Method for prediction of clinical outcome of musculo-invasive bladder cancer following combination therapy
SUBSTANCE: invention refers to medicine, specifically to oncology, and concerns the prediction of the clinical outcome of musculo-invasive bladder cancer. The method involves measuring the activity of 26S proteasomes and NF-kB and p50 amounts in the tumour tissue. If the 26S proteasome activity is more than 18-1,000 units/mg of protein, and if the NF-kB p65/p50 coefficient is more than 1.0, a low probability of the recurrence is predicted; if the 26S proteasome activity is less than 18-1,000 units/mg of protein, and if the NF-kB p65/p50 coefficient is less than 1.0, a high probability of the tumour recurrence is predicted. The invention can be used for detecting the progression of the disease in the form of the tumour recurrence following the transurethral resection of the bladder tumour and M-VAC polychemotherapy.
EFFECT: using the invention provides the higher accuracy and information value for predicting the clinical outcome of the bladder cancer.
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The invention relates to medicine, specifically to Oncology, relates to methods for predicting outcomes of muscle-invasive bladder cancer and can be used to determine the progression of the disease in the appearance of the tumor relapses after conducting transurethral tumor resection and chemotherapy scheme M-VAC.
Currently used traditional methods of predicting the course of disease, primarily involve the analysis of the parameters characterizing the clinical features of the disease, namely the size of the tumor, the nature and extent of the process. According to the literature there are data on the prognostic significance of parameters such as the presence of microsatellite DNA sequences, mutations of proanagen, oncosuppressor, cell cycle regulation of angiogenic factors [3, 4, 7] and adhesion molecules  regarding the risk of recurrence of the disease. However, these data are controversial.
Closest to the claimed is a method of forecasting, which includes the determination of the polymorphism of the GSTP1 gene (glutatine S-transferase), which is associated with the metabolism of xenobiotics (RF Patent No. 2393772, publ 23.03.2009). Polymorphic variants of the gene associated with the change of enzyme activity, which can affect the ri is to the development of recurrence of malignant tumors. From lymphocytes of peripheral venous blood secrete DNA, carried out PCR analysis with subsequent restriction of the amplified fragments polymorphic locus Ile105Val GSTP1 gene. Upon detection of patients with superficial forms RMP homozygous genotype Val/Val predict a high risk of recurrence of bladder cancer (RMP). Homozygous genotype Ile/Ile is associated with minimal risk for the development of recurrence RMP.
The disadvantages of this method are the low sensitivity, accuracy, and comprehension, as well as the limited scope of the method, due to the lack of communication polymorphic variants of genes with enzyme activity, which is associated not only with them, but may be changed due to post-translational modifications of the polypeptide chain, and do not take into account the biological behavior of the tumor itself, which can have a pronounced effect on carcinogenesis.
A new technical challenge - improving the accuracy and usefulness of the prediction of disease progression in the risk of recurrence.
To solve the problem in the method for predicting the outcome of muscle-invasive bladder cancer after combined treatment by studies of the enzymes of metabolism determine the activity of the 26S proteasome and the content of NF-KB p65 and p50 in the tissue of the op is Holi and activity of 26S proteasomes more 18-1000 U /mg protein and when you factor NF-kB p65/p50 more than 1.0 predict a low likelihood of recurrence; when activity of 26S proteasomes less 18-1000 U /mg protein and factor NF-KB p65/p50 less than 1.0 predict a high probability of relapse development.
The method is as follows.
Spend fluorometrically the determination of the activity of the 26S proteasome, as well as enzyme-linked immunosorbent determining the expression of NF-kB (nuclear factor kappa-light-chain-enhancer of activated B cells - nuclear transcription factor kB) p65 and p50 in the tumor tissue. To do this, the operating material take samples of tumor tissue within 2-3 hours after the operation, clear of areas of necrosis, hemorrhage and placed in liquid nitrogen. Tissue samples to determine the activity of enzymes stored at -70°C and thawed no more than 1 time for research activity and maintenance of markers. Before determining the activity of the investigated enzymes frozen tumor tissue homogenized to a powder in liquid nitrogen. 26S proteasome allocate method vysalivaniya using ammonium sulfate to 40% saturation . Chymotrypsinogen activity of 26S proteasomes, ongoing chymotrypsinogen centers of proteasomes, judged by the hydrolysis fluorogenic of oligopeptides Suc-LLVY-AMC (Sigma, USA) . The resulting product register on fluorimetry "Hitachi-850 (Japan) at the wavelength of excitation 380 nm and emission 440 nm. To evaluate the activity of impurity PR is teas in the samples used specific inhibitor of proteasome - MG132. Per unit activity of proteasomes take the amount of enzyme, which hydrolyzes 1 nmol Suc-LLVY-AMC for 1 min Specific activity of proteasomes expressed in units of activity per 1 mg of protein. The protein content determined by the method of Lowry.
