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1.
Talanta ; 175: 143-149, 2017 Dec 01.
Article in English | MEDLINE | ID: mdl-28841971

ABSTRACT

New sampling format of whole cows' milk in strip-dried form was proposed. Few methodological issues of whole milk progesterone ELISA using samples dried on a membrane carrier in a form of strip were investigated and optimized: width of a strip, shape of punched/cut-off part of membrane, sample application method. It was shown that distribution of the hormone along narrow strip was even except the initial part of a strip (the first 0.5 × 0.5cm piece) where recovered concentration of progesterone was higher. Storage stability of progesterone in strip-dried whole cows' milk samples at 4°C, ambient temperature, 37°C and 60°C was investigated. Rising of the detected progesterone concentration over storage period at elevated temperatures was observed predominantly in milk samples with low hormone concentration (from non-pregnant cows). Strip-dried whole milk samples can be used for collection, transportation, storage and ELISA analysis of progesterone level which is correlated with reproductive status of cows.


Subject(s)
Desiccation/instrumentation , Enzyme-Linked Immunosorbent Assay/instrumentation , Milk/chemistry , Progesterone/analysis , Specimen Handling/instrumentation , Animals , Cattle , Female , Reagent Strips/analysis , Temperature
2.
Talanta ; 132: 685-9, 2015 Jan.
Article in English | MEDLINE | ID: mdl-25476365

ABSTRACT

New rapid method of lateral flow enzyme immunoassay (LFEIA) for progesterone detection in whole cows' milk was developed. The test system utilized horseradish peroxidase as a label along with the substrate solution containing 3,3',5,5'-tetramethylbenzidine and dextran sulfate to obtain an insoluble blue colored product of the enzyme reaction on a surface of analytical membrane (test and control lines). Several aspects of LFEIA were optimized: time of the signal detection, membrane materials and assay conditions. Resulting competitive LFEIA can be performed within 15 minutes with the limit of progesterone detection of 0.8 ng/ml. Progesterone concentration in whole milk samples was determined by LFEIA and enzyme-linked immunosorbent assay (ELISA). The results obtained were in good correlation (R=0.97, n=46). Thus new sensitive LFEIA can be successfully used for on-site monitoring of oestrus status of cows' reproductive system and for early none-pregnancy detection. The method is fast, easy to perform and needs no preliminary sample preparation.


Subject(s)
Immunoenzyme Techniques/methods , Milk/chemistry , Progesterone/analysis , Animals , Benzidines/chemistry , Dextran Sulfate/chemistry , Horseradish Peroxidase/chemistry , Immunoenzyme Techniques/instrumentation , Limit of Detection
3.
Biochemistry (Mosc) ; 74(9): 986-93, 2009 Sep.
Article in English | MEDLINE | ID: mdl-19916909

ABSTRACT

A highly sensitive express immunochromatography method for molecular diagnosis of plant virus infections was elaborated on the example of a model object - tobacco mosaic virus (TMV). The analysis time does not exceed 5 min, and the lower limit of TMV detection in non-clarified leaf extract (2-4 ng/ml) is comparable with the sensitivity of the enzyme-linked immunosorbent assay of the virus. A single measurement requires 0.1-0.2 ml tested solution (extract from 10-20 mg of leaf material). The sensitivity of TMV determination in the leaf tissue extract was increased by more than one order of magnitude using signal enhancement by silver and is 0.1 ng/ml. In this case, analysis time did not exceed 25 min. The simplicity of this method makes it especially convenient in express diagnosis of numerous analyzed specimens. The prototype of a diagnostic kit for serial analyses of plant viral infections both in laboratory and field conditions was elaborated.


Subject(s)
Chromatography, DEAE-Cellulose/methods , Plant Diseases/virology , Tobacco Mosaic Virus/isolation & purification , Antibodies, Viral/analysis , Enzyme-Linked Immunosorbent Assay , Metal Nanoparticles , Tobacco Mosaic Virus/immunology
4.
Biochemistry (Mosc) ; 67(12): 1356-65, 2002 Dec.
Article in English | MEDLINE | ID: mdl-12600264

