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Biomedical subjects

A P Savitskiĭ

Publications and source records attributed to A P Savitskiĭ.

At least 19 recordsLinked to original sources
PubMed1999

[Luminescent ultramicroanalysis for the simultaneous determination of biological and chemical substances: the methodology and the ways for solution].

The next revolution in in-vitro bioaffinity assays will be associated with the minimization of the liquid phase of an experiment and with the extremely rapid detection of a large number of samples. The ultra detection unit is one of the essential elements of such devices. The fluorescence detection systems are most promising. The fluorescence detection limit of europium ions is as low as 10(-15) M in the complex with beta-diketone. The authors' approach to designing new highly effective chelates is to synthesize a