Diagnostic technique for membrane toxicity
SUBSTANCE: method involves taking blood at 1 ml per a sample into the Vakutainer evacuated test tube with sprayed ethylene diaminetetraacetic acid 2.2 mg; a reference sample 1.0 ml and a sample 1.0 ml with weighted xenobiotics are taken from the total volume into the Eppendorf tubes; 30-minute incubation is followed by using a uniform procedure to prepare blood smears by applying on a dry slide to prepare a smear to be stained according to Leishman; blood corpuscles are analysed by the Zeiss light microscope at a magnification 1500. A microscope field showing echinocytes with styloid processes of the membrane not found in the reference sample is a sign of the xenobiotic membrane toxicity.
EFFECT: invention enables an instant assessment of the membrane toxicity of the substance.
The invention relates to medicine, namely membranology, and can be used to test embryotoxicity xenobiotics used in therapeutic and orthopedic dentistry as restorative and reconstructive materials, as well as in toxicology and pharmacology along with traditional methods of evaluating the toxicity of substances, medicines.
There is a method of determining the potential cytotoxicity of biomaterials on cell cultures of mouse fibroblasts (L-929), based on the determination of qualitative and quantitative changes in cell cultures after incubation with a filling material.
The disadvantage of this method is that it is time-consuming, costly and not sufficiently specific.
Known "Method of determining embryotoxic properties of xenobiotics" (Patent RF №2073244, date of publication - 10.02.1997) using the reaction spontaneous rosethorne lymphocytes before and after administration of the test substances in experimental animals, which is close to the estimated effect and the achieved result.
The disadvantage of this taken as a prototype of this method is the length, complexity, the multiple steps involved, the need for the use of experimental animals with subsequent treatment of the tissue, receiving only delayed is the result.
The aim of the invention is to develop a method of assessing embryotoxicity xenobiotics, which allows a rapid assessment of the substance.
The technical result is achieved by the fact that in a vacuum tube Vakutainer with powder 2.2 mg ethylenediaminetetraacetic acid blood is taken at the rate of 1 ml per sample, then the total volume taken by 1.0 ml tube (Eppendorf, control and contains a sample of xenobiotics, after 30-minute incubation on a standardized method to prepare blood smears on dry glass slide put a drop of blood, get a PAP smear and paint it according to the method of Leishman and conduct the study of the formed elements of the blood with the help of light microscope "Zeiss" with increasing 1500. Indicator of embryotoxicity of the xenobiotic is the emergence of the field of view of echinocytes - erythrocytes with subulate appendages membrane absent in the control.
The technical result, which directed the establishment of this invention is the development of rapid, specific, cost-efficient method of determining embryotoxicity xenobiotics.
The method is as follows: in the vacuum tube Vakutainer with powder 2.2 mg ethylenediaminetetraacetic acid took 4 ml of blood was then collected in 1.0 ml of 4 Eppendorf tubes: control (1)and containing 25 mg of hanging the cured dental composites and adhesive Filtek Ultimate (2), Filtek Bulk Fill (3) and Single Bond Universal (4). After 30-minute incubation on a standardized methodology was preparing blood smears on dry glass slide was put a drop of the studied blood and distributed it using pure grinding glass, holding at an angle of 45°. Blood smears were located at a distance of 1.0-1.5 cm from the edges of the glass, occupying almost the whole of its length, and ended the "whisk". Received drugs (1-4) was dried in the air, marked, stained by the method of Leishman, washed with distilled water, dried in the air. Criteria of a good color was pink erythrocytes, violet staining granularity of neutrophils on a pink background, gentle Ashrafinia grain monocytes. Morphological examination of blood cells was performed using a light microscope "Zeiss" with increasing 1500. Indicator of embryotoxicity of the xenobiotic is the emergence of the field of view of echinocytes - erythrocytes with subulate appendages membrane absent in the control. In the sample 2 with Filtek Ultimate, their number amounted to 1%, in the sample 3 with Filtek Bulk Fill is 3%, and the sample 4 with a Single Bond Universal number of echinocytes was 12%. Thus, due to the research of the adhesive has a sufficiently strong embryotoxicity properties that require hermetic isolation from contact with the tissues Polo the tees mouth.