ABSTRACT

The interaction between two different monoclonal antibodies (Mabs) and their corresponding bispecific antibodies (Babs) with immobilized antigens was investigated using an optical biosensor (IAsys). The analyzed panel of affinity-purified antibodies included two parental Mabs (one of which was specific to human IgG (hIgG), and another one to horseradish peroxidase (HRP)), as well as Babs derived thereof (anti-hIgG/HRP). Babs resulting from the fusion of parental hybridomas bear two antigen-binding sites toward two different antigens and thus may interact with immobilized antigen through only one antigen-binding site (monovalently). Using an IAsys biosensor this study shows that the bivalent binding of Mabs predominates over the monovalent binding with immobilized HRP, whereas anti-hIgG parental Mabs were bound monovalently to the immobilized hIgG. The observed equilibrium association constant (K(ass)) values obtained in our last work [1] by solid-phase radioimmunoassay are consistent with those constants obtained by IAsys. The K(ass) of anti-HRP Mabs was about 50 times higher than that of anti-HRP shoulder of Babs. The dissociation rate constant (k(diss)) for anti-HRP shoulder of Babs was 21 times higher than k(diss) for anti-HRP Mabs. The comparison of the kinetic parameters for bivalent anti-HRP Mabs and Babs derived from anti-Mb/HRP and anti-hIgG/HRP, allowed to calculate that 95% of bound anti-HRP Mabs are bivalently linked with immobilized HRP, whereas only 5% of bound anti-HRP Mabs are monovalently linked. In general, the data obtained indicate that Babs bearing an enzyme-binding site may not be efficiently used instead of traditional antibody-enzyme conjugates in the case of binding of bivalent Mabs.


Subject(s)
Antibodies, Monoclonal/chemistry , Biosensing Techniques/instrumentation , Biosensing Techniques/methods , Antigen-Antibody Reactions , Antigens/chemistry , Binding Sites , Dose-Response Relationship, Drug , Humans , Hybridomas/immunology , Immunoglobulin G/chemistry , Immunoglobulin G/metabolism , Kinetics , Time Factors
5.
Bioorg Khim ; 27(4): 265-74, 2001.
Article in Russian | MEDLINE | ID: mdl-11558260

ABSTRACT

The ability of bispecific antibodies (Babs) formed by fusion of hybridomas and parent monoclonal antibodies (Mabs) to interact with the solid phase-adsorbed antigens was studied. Mabs specific to the three different antigens [horseradish peroxidase (HRP), human IgG (hIgG), and human myoglobin (Mb)] as well as Babs with the double specificity [antimyoglobin/antiperoxidase (anti-Mb/HRP) and anti-hIgG/antiperoxidase (anti-hIgG/HRP)] were used. It was shown by radioimmunological and immunoenzyme assays that parent Mabs bind to solid phase-adsorbed antigens considerably more effectively than Babs. The observed equilibrium binding constant (Ka) of antiperoxidase parental Mabs to immobilized HRP is 21 and 38 times higher than Ka for Babs binding sites (anti-Mb/HRP and anti-hIgG/HRP, respectively) to peroxidase. It was calculated that about 90-95% of all bound parental antiperoxidase Mabs were associated with immobilized HRP bivalently, and only about 5-10% were bound monovalently. On the contrary, parental Mabs against hIgG bind to the sorbed antigen essentially only monovalently. It was also shown that the avidity of anti-Mb/HRP Babs significantly increased when two antigens, Mb and HRP, were simultaneously adsorbed on the solid phase. These data imply that Babs bearing an enzyme-binding site (for example, binding to HRP) cannot be more effective than standard conjugates (e.g., enzyme-conjugated antibodies) in heterogeneous noncompetitive immunoassays.


Subject(s)
Antibodies, Bispecific/immunology , Antigen-Antibody Complex/immunology , Horseradish Peroxidase/immunology , Immunoglobulin G/immunology , Myoglobin/immunology , Animals , Binding Sites, Antibody/immunology , Enzyme-Linked Immunosorbent Assay/methods , Humans , Mice , Radioimmunoassay/methods , Sensitivity and Specificity
6.
Clin Chim Acta ; 309(1): 57-71, 2001 Jul 05.
Article in English | MEDLINE | ID: mdl-11408007

ABSTRACT

BACKGROUND: Bindings of mouse monoclonal antibodies (mAbs) and affinity purified bispecific antibodies (bAbs), derived thereof, to antigens adsorbed on immunoplates have been compared, using ELISA and RIA methods. METHODS: The analysed panel of antibodies included mAbs specific to human myoglobin (Mb), human IgG (hIgG) and horseradish peroxidase (HRP) and biologically produced bAbs with double specificity to Mb and HRP, and to hIgG and HRP. RESULTS: The degree of difference between different mAbs and corresponding bAbs varied markedly from antibody to antibody, depending on whether the parental mAbs could bind immobilized antigens bivalently. The observed equilibrium binding constant (K(obs)) for anti-HRP mAbs was 21-38 times higher that of anti-HRP site of bAbs (anti-hIgG/HRP or anti-Mb/HRP, respectively), due to bivalent binding of mAbs. Anti-Mb mAbs also bound bivalently with immobilized Mb. On the contrary, anti-hIgG mAbs bound monovalently with immobilized hIgG in the same conditions. The avidity of anti-Mb/HRP bAbs increased, if both antigens were simultaneously adsorbed on a solid phase. CONCLUSIONS: The obtained data indicate that the use of bAbs in heterogeneous immunoassays instead of traditional mAb-enzyme conjugates hardly can provide the significant gain in assay performance if parental mAbs bind bivalently.


