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Biomedical subjects

A P Osipov

Publications and source records attributed to A P Osipov.

At least 19 recordsLinked to original sources
PubMed2002

Analysis of bispecific monoclonal antibody binding to immobilized antigens using an optical biosensor.

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.

Antibodies, Monoclonal↗
PubMed2001

The comparison of the ability of monoclonal antibodies directed to different proteins (human IgG, human myoglobin and HRP) and bispecific antibodies derived thereof to bind antigens immobilized on a surface of a solid phase.

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.

Adsorption↗
PubMed1996

Chemiluminescent immunoenzyme biosensor with a thin-layer flow-through cell. Application for study of a real-time bimolecular antigen-antibody interaction.

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.

Antigen-Antibody Reactions↗
PubMed1993

New immunoassay technique using antibody immobilized on a membrane and a flow cuvette as reaction vessel.

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.

Flow Injection Analysis↗
PubMed1992

[The treatment of acute appendicitis].

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.

Acute Disease↗
PubMed1989

An investigation on the catalytic mechanism of enhanced chemiluminescence: immunochemical applications of this reaction.

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.

Haptens↗
PubMed1989

[Completion of choledochotomy].

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.

Adult↗
PubMed1988

[The problem of relaparotomy in emergency surgery].

For the recent 15 years the authors have made 6220 operations on organs of the abdominal cavity, among them there were 111 relaparotomies in 96 patients (1.78%). An atypical course of the postoperative period was an early symptom of unfavorable state of the stomach. Thirty patients died (31.25%). The authors consider that a delayed first operation in urgent surgery is one of the main causes resulting in relaparotomy and appeal to properly perform the sanitary-instructive work, dispensary observation and planned sanitation of surgical patients.

Abdomen, Acute↗
PubMed1987

[Immunoenzyme analysis of human IgG in the kinetic mode].

A kinetic sandwich enzyme-linked immunosorbent assay for the detection of human IgG (used as a model antigen) has been developed. Rabbit antihuman IgG has been used both for coating polystyrene microtitration plates and for the preparation of the conjugate of anti-human IgG with horse-radish peroxidase. The kinetics of the reaction of the antigen and the antibody-peroxidase conjugate with the reagents immobilized on polystyrene plates has been studied. The assay is optimized with respect to its sensitivity and the duration of intervals for every stage of the assay. The optimal time of the assay is about 10-15 minutes. The correlation between sensitivity and the duration of every stage of the assay has been established.

Adsorption↗