Preparation of nuclear extracts for determination of NF-kB (p50 and p65 forms) is carried out in accordance with the recommendations of the manufacturer Caymanchem (USA). Frozen tissue (100 mg) were homogenized in liquid nitrogen, then resuspending. The homogenate was centrifuged at 2400g and 4°C to obtain a residue, which resuspended in 50 μl of 50 mM Tris-HCl buffer (pH 7.5) and then centrifuged at 14000g and 4°C. the resulting supernatant used to determine the expression of transcription factors. The expression of activated forms of NF-kB p50, P65 determined in nuclear extracts by ELISA. The results of the determination of the content of transcription factors expressed in arbitrary units per mg of protein in the hole. And activity of 26S proteasomes more 18-1000 U /mg protein and when you factor NF-KB p65/p50 more than 1.0 predict a low likelihood of recurrence; activity of 26S proteasomes less 18-1000 U /mg protein and factor NF-KB p65/p50 less than 1.0 predict a high probability of relapse development.
This approach to predict the risk of recurrence for bladder cancer due to R the house premises.
One of the pathogenetic mechanisms underlying the development and progression of bladder cancer, is the change in the activity of proteasome system and expression regulated by these substrates, including NF-kB. It should be noted that regulatory step in many physiological and pathological processes is an intracellular protein degradation in the proteasome. 26 proteasome is able to activate NF-kB due to the rapid degradation of its own inhibitor IkB, which promote the development p105 - precursor of the p50 subunit of NF-kB [5; 10]. It is known that post-translational modification p105 - predecessor NF-kB p50 is performed using the 20S proteasome without the process of ubiquitination . It is shown that the expression of NF-kB and the products of its regulation high in the case of transitional cell carcinoma of the bladder. The level factor associated with lymphogenous metastasis of tumors [8; 11]. Great importance is given to the ratio of NF-kB p65/p50 , which is associated with the presence of the active form of factor.
The information content of the selected criteria is justified by the presence of communication activity of 26S proteasome and the expression of NF-kB p50 and p65 with recurrences of bladder cancer, which was carried out in 54 patients RMP T2-3N0M0(the average age 61,8±1.5 years), treated in the Department of General Oncology fsbi " research Institute on Oncology the GII WITH the RAMS from 2006 to 2012. In all patients the diagnosis was morphologically verified. Treatment consisted of 2 courses of systemic chemotherapy in the neoadjuvant mode according to the standard scheme MVAC (methotrexate 30 mg/m2intravenous in the 1st, 15th and 22nd day, vinblastine 4 mg/m2in the 1st, 15th and 22nd day of intravenous cisplatin 70 mg/m2in the 2nd day intravenous doxorubicin 30 mg/m2intravenously at day 2), a break between courses was 4-6 weeks. Subsequently these patients underwent transurethral resection (TUR) of the bladder and another 2 courses of adjuvant chemotherapy according to the above scheme.
Tumor tissue was taken from the operational material no later than 2-3 hours after surgery, frozen in liquid nitrogen. Then in tumor tissue was determined by the activity of the 26S proteasome and the expression of NF-kB p50 and p65.
The average term monitoring of patients included in the study was 22 months. The progression of the process was noted in 19 patients (35%) due to the development of recurrence of the tumor.
To assess the prognostic significance of the activity of 26S proteasome and the expression of NF-kB P50 and P65 in the tumor tissue for bladder cancer used univariate analysis of the prognostic significance of signs (Survival Analysis, Statistica 6.0). Figures 2-year relapse-free survival was analyzed using build distorting the x survival by the method of Kaplan-Meier. Comparison of survival rates in the groups was carried out according to the criterion of Gehan-Wilcoxon .