The method can be used for rapid assessment of embryotoxicity various xenobiotics.
1. GOST RISO 10993-5-2009.
2. "Method for determining embryotoxic properties of xenobiotics" (Patent RF №2073244, date of publication - 10.02.1997).
The way the rapid assessment of embryotoxicity of xenobiotics, including the study of membrane structures of reactive cells under the influence of the analyte, characterized in that the vacuum tube Vakutainer with powder 2.2 mg ethylenediaminetetraacetic acid blood is taken at the rate of 1 ml per sample, then the total volume taken by 1.0 ml tube (Eppendorf, control and contains a sample of xenobiotics, after 30-minute incubation on a standardized method to prepare blood smears on dry glass slide put a drop of blood, get a PAP smear, paint his method Leishman and conduct the study of the formed elements of the blood with the help of light microscope "Zeiss" with increasing 1500, the emergence of the field of view of echinocytes - erythrocytes with subulate appendages membrane absent in the control, is indicative of embryotoxicity of the xenobiotic.
SUBSTANCE: blood cell sample is used to prepare slides of several layers of agarose one of which contains individual non-dividing nucleated cells. Damaging the cell DNA is carried out by phosphate-buffer saline with pH 7.4-7.5 with ozone-oxygen mixture flown through preliminary for 5 min with the ozone concentration of 500-1000 mcg/l. The cells are broken down at pH=10; the DNA is denatured by placing the slides into an alkaline solution at pH>13; the damaged DNAs of the broken down cells are subject to an electrophoresis in the solution of pH>13 at voltage 27 V, current intensity 260-270 mA, field density 2.0 V/cm. The fluorescent stained damaged DNAs are recorded by photographing, software-processed; a percentage ratio of the damage DNAs to their total number is calculated; if the derived ratio is more than 10%, the induced DNA damages are stated.
EFFECT: method enables diagnosing malignant new growths, selecting drug preparations and geriatric protectors in radio-, chemo- and ozone therapy.
SUBSTANCE: tumour tissue is detected for total proteasome activity, 20S proteasome activity and NF-κB p50, HIF-1α expression, and discriminant functions Y1, Y2 are calculated by equations. If observing Y1>Y2, the absence of lymphogenic metastases is predicted, while Y1<Y2 enables predicting developing lymphogenic metastases.
EFFECT: higher prediction accuracy.
FIELD: veterinary medicine.
SUBSTANCE: method comprises carrying out serological studies and identifying animals reacting positively. Additionally, after carrying out of serological studies the preparation in the form of a mixture of the medical solution formaldehyde with isotonic sodium chloride solution is administered to the pigs of all age groups at a ratio of parts by weight 2-6:994-998. The preparation is administered twice intramuscularly at a dose of 5-6 ml per head with an interval of 7-8 days between injections. 10-12 days after the preparation is administered the additional serological studies are carried out and the infected animals are identified.
EFFECT: method enables to increase the percentage of identifying animals in relation to actinobacillus pleuropneumonia.
3 tbl, 3 ex
SUBSTANCE: invention represents method for prediction of threshold retinopathy of prematurity in infants having no ophthalmic signs of the disease characterised by the fact that blood serum is analysed for vascular endothelial growth factor (VEGF) up to the 33rd week of gestation; if the VEGF level is 1300 pg/ml or more, developing threshold retinopathy of prematurity is predicted.
EFFECT: enabling correction of therapeutic approach to premature newborns at the stages of nursing.
2 dwg, 1 tbl, 5 ex
SUBSTANCE: invention represents a diagnostic technique for external genital endometriosis involving blood serum analysis, differing by the fact that the blood serum is analysed for high-density lipoproteins, and if the derived value is 0.77 mmole/l and higher, external genital endometriosis is diagnosed.
EFFECT: invention enables providing higher accuracy and simplifying a diagnostic procedure for external genital endometriosis.