Subject(s)
Antibodies, Bispecific/metabolism , Antibodies, Monoclonal/metabolism , Antigen-Antibody Reactions/immunology , Horseradish Peroxidase/metabolism , Immunoglobulin G/metabolism , Myoglobin/metabolism , Adsorption , Animals , Antibodies, Bispecific/immunology , Antibodies, Monoclonal/immunology , Antibody Affinity/immunology , Antigens/immunology , Antigens/metabolism , Binding Sites, Antibody/immunology , Enzyme-Linked Immunosorbent Assay , Horseradish Peroxidase/immunology , Humans , Hybridomas , Immobilized Proteins , Immunoglobulin G/immunology , Mice , Myoglobin/immunology , Radioimmunoassay
8.
Biosens Bioelectron ; 11(9): 881-7, 1996.
Article in English | MEDLINE | ID: mdl-8688165

ABSTRACT

A simple flow enzyme system for real-time continuous monitoring of interaction of biological molecules has been developed. It relies upon a thin-layer flow-through cell placed directly into the measuring compartment of the luminometer. One ligand (antibody) is immobilized on the inner surfaces of the flow cuvette, and a second ligand (antigen) labeled with a peroxidase molecule moves through the flow cell. The quantity of the complex on the surface of the cell may be monitored by measurement intensity of chemiluminescence in the reaction of peroxidase label with substrates (p-iodophenol, luminol and hydrogen peroxide). In such a way one can detect in a real-time regime the kinetics of association (or dissociation) of the complex labeled ligand-receptor on the surface of the cuvette. Due to the small thickness of the flow cell the diffusion limitations of interaction for two kinds of biomolecules (soluble and immobilized) are negligible, so the resulting intensity of chemiluminescent signal reflects the kinetics of interaction between soluble and immobilized components. The system may be successfully used for molecular recognition studies, analyzing the kinetics of bimolecular interaction and for concentration determination.


Subject(s)
Antigen-Antibody Reactions , Biosensing Techniques , Flow Injection Analysis , Immunoenzyme Techniques , Luminescent Measurements
9.
J Immunol Methods ; 157(1-2): 11-7, 1993 Jan 04.
Article in English | MEDLINE | ID: mdl-8423353

ABSTRACT

Immunoenzymatic detection systems have been developed using human IgG as a model antigen. A membrane with covalently immobilized specific antibodies was placed into a specially constructed ultranarrow flow cuvette and solutions containing the antigen and antibody-peroxidase conjugate were then successively passed through the flow capillary cell. After washing, the membrane was placed into the substrate solution and the intensity of developed colour on the membrane was recorded visually or by a reflection spectrophotometer. The lower detection limit was about 5 x 10(-11) M and the overall analysis time was 10 min. Photoimmobilization was used to immobilize the antibody and thereby permitting control of the protein surface concentration on the membrane as well as the dimensions and shape of the activated region.


Subject(s)
Immunoenzyme Techniques , Flow Injection Analysis , Humans , Immunoglobulin G/analysis
10.
Khirurgiia (Mosk) ; (2): 54-7, 1992 Feb.
Article in Russian | MEDLINE | ID: mdl-1388214

ABSTRACT

The article deals with the experience in the treatment of 3,368 patients with acute appendicitis from 1979 to 1989. Among them were 2,398 adults and 976 children. Complications developed in 190 (5.6%) patients, 5 (0.15%) patients died. The mortality rate among children was 0.3%. Complications and deaths were connected with neglected forms of the disease. 695 (20.6%) patients were admitted later than 24 hours after the onset of the disease. The patients diagnosed the disease themselves and resorted to self-treatment. Health education measures were poorly effective; it is suggested that this work should be started from early childhood. A permanently functioning seminar on acute surgical diseases was organized at the institution for doctors not practicing in surgery, but diagnostic errors were made at the prehospital stage in 117 cases by district therapists, pediatricians, and the staff of emergency aid teams. There were also cases in which operation was undertaken late after admission to the hospital and cases of hyperdiagnosis. The late-term results were studied in 109 patients without morphological changes in the removed vermiform processes, in 51.4% of them other diseases were encountered in the long-term period.