When evaluating the prognostic significance of enzyme activity systems of intracellular proteolysis in tumor tissue for bladder cancer revealed that a statistically significant factors in relation to disease-free survival, are the activity of the 26S proteasome and the expression of NF-kB p50 and p65. In Fig.1 presents the performance of 2-year disease free survival rates depending on the activity of 26S proteasomes in the tumor, Fig.2 - depending on factor NF-kB p65/P50 in the tumor, which confirms the high likelihood of relapse in the group of patients with the activity of 26S proteasomes more 18-1000 U /mg protein and when you factor NF-kB p65/p50 more than 1.0.
When activity of 26S proteasomes in the tumor over 18*1000 IU/mg protein 3 (13%) patients out of 23 was revealed recurrence of the process. Then, as if enzyme activity of less than 18*1000 IU/mg protein, at 16 (51%) patients out of 31 were diagnosed with recurrence. Disease-free survival in the first case amounted to 86.9 per cent, which was significantly higher compared with the second group of patients, where this indicator was equal to 49% (p<0,05). The sensitivity of the method for determining the activity of the 26S proteasome to assess the risk of recurrence of the tumor was 84%, specificity 57%, predictive value Polo is sustained fashion of the - 51%.
When the ratio of NF-kB p65/p50 above 1.0 in 3 (15%) patients out of 20 were revealed recurrence of bladder cancer, disease-free survival was equal to 84%. In 12 (60%) patients out of 20 when you factor NF-kB p65/p50 less than 1.0 were identified recurrence of the tumor after treatment. Relapse-free survival rate in this case was 40%. When comparing relapse-free survival in groups of patients with different threshold ratio NF-kB p65/p50 was statistically significant difference on this measure (p<0,05). The detection sensitivity of the ratio NF-kB p65/p50 relative risk of recurrence of the tumor was 80%, specificity of 68%, the positive predictive value of the result is 60%.
Example 1. Patient G., 60 years old, came in fsbi research Institute of Oncology WITH the RAMS, where the results of a comprehensive survey was diagnosed with bladder cancer T2N0M0. Received 2 courses of systemic chemotherapy in the neoadjuvant mode according to the standard scheme MVAC. In subsequent years this patient was performed TUR of the bladder and another 2 courses of adjuvant chemotherapy according to the above scheme. Histological examination of the surgical material is diagnosed with transitional cell bladder cancer moderate degree of differentiation. The activity of the 26S proteasome in the tumor is fabric made 17-1000 IU/mg protein, factor NF-kB p65/p50 was 0.9 units, which corresponded to an adverse prognosis in relation to risk of tumor recurrence. Dynamic observation of patients in the 14 months after treatment was diagnosed with a relapse, about which has been further treatment. And 20 months after treatment was detected relapse of the disease, and the patient was held cystectomy. The total period of observation was 3 years old.
Patient I., 55 years old, appealed to the state scientific research Institute of Oncology WITH the RAMS, where the results of a comprehensive survey was diagnosed with bladder cancer T3N0M0. Received 2 courses of systemic chemotherapy in the neoadjuvant mode according to the standard scheme MVAC. Further treatment was to conduct a TOUR of the bladder and another 2 courses of adjuvant chemotherapy according to the above scheme. Histological examination of the surgical material is diagnosed with transitional cell bladder cancer moderate degree of differentiation. The activity of 26S proteasomes in tumor tissue was 28-1000 IU/mg protein, factor NF-kB P65/P50 was equal to 1.2 units, which corresponded to a favorable prognosis in relation to risk of tumor recurrence. The period of dynamic observation of patients was 36 months. Data for the program is the financing of the disease was not detected.
Thus, the proposed method for predicting the outcome of muscle-invasive bladder cancer together with the main clinico-morphological parameters allows for a more accurate and informative to predict the occurrence of relapses, which gives the opportunity to adjust our tactics in patients.
Sources of information
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A method for predicting the outcome of bladder cancer in the study of enzymes of metabolism in the tumor, characterized in that the research activity of the 26S proteasome and the content of NF-kB p65 and p50; activity of 26S proteasomes more 18-1000 U/mg protein and when you factor NF-kB p65/p50 more than 1.0 predict a low likelihood of recurrence; activity of 26S proteasomes less 18-1000 U /mg protein and when you factor NF-kB p65/p50 less than 1.0 predict a high likelihood of tumor recurrence.