SUBSTANCE: summary of declared method consists in determining the anticholinesterase effect of solutions of the examined anticholinesterase compounds (AnCEC) of a preparation of cholinesterase - the enzyme propionyl cholinesterase of calamari brain tissue (PCE), namely in estimating a percentage PCE activity inhibition in solutions of low (0.04 U/ml) and high (4 U/ml) concentrations. A toxicity hazard of the examined AnCEC solutions is assessed by a difference of their effect on PCE activity in the solutions of low and high concentrations. If a percentage of PCE activity inhibition in the solution of high concentration goes to zero or does not exceed 5-10%, the examined solution is stated to contain high-toxicity AnCEC of the 1st hazard class. If the inhibitory effect size remains constant or falls by no more than 10%, the presence of low-toxicity AnCEC is stated.
EFFECT: using the declared method enables assessing the toxicity hazard of the anticholinesterase compounds for 30-60 minutes in various situations, where their composition is unknown or they present in the form of solutions of an unknown concentration.
1 tbl, 2 ex
SUBSTANCE: invention refers to medicine, namely to cardiology, and concerns diagnosis of a functional class of chronic cardiac failure (CCF). The diagnosis is implemented by developed formula taking into account assessed variations of a left ventricular end diastolic and a diastolic septal wall thickness determined by echocardiography, as well as variations of NT-terminal fragments of a brain natriuretic peptide precursor. The functional class of CCF is verified by a relation of expected values and the functional class NYHA of CCF.
EFFECT: method enables the adequate diagnosis of the functional class of CCF, including in the patients with counterindications for physical and pharmacological loading tests, as well as with no equipment required for stress tests.
4 ex, 1 tbl
SUBSTANCE: invention refers to medicine, namely to oncohaematology, and can be used for prediction of the clinical effectiveness in the patients with non-Hodgkin lymphomas with bone marrow involvement. That is ensured by determining a lymphocyte volume and electric conductivity in patient's peripheral blood before the treatment and every next course of the chemotherapy with calculating a relation of the values. If observing a decrease of the relations as compared to the initial value, the positive clinical effectiveness and the absence of generalisation symptoms are predicted. The absence of the decrease or increase of the relations shows the absence or low clinical effectiveness and the presence of the symptoms of a continued tumour growth.
EFFECT: using of given method enables determining the sensitivity of the patients suffering non-Hodgkin lymphomas to the chemotherapy and performing the immediate and remote prediction of the clinical course of the disease.
1 tbl, 3 ex
SUBSTANCE: invention refers to medicine, and can be used for fecal sample automatic analysis. The fecal sample automatic analyser comprises an automatic controller, a sample container, a thinning device, an agitating and mixing device, an analysing device, a suction and cleaning device piped to the analysing device. The analysing device comprises a physical analysis unit and a chemical analysis unit. The suction and cleaning device comprises the sample suction needle, a thinner receiving unit and a peristaltic sample suction pump connected between the sample suction needle and the thinner receiving unit. The analysing device is connected between the sample suction needle and the peristaltic sample suction pump; when the sample suction needle is inserted into the sample container, and the peristaltic sample suction pump is positively rotated, the sample suction needle sucks the samples in from the sample container and supplies the fecal samples to the analysing device. The peristaltic sample suction pump is configured for reverse rotation and suction of the thinner from the thinner receiving unit for cleaning the analysing device and connection pipes after an analysis procedure.
EFFECT: invention provides reducing environment and laboratory contamination in the process of analysis, as well as higher performance.
15 cl, 6 dwg
FIELD: veterinary medicine.
SUBSTANCE: sample is placed in a glass, poured with the flotation liquid, stirred, filtered, the slurry is allowed to equilibrate for 15-20 minutes to detect protozoa oocysts or for 40 minutes to detect helminth eggs. After that the droplets of the surface film are transferred to the glass slide and microscopic examination is performed. Additionally, before addition of the flotation liquid, for dissolving the fat cells the aqueous solution of industrial alcohol is added and stirred. The ratio of water and alcohol is 1:1. It is allowed to equilibrate for 3-5 minutes.
EFFECT: method enables to detect simultaneously protozoa oocysts and helminth eggs of gastrointestinal tract of animals in milk period.
SUBSTANCE: lactic acid is transferred from a sample into a solution and voltammetric accumulation of lactic acid in the stirred solution is performed while bubbling with an inert gas for 30 s and with electroaccumulation potential of 1.2-1.4 V relative to a saturated silver chloride electrode on a background electrolyte of 0.1 M Na2HPO4, followed by detection of cathode peaks in differential mode of recording voltamperograms with potential sweep rate of 30-40 mV/s; concentration of lactic acid is determined from peak height in the potential range of 0.25-0.40 V by a standard addition method.
