being found, persistent infection is to be diagnosed. No factors being found, recovery is to be predicted.

EFFECT: high accuracy of prediction.

1 tbl

The invention relates to an analytical device for determining analytes in a liquid dairy product using capillary migration of the specified dairy product, comprising a solid substrate having first and second end, which strengthened sequentially, starting from the first end of the membrane for the purification of the analyzed liquid membrane immobilized on which one or more exciting substances, and absorbent membrane

The invention relates to the definition present in the liquid sample being analyzed component in the form of chemical, biochemical or biological complex

The invention relates to the field of immunology, in particular clinical immunology and relates to a method of immunocorrective therapy preclinical and clinically significant forms of immunological failure and rapid screening tools for immune

The invention relates to medicine, namely to rheumatology, and relates to a method of diagnosis of purpura Seleina's disease

FIELD: chemistry.

SUBSTANCE: in order to produce a biospecific polymer sorbent for extracting proteinase, an aqueous solution undergoes radical polymerisation, where said solution contains 0.1-1.5 wt % ovomucoid from duck egg albumen, acylated with acyl chloride of acrylic or methacrylic acid, 7.0-20.0 wt % hydrophilic monomer consisting of acrylamide, methacrylamide or N-vinylpyrrolidon, and 0.7-15.0 bifunctional cross-linking agent. The hydrophilic monomer is further mixed with N-ethylbromide dimethylaminoethyl methacrylate, taken in amount of 45-55 wt % with respect to weight of the hydrophilic monomer.

EFFECT: addition of N-ethylbromide dimethylaminoethyl methacrylate to the hydrophilic monomer increases efficiency of sorption of proteinase from biological fluids.

1 tbl, 2 ex

FIELD: medicine.

SUBSTANCE: tissue sections are incubated simultaneously with mixed primary and secondary antibodies. Primarily, the primary antibodies are applied on the tissues, and in 15 seconds of the incubation process - the secondary antibodies without washing with a buffer. It is followed with combined incubation for 4 minutes. A substrate-chromogen solution for the incubation is prepared immediately before the application, the sections are washed with a buffer solution and distilled water heated to temperature 37°C.

EFFECT: method allows fast and precise tumour histogenesis, detection of tumour cells within resection boundaries.

6 dwg, 2 tbl, 6 ex

FIELD: medicine.

SUBSTANCE: invention describes a method of immunochemical streptomycin detection wherein on a surface of a piezoelectric immunosensor, a receptor coating with a streptomycin-protein (bovine serum albumin) conjugate is immobilised, a settled number of streptomycin antibodies is added to a sample and kept for 2-3 min to produce an immune complex which is introduced in a cell in a phosphate buffer solution of pH = 7.1-7.5 and to record the sensor oscillation frequency variations when reacting with the streptomycin-protein conjugate with streptomycin antibodies, an analytical signal is inversely proportional to the concentration of streptomycin in the analysed sample, while the concentration is derived from the calibration chart, the receptor coating is regenerated by applying 0.03 mM potassium thiocyanate solution on the surface.

EFFECT: higher sensitivity of streptomycin detection in composite mixtures, ensured multiple use of the immunosensor after regeneration of the bioreceptor coating that reduces analysis price.

13 dwg, 1 tbl

FIELD: medicine.

SUBSTANCE: specific enzymatic hydrolysis of proteins immobilised on a substrate surface of a scanning probe microscope is enabled by the fact that proteins are exposed to ultrasound and a site-specific enzyme in a buffer solution of such pH value and temperature to ensure enzyme activity. Before the enzymatic hydrolysis, the substrate of the scanning probe microscope is immersed in 1-5% acetic acid and exposed to high-frequency ultrasound 1-3 MHz and power 1 Wt/cm2 and more at such temperature of the solution to ensure breaking of the protein-surface bonds. Then, the solution containing desorbed proteins is frozen and lyophilised. The substrate of the scanning probe microscope may consist of silicon, glass, mica, sapphire. The substrate surface of the scanning probe microscope whereon protein molecules are activated, is chemically activated by silane.

EFFECT: use of the method allows lowering a detection threshold of protein molecules immobilised on the substrate surface of the scanning probe microscope by the use of a mass spectrometer.

2 ex

FIELD: medicine.