SUBSTANCE: invention refers to biotechnology, particularly to a microorganism test in various objects and media. The method provides conjugating electrically Fe0, MgFe2O4 or Fe3O4 labelled bacteria in an aqueous medium at specified parameters. Unbounded nanoparticles are separated with using a magnetic field, and a working electrode made of gold, platinum or graphite-bearing materials and having a surface pre-modified by antibodies specific to a bacterial strain to be tested is immersed into an analysed solution. The electrode is kept at the specified parameters to form an immune complex on its surface and washed in a buffer solution containing normal horse serum and Tween-20. The electrode is removed from the solution and placed into a cell containing LiClO4, dissolved in acetonitrile, dimethyl formamide or dimethyl sulphoxide; the bacterial count is determined by a value of analytical oxidation of the nanoparticles localised in the immune complex on the surface of the working electrode.
EFFECT: invention enables providing higher analysis sensitivity, better output and simplified analysis procedure.
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FIELD: medicine, pharmaceutics.
SUBSTANCE: invention refers to medicine, experimental and clinical pharmacology. The substance of the method: oxidised dextran is dissolved in a tris-acetate buffer solution with pH 5.0-5.5; the prepared solution is added with biotin hydrazide in a relation to oxidised dextran equal to 1:(20-25); the prepared solution is heated to 80-90°C and settled for 30-60 minutes.
EFFECT: preparing the high-quality biotinylated derivative of oxidised dextran providing higher sensitivity of its immune immunoassay in blood serum.
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SUBSTANCE: method involves blood serum analyzing for corticotropin-releasing hormone, substance P, neurokinin A and B, interleukin 6 and 8, relaxin and cortisol in females with full-term pregnancy by enzyme immunoassay at the first stage of labour in order to calculate a prognostic probability P by formula: P=(-0.0011229·CRH+(-0.15119·NA)+(-0.0071961·NB)+(-0.021071·IL6)+0.0032637·IL8+(-0.00085785·pF2α)+9.5773·REL+(-0.0098088·SP)+0.0058058·cort)·2.719. If the probability P is more than 0.51, a high risk of dysthyroidism requiring the urgent surgical labour completion is predicted.
EFFECT: higher-accurate prediction of dysthyroidism that enables the timely formation of a high-risk group, the differentiation of a dysthyroidism nature thereby selecting an adequate approach to the patient.
SUBSTANCE: if a pregnant woman suffers an aggravated cytomegaloviral infection, peripheral blood is examined to measure cytomegalovirus IgG antibody titres and histochemical product activity on carbonic anhydrate in red cells; if the IgG titre is 1:1600, and the activity on carbonic anhydrate reaction up to 0.015±0.002 standard units in 65.0±2.2% red cells, reducing oxyhaemoglobin up to 95.0±1.5%, threatening hemic hypoxia is stated.
EFFECT: method enables the preclinical assessment of the threatening hemic hypoxia.
SUBSTANCE: invention relates to field of medicine and can be used to estimate stability of membranes of peripheral blood erythrocytes in pregnant women with cytomegalovirus infection in the third trimester of gestation. Method is characterised by the following: titre of antibodies to cytomegalovirus, eximerisation factor in lipid bilayer and in protein-lipid layer are determined, microviscosity in membranes of erythrocytes is estimated and activity of adenosine triphosphate in erythrocytes is determined. If titre of antibodies to cyclomegalovirus is 1:1600, activity of adenosine triphosphate reduces to 0.40±0.008 mcmol/ml, microviscosity in membranes of erythrocytes is impaired, eximerisation factor in lipid bilayer reduces to 0.59±0.006 Fe/Fm, and in protein-lipid layer - to 0.70±0.004 Fe/Fm, threat to stability of membranes of erythrocytes is estimated.
EFFECT: estimation of stability of membranes of erythrocytes in peripheral blood in pregnant women with cytomegalovirus infection in the third trimester of gestation.
SUBSTANCE: method involves immunising with 5×107 microbial cells Vibrio cholerae 5879; 10 days later, a common pool of anticholeraic Ig is recovered from enteric washing liquid. That is combined with producing the primary enterocyte culture from intact mice. Thereafter, the doubly-diluted anticholeraic Ig are applied in an amount of 3 ml on a monolayer of erythrocytes formed in wells of culture panels. The latter are added with alive choleraic vibrions of the strain Vibrio cholerae 5879, 40 m.c. per 1 erythrocyte and incubated for 3 hours. Then, the panels are washed three times, fixed with ethanol for 20 minutes and stained by Romanowsky-Giemsa. Local humoral anticholeraic immunity is assessed under the microscope by the number of vibrions stained in dark blue and attached to one erythrocyte.