EFFECT: method is simple, does not require a large amount of reactants and labour costs.
2 ex, 1 tbl
SUBSTANCE: method consists in the application of glutationreductase and catalase enzymes as test-objects for determination of the anti-oxidant activity by the ratio of the rate of enzymatic reaction on the test-object after the substance addition and the rate of enzymatic reaction before the substance addition, which must be larger than 1. Preliminarily, before addition to an incubation medium, the samples of essential oil of Siberian fir are diluted with dimethylsulphoxide in a ratio of 1:1.
EFFECT: increased accuracy of determination.
2 ex, 1 tbl
SUBSTANCE: invention relates to a method of early detection of muscular degenerative diseases and to a method of prediction and/or determination of a therapeutic efficiency of a therapeutic preparation and/or a method of disease therapy by measurements of tetranor-PGDM (11,15-dioxo-9α-hydroxy-2,3,4,5-tetranorprostan-1,20-dioic acid) in a subject's urine sample and comparison of its content relative to a sample, separated from a healthy individual. A muscular degenerative disease is identified in a patient if the concentration or content of tetranor-PGDM in a sample, separated in the patient is higher than the concentration or content of tetranor-PGDM in a sample, separated from a healthy individual. An efficiency of the therapeutic preparation and/or the method of therapy of the muscular degenerative disease is determined by comparison of the content of tetranor-PGDM in the sample, separated from the patient with the muscular degenerative disease before and after introduction of the therapeutic preparation. If the measured content of tetranor-PGDM in the sample considerably or inconsiderably decreases after the introduction of the therapeutic preparation, the method of therapy is efficient. The invention also relates to a set for diagnostics of muscular degenerative diseases, which includes an antibody to tetranor-PGDM, labelled tetranor-PGDM and optionally, at least, one compound, selected from a group, consisting of an antibody to immunoglobulin, a diluting solution for the sample, a diluting solution for the antibody and labelled tetranor-PGDM, tetranor-PGDM standard of the known concentration, a substrate for an enzyme immunoassay and a stopping solution for the enzyme immunoassay.
EFFECT: invention is efficient and simple in implementation.
7 cl, 1 tbl, 1 ex, 2 dwg
SUBSTANCE: method of codeine identification includes separation of mixture components by a method of thin-layer chromatography with separation of a chromatographic zone of codeine by a reagent of Dragendorff, and quantitative determination of codeine is carried out in an area of its manifested zone directly in a solid phase by measuring diffusion reflection coefficient at a wave length of 520 nm.
EFFECT: reduction of time of identification and quantitative determination of codeine, reduction of the process labour consumption, reduction of a probability of the target component loss.
2 tbl, 3 ex
FIELD: medicine, pharmaceutics.
SUBSTANCE: invention relates to chemical-pharmaceutical industry and represents a preparation for involving a mesenchymal stem cell of the bone marrow into peripheral blood from the bone marrow, which is introduced into the blood vessel or muscle and which contains any of components: (a) protein HMGB1; (b) HMGB1 protein-secreting cell; (c) a vector, into which HMGB1 protein-coding DNA is inserted; (d) protein HMGB2; (e) HMGB2 protein-secreting cell; (f) a vector, into which HMGB2 protein-coding DNA is inserted; (g) protein HMGB3; (h) HMGB3 protein-secreting cell; and (i) a vector, into which HMGB3 protein-coding DNA is inserted.
EFFECT: elaboration of the preparation for involving the mesenchymal stem cell of the bone marrow into peripheral blood from the bone marrow.
3 cl, 6 ex, 1 tbl, 14 dwg
SUBSTANCE: invention refers to medicine, and describes a method for testing a planned or known immunomodulatory agent for T-cell activation which involves the stage of contacting a peripheral blood mononuclear cell (PBMC) culture with the pre-determined amount of the planned or known immunomodulatory agent in vitro and observing the T-cell activation in the PBMC culture using a readout system, when contacting the planned or known immunomodulatory agent, wherein the PBMC culture density at the stage of pre-culture makes at least 2×106/ml, preferentially at least 5×106/ml, more preferentially at least 107/ml, or at least 4×105/cm2, preferentially at least 106/cm2, most preferentially at least 2×106/cm2; the PBMC pre-culture is cultured for at least 12 hours.