Subject(s)
Appendicitis/surgery , Acute Disease , Age Factors , Appendectomy/statistics & numerical data , Appendicitis/diagnosis , Appendicitis/epidemiology , Appendicitis/mortality , Diagnostic Errors , Humans , Patient Education as Topic , Postoperative Complications/epidemiology , Quality of Health Care , Retrospective Studies , Russia/epidemiology , Sex Factors
14.
Med Tekh ; (4): 8-13, 1990.
Article in Russian | MEDLINE | ID: mdl-2215181

ABSTRACT

The authors review the thermodynamic bases and physiological limitations of the applicability of thermal engines for driving artificial heart ventricles. Show that the thermodynamic characteristics of Stirling and Brighton cycles do not make it possible to effectively use cycle-based engines in the artificial heart. A steam engine operating in accordance with the Rankine cycle may be regarded as an optimum type engine for that purpose. Demonstrate that according to the rules of physiology, use should be made of a separate driving of artificial heart ventricles by two independently operating steam engines. Provide the characteristics of the Soviet artificial heart "MIKRON" acceptable for implantation into the orthotopic position.


Subject(s)
Heart, Artificial , Equipment Design , Humans , Medical Laboratory Science , Thermodynamics
17.
Stomatologiia (Mosk) ; 68(4): 28-9, 1989.
Article in Russian | MEDLINE | ID: mdl-2530653

ABSTRACT

The study of cicatrization after the teeth extraction in workers of viscous rayon industry which had had a long contact with sulfuric carbon revealed a lower regeneration potential and increased complication incidence as compared to control patient group. Preventive measures are designed providing for control check-ups 3 to 5 days after the intervention and the use of rehabilitation period in presurgical preparation of the patients.


Subject(s)
Alveolar Process/drug effects , Cellulose/adverse effects , Textile Industry , Textiles , Tooth Extraction , Wound Healing/drug effects , Alveolar Process/physiology , Dry Socket/physiopathology , Female , Humans , Male , Middle Aged , Russia , Time Factors , Wound Healing/physiology
18.
J Biolumin Chemilumin ; 4(1): 164-76, 1989 Jul.
Article in English | MEDLINE | ID: mdl-2678913

ABSTRACT

The mechanism of peroxidase-catalysed oxidation of luminol by H2O2 was studied. The stopped-flow technique was used to measure the rate constants for the reactions between the oxidized forms of peroxidase with luminol and the following substrates: p-iodophenol, p-bromophenol, p-clorophenol, o-iodophenol, m-iodophenol, luciferin, and 2-iodo-6-hydroxybenzothiazole. The correlation between kinetic parameters and the degree of enhancement was established. The effect of charged synthetic polymers and specific antibodies on the peroxidase activity in the enhanced chemiluminescent reaction was also studied. The close approach of an effector molecule to the active site of the enzyme was found to inhibit the enhanced chemiluminescent reaction. Novel homogeneous methods of luminescent immunoassay (LIA) for (1) antibodies to insulin, (2) insulin and (3) antibodies to trinitrophenyl group are proposed on the basis of regulatory facilities of the enhanced chemiluminescent reaction. Based on the enhanced chemiluminescent reaction a peroxidase flow-injection assay was developed and successfully tested in the flow-injection enzyme immunoassays for human IgG and for thyroxin (T4). The immunoassay proposed has a detection limit of 10(-9) M for IgG and 10(-11) M for T4, the overall time of the assay being 5-15 min.


Subject(s)
Immunochemistry , Immunoenzyme Techniques , Luminescent Measurements , Haptens , Horseradish Peroxidase , Humans , Immunoglobulin G/analysis , Insulin/analysis , Insulin Antibodies/analysis , Kinetics , Luminol , Oxidation-Reduction , Substrate Specificity , Thyroxine/analysis
20.
Vestn Khir Im I I Grek ; 142(2): 27-30, 1989 Feb.
Article in Russian | MEDLINE | ID: mdl-2728232

ABSTRACT

An analysis of 144 operations using external drainage of the common bile duct per 926 cholecystectomies and using T-shaped drains (in 90 patients drains of vulcanized latex [Revultex] proposed by the authors were used) has brought the authors to a conclusion that the temporary external drainage of the common bile duct is the method of choice for the completion of choledochotomy, when it is not possible to carefully visually control the hepatocholedochus lumen. The T-shaped cast latex drains are thought to be most adequate for the operations.


Subject(s)
Cholecystectomy/methods , Choledochostomy/methods , Cholelithiasis/surgery , Cholestasis, Extrahepatic/surgery , Drainage/methods , Gallstones/surgery , Adult , Aged , Choledochostomy/instrumentation , Cholelithiasis/complications , Drainage/instrumentation , Female , Gallstones/complications , Humans , Latex , Male , Middle Aged
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