EFFECT: method enables assessing specific local humoral immunity and intensifying the anti-adhesive activity of the common pool of anticholeraic immunoglobulin.
4 cl, 2 ex, 2 tbl
SUBSTANCE: there is a suspension of live mononuclear cells of peripheral blood of an individual, applied onto a plastic dish, which comprises at least one microwell in a matrix of microwells. At the same time at least one microwell in the specified matrix of microwells comprises one cell in the volume of at least one nanolitre. The matrix of microwells is put in contact with a substrate, besides, the substrate is previously treated by at least two agents for detection, and each detection agent is bound with a secreted cytokine of the specified cell. They measure the level of each specified secreted cytokine on the specified substrate, at the same time the level corresponds to quantity of the specified secreted cytokine of the specified single cell. They determine speed of secretion of each specified secreted cytokine from the specified cell, thus determining characteristics of the specified secreted cytokines. They measure dynamic change of the specified characteristics of secreted cytokines.
EFFECT: development of immunological characteristics of diseases.
41 cl, 4 tbl, 21 dwg, 4 ex
SUBSTANCE: invention represents an instant diagnostic technique for acute intestinal infections (AIIs), involving detecting indication markers of the AII aetiology with the use of laboratory immunology tests, differing by the fact that the AII aetiology is stated in children of an early age category, preferentially in the newborn children; that is accompanied by measuring the concentration of cytokine, interleukin IL-10 in coprofiltrate and diagnosing chronic placental insufficiency (CPI); a probability (P) of the bacterial AII aetiology is calculated; the value P of more than 50% testifies to the bacterial AII aetiology, while the value P being less than 50% shows the absence of the bacterial AII aetiology, and enables considering the diagnostics second stage to be necessary, which implies measuring the concentration of cytokine, interleukin IL-4 in coprofiltrate; the time of latching the newborn child to the breast is established with considering the type of feeding; that is combined with calculating a probability (P) of the viral or viral-bacterial AII aetiology, with the value P of more than 50% testifying to the viral AII aetiology, while the value being less than 50% makes it possible to state the viral-bacterial AII aetiology.
EFFECT: more accurate diagnosing of the aetiology of acute intestinal infection and simplifying the diagnostic procedure.
SUBSTANCE: invention relates to field of medicine, namely to medical diagnostics, and describes method of qualitative differential diagnostics of benign and malignant neoplasms of lip mucosa by content of biomarkers in patient's oral fluid. Method includes sampling patient's oral fluid, centrifugation of patient's oral fluid at 3000 rev/min for 15 minutes, dilution of oral fluid with physiological solution in ratio 1:100, re-centrifugation at 3000 rev/min for 15 minutes, placement of ready for analysis material of patient's oral fluid into pan and carrying out standard method of enzyme immunoassay on die on the basis of monoclonal antibodies. As independent biomarkers selected are matrix metalloproteinase 8/matrix metalloproteinase 8 (MMP 8/MMP 8) and metalloproteinase 9/matrix metalloproteinase 9 (MMP 9/MMP 9). Method is characterised by high accuracy and sensitivity of differential express-diagnostics of benign and malignant neoplasms of lip mucosa, simplification of patient's preparation for sampling and storage of diagnostic material, possibility of obtaining result on the day of patient's visit, high probability of identification of benign and malignant neoplasms in patient's organism both at early and at later stages of the disease.
EFFECT: invention can be used in differential diagnostics of benign and malignant neoplasms of oral cavity organs under conditions of stomatological, oncologic and surgical medical institutions.
3 cl, 6 ex
SUBSTANCE: method for elution is implemented as follows: 10% tularemic, plague or brucellosis magnetic immune sorbent (MIS) 0.1ml is incubated with microbial suspensions of tularemic, plague or brucellosis agents for 30 min; the supernatant is removed; the MIS is incubated with an eluting solution 0.5 ml for 10 min; the eluate pH is reduced to the physiological value; an indirect hemagglutination reaction or a latex agglutination reaction is carried out.
EFFECT: method enables eluting the pathogen from a magnetic immobilised matrix qualitatively to be analysed by serological responses and keeping the antibody functionally active on the magnetic matrix for re-use.
3 ex, 1 tbl
SUBSTANCE: chemical elements are measured in human blood by X-ray fluorescence using synchrotron emission; a ratio of the total chemical elements with the prooxidant properties to the total chemical elements with the antioxidant properties is calculated by formula:
EFFECT: higher objectivity and reliability of assessing the antioxidant protection of the human body.