EFFECT: invention provides the improved agent for testing the immunomodulatory agents in vitro.
10 cl, 16 dwg, 13 ex
SUBSTANCE: method concerns quantitative picamilon measurement. Solutions of a measured substance (concentration of 0.00002 g/ml) and a reference sample are prepared. A solvent for preparing the measured solutions is presented by 0.1 M sodium hydroxide. The reference sample is methyl orange. An optical density of the solution of the measured substance and the reference sample is measured in a spectrophotometer at an analytical wave length of 261 nm. The results are calculated by formula with recalculation factor 0.7505 introduced therein.
EFFECT: method allows for increasing reproducibility of the measurement results, reducing cost price, labour-intensiveness and analysis error, and unifying the analysis procedure.
SUBSTANCE: method involves extraction preparation of a sample of biological material, centrifuging and performing analysis on a capillary electrophoresis system in a quartz capillary with an effective length of 0.5 m and inner diameter of 75 mcm, wherein analysis is performed using an aqueous main electrolyte which contains 0.28% boric acid and 0.04% sodium tetraborate, with positive voltage polarity and detection wavelength of 254 nm.
EFFECT: rapid and reliable quantitative determination of indolyl-acetic acid by capillary electrophoresis using nontoxic and readily available reactants for analysis.
6 ex, 1 tbl, 1 dwg
SUBSTANCE: solutions of an analyte (bendazole) and a comparison sample are prepared. The solvent used to prepare the test solutions is 0.1M hydrochloric acid solution. The comparison sample used is benzoic acid or phenolphthalein. Optical density of the analyte (bendazole) solution and the comparison sample is measured on a spectrophotometer with analytical wavelength of 270 nm. The results are calculated using a formula by entering a calculation factor of 0.181 into said formula when determining using benzoic acid and 0.293 when determining using phenolphthalein.
EFFECT: method increases reproducibility of measurement results, reduces cost, labour input, analytical error and enables unification of the analysis method.
SUBSTANCE: method of extracting novocaine from aqueous media with a mixture of phenetole and ethyl acetate, characterised by that it involves preparing an aqueous salt solution of novocaine by placing aqueous novocaine solution with known concentration into a measuring flask, making up to volume with a saturated solution of a salting-out agent in form of ammonium sulphate, extracting novocaine with a mixture of phenetole and ethyl acetate, taken in ratio of 1:1, by adding to the obtained aqueous salt solution of novocaine an extraction agent in form of a mixture of phenetole and ethyl acetate (1:1) with the ratio of the volume of the phases of the aqueous salt solution of novocaine to the extraction agent of 5:1, extracting on a vibratory blender until inter-phase equilibrium is established, after demixing of the system, separating the aqueous salt solution from the organic phase and analysing by UV spectrophotometry, measuring optical density of the aqueous salt solution on a UV spectrophotometer at wavelength of 291 nm and on a calibration curve which is plotted in coordinates of optical density of the aqueous salt solution and concentration of novocaine, finding content of novocaine in the aqueous medium; using the concentration to calculate the distribution coefficient and degree of extraction of novocaine.
EFFECT: method enables to completely extract novocaine from aqueous media, intensifies the extraction process, provides rapid and accurate determination of concentration of novocaine.
FIELD: microbiology, pharmaceutical agents.
SUBSTANCE: invention relates to method for determination of genome response of specific cells to vegetable extract action. Method for drug screening includes 1) treatment of specific cells with vegetable extract; 2) protein or RNA isolation from said treated cells; 3) identification of isolated protein or RNA; 4) determination of compound(s) in said vegetable extract; 5) treatment of said cells with determined compound(s); 6) protein or RNA isolation from said treated cells; and 7) determination of compounds providing expression or suppression of said protein or RNA which have another concentration than in untreated said specific cells.
EFFECT: identification of individual compound(s) for screening of new pharmaceutical agents or new pharmaceutical application of existing drugs.