SUBSTANCE: patient's blood serum is analysed by ELIZA, determining concentrations of antibodies, specific to components of male reproductive tissues. As components of male reproductive tissues, used is nuclear antigen of pachytene spermatocytes of rodents. If values of antibody concentration, expressed in units of optic density, equals or is higher than 0.37, presence of reproductive dysfunction in men is diagnosed.

EFFECT: invention makes it possible to obtain accurate evaluation of presence of reproductive dysfunction in men due to elaborated accurate quantitative diagnostic criterion.

10 ex

FIELD: medicine.

SUBSTANCE: pharmaceutical preparation derinate representing sodium deoxyribonucleate is sorbed in microplate wells. Then, an analysed sample containing a component C3 of unknown activity and a solution of reagent R3 (component C3 activity deficient human blood serum) is added in the wells. It is followed with incubation, and after washing and drying of the dish, a conjugate of enzyme with component C3 antibodies and a substratum of this enzyme are added into the wells. The component C3 activity is evaluated by the amount of the prepared product of enzymatic reaction. A kit comprises a flat-bottomed microplate with sorbed derinate, the conjugate of enzyme with human complement component C3 antibodies, the reagent R3 (component C3 activity deficient human blood serum) and the substrate buffer.

EFFECT: method allows the reliable and high sensitive component C3 test.

1 dwg, 3 ex

FIELD: medicine.

SUBSTANCE: pharmaceutical preparation derinate representing sodium deoxyribonucleate is sorbed in microplate wells. Then, an analysed sample containing a component C3 of unknown activity and a solution of reagent R3 (component C3 activity deficient human blood serum) is added in the wells. It is followed with incubation, and after washing and drying of the dish, a conjugate of enzyme with component C3 antibodies and a substratum of this enzyme are added into the wells. The component C3 activity is evaluated by the amount of the prepared product of enzymatic reaction. A kit comprises a flat-bottomed microplate with sorbed derinate, the conjugate of enzyme with human complement component C3 antibodies, the reagent R3 (component C3 activity deficient human blood serum) and the substrate buffer.

EFFECT: method allows the reliable and high sensitive component C3 test.

1 dwg, 3 ex

FIELD: medicine.

SUBSTANCE: method of early tuberculosis diagnostics in HIV-infection involving immunologic marker monitoring by immune-enzyme assay is used to describe a complex of serum cytokines: tumour necrosis factor alpha, interleukin - 10 and a soluble receptor interleukin - 6 to fix the level, and while observing certain values of immunological markers, tuberculosis is diagnosed in the HIV-infected patients.

EFFECT: method allows higher quality and reduced length of early diagnostics of tuberculosis infection in HIV-infection.

1 dwg, 4 tbl

FIELD: medicine.

SUBSTANCE: invention refers to a method of human blood serum analysis for a soluble form of antigen CD50 dimer involving the use of CD50-specific monoclonal antibodies whereat a tray reaction includes tetramethyl benzidine as a substratum.

EFFECT: invention provides detecting the soluble form of antigen CD50 dimer in human blood serum.

1 cl, 2 ex

FIELD: medicine.

SUBSTANCE: invention refers to medicine and veterinary science, namely to microbiology and immunology. The method involves cultivation of Brucella abortus I-206 L-strain on a dense nutrient medium for L-brucellas at 37°C for 3-5 days. Thereafter, a microbial mass is washed of with buffered 0.9% normal saline of pH 7.2±0.2 and inactivated by adding 2.5% formalin. A bacterial suspension is kept for one day at 37°C with specific sterility being controlled. The bacterial suspension is reduced to concentration 4.5·1010-5·10 m.c. in ml with buffered 0.9% normal saline (pH 7.2±0.2) and thermally treated on a water bath at 100°C for 45-50 minutes. The suspension is centrifuged at 7000 rpm for 50-60 minutes; the supernatant is separated and frozen-dried. The method allows producing the preparation exhibiting high specificity and activity, available for preparing based immunobiological preparations and a test-system for human and animal blood serum examination for L-brucella antibodies.

EFFECT: method is technological, accessible, does not require using expensive devices and equipment.

2 ex

FIELD: medicine, ophthalmology.

SUBSTANCE: in lacrimal liquid one should detect the content of interleukin 8 (IL-8) and that of interleukin 1 beta (IL-1β) to calculate prognostic coefficient (PC) due to dividing the first value by the second one by the following formula: At PC value being below 10.0 one should predict favorable disease flow, and at PC value being above 10.0 - unfavorable flow.

EFFECT: higher accuracy of prediction.

2 ex

Up!