SUBSTANCE: photometric measurement of whole blood is taken in the visible and near IR wavelength region; spectral transmission factors of a whole blood cell are measured, while the primary haemoglobin derivative concentration values are determined by minimising a disparity The invention can be used to control the blood gas in resuscitation, toxicology, in intensive care to determine an effect on the haemoglobin composition of physiological, pathological and ecological factors, as well as in maternity hospitals, obstetric and child care hospitals to control haematological factors of newborn children and to monitor hyperbilirubinaemia.
EFFECT: more informative blood count.
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SUBSTANCE: invention refers to medicine and concerns a method for carrying out an immunochromatographic assay with a dissociating fluorescent tag, wherein complexes comprising molecules of an antigen or antigens, specific antibodies and tag molecules are formed on a membrane test strip. After the assay is completed, the tag is extracted from the working membrane of the test strip, and a fluorescent signal is measured in a liquid volume.
EFFECT: method according to the invention provides creating the optimum conditions for tag fluorescence, as well as eliminating the harmful influence of the membrane which is impermeable to decrease an intensity of both exciting and emitted light.
1 dwg, 1 ex
SUBSTANCE: method involves: peripheral blood is examined for cytomegalovirus titre, blood pH and oxyhaemoglobin on a biochemical analyser; if observing an increase of the cytomegalovirus titre to 1:1600, a decrease of blood pH to 7.25±0.03, and oxyhaemoglobin - to 90.15±0.35% with the reference of 95.20%, a threatening formation of hemic hypoxia is stated.
EFFECT: higher assessment accuracy.
SUBSTANCE: carried out are: determination of the serum concentration of a factor of a tumour necrosis alpha (TNF-α), evaluation of a relative content of Th17-lymphocytes (%CD3+CD4+CD161+ ot CD3+CD4+) and an activity of succinatedehydrogenase in a population of regulatory T-cells (CD3+CD4+CD127low) (SDH-Treg) before and on the following day after infliximab introduction. A prognostic factor of the medicine efficiency K is calculated by formula. If the factor value is K<0.5, a minimal therapeutic effect of infliximab is predicted. When K=0.5-1.5, a moderate therapeutic effect, which demands correction of the medicine introduction plan, is predicted. If K is from over 1.5 to 2.5, a positive effect from introduction of infliximab is predicted.
EFFECT: possibility to predict long-term, not less than 1 year, effect from the infliximab therapy at any stage of therapy.
4 dwg, 2 tbl, 3 ex
SUBSTANCE: invention relates to analytical chemistry, in particular to aspects, related to methods of the nonenzymatic determination of the presence or a quantity of carbohydrates in a sample, and describes a method, device and set for the sample analysis to determine the presence or a quantity of an analysed substance, in particular a carbohydrate, in particular, sugar, in the sample with the application of fabric. The method includes the application of the sample on a synthetic fabric; chemical modification of the said carbohydrate, present in the sample, with an oxidising agent with a sufficient oxidising potential in order to cleave the carbohydrate between two hydroxyl groups; inactivation of the said oxidising agent, which would prevent the identification of the chemically modified carbohydrate; identification of the presence or the quantity of the said chemically modified carbohydrate by means of a copper-containing compound to obtain the visible change of colour.
EFFECT: invention can be used for the determination of the presence of a carbohydrate on the surface to indicate the surface contamination, in particular contamination with a substance, promoting the microbial growth.
20 cl, 2 dwg, 7 tbl, 4 ex
SUBSTANCE: invention relates to medicine, namely to method of determining expression of inflammatory process in case of osteoarthrosis. Essence of method consists in the following: carried out is luminol-dependent iron-induced chemi-luminescence of model system, which has the following composition: 2.72 g of KH2PO4, 7.82 g of KCl, 1.5 g of sodium citrate C6H8O7Na3*5,5H2O per 1 liter of distilled water, pH 7.45 with 0.2 ml of 10-5 M luminol solution, after that, intensity of model system luminescence is determined in presence of synovial fluid before and after addition of synovial fluid. Degree of suppression of intensity of model system chemi-luminescence is calculated by formula. If its value is from 1.71 to 6.48%, high activity of inflammatory process is determined, if its value is from 6.49 to 21.55%, medium activity is determined, from 21.56 to 55.46 - small activity.
EFFECT: application of the method makes it possible to reduce time of determination and increases accuracy of assessment of inflammatory process degree in case of osteoarthrosis.
1 tbl, 1 dwg, 3 ex
SUBSTANCE: diagnostic test element (110) for the detection of an analyte in a sample (126) of a body fluid, in particular whole blood with the volume not less than 2 microlitres, contains a test field (116) with a reagent-indicator, where the reagent-indicator is capable of feeling a detected change, in particular an optic change, in case of the analyte presence. The test field (116) includes a detector layer (118), containing the reagent-indicator, where the detector layer contains particles (137). 90% of all particles (137) of the indicator layer (118) have the actual size smaller than 10 microns. The diagnostic test element (110) contains a bearing element (112), which has a transparent region (114), where the test field (116) with its detection side (120) is partially applied on the transparent region (114).
EFFECT: detected change is an optically detectable change, where an optic detector with spatial resolution is applied for the detection of the detected change.
9 cl, 8 dwg, 1 tbl, 2 ex
SUBSTANCE: invention relates to immunology and medical diagnostics and represents method of carrying out immunochromatographic analysis for serodiagnostics. Claimed invention is intended for immunochromatographic determination of antibodies to causative agents of infectious diseases or other antigens, for instance, allergens, in liquid samples. Claimed method of identifying antibodies is characterised by the following: solution for sample dilution contains specific antibodies against test strip of antigen (or antigens) immobilised in analytic zone. Used concentration of antibodies in diluting solution is lower than lower limit of determination of antibodies by said method with application of diluting solution, which does not contain specific antibodies, which results coloration of analytic zone of test strip is not observed if specific antibodies are absent in sample. If sample contains specific antibodies, presence of additional quantity of specific antibodies in diluting solution results in enhancement of intensity of coloration of test strip analytic zone.
EFFECT: application of invention makes it possible to register total concentration of specific antibodies in sample and in diluting solution, which results in reduction of limit of analysis identification due to displacement of working range of concentrations, determined by test, into the area of lower values.
3 dwg, 1 ex
SUBSTANCE: invention refers to medicine and represents a method for accelerated staining of histological specimen for identifying tuberculosis mycobacteria involving the preparation of the histological specimen according to a common technique, the Ziehl-Nelsen staining differing by the fact that the standard preparation of the specimen for staining is followed by a de-waxing procedure by placing them successfully into xylol and alcohols of the decreasing concentration for 5 minutes in a thermostat at 54°C and then placing into water, then in a 1% periodic acid for 2 minutes and washing in flowing water for 10 seconds; thereafter, Ziehl carbol-fuxine is poured on a section coated with a filter paper and heated on an alcohol burner lamp until evaporating for 2 minutes; the stain is left on the section after the heating procedure is completed, for 3-5 minutes; the filter paper is removed, and the sections are washed in water, differentiated in a 1% acid buffer, washed in main water for 1 minute; thereafter Leffler methylene blue is placed on the section and heated on the alcohol burner lamp until evaporating, washed in water, differentiated in a 1% acid buffer and washed with a plenty of water, de-watered in alcohols of the increasing concentration for 1 minute, placed in xylol and balm.
EFFECT: developing the method for the accelerated stain of the histological specimen.
FIELD: medicine, analytical biochemistry.
SUBSTANCE: invention relates to laboratory methods of investigations. Method involves sampling specimen from patient to be inspected, extraction of serotonin and histamine from a specimen, chromatography of extract and determination of concentration of serotonin and histamine by the fluorescence intensity value. Saliva is used as biological fluid. Saliva by volume 1 ml is extracted with 4 ml of 1 N hydrochloric acid solution, 2 g of anhydrous potassium carbonate and 5 ml of mixture of butanol and chloroform in the ratio 3:2 are added, extract is shaken up and centrifuged. Organic phase (4 ml) is sucked off from extract and passed through chromatography column (diameter is 3 mm, height is 16 mm) filled with ion-exchange resin KB-4 or KB-4P-2 or Bio Rex-70 in H+-form, size of granules is 0.1 ± 0.02 mm. Histamine is eluted with 4 ml of 0.1 N hydrochloric acid at the rate of eluting solution 0.4 ml/min. Histamine concentration is determined by reaction with ortho-phthalic aldehyde dissolved in ethanol. Serotonin concentration is determined by reaction with ninhydrin in organic passed through column. Method provides assaying the saliva concentration of serotonin and histamine with high precision.
EFFECT: improved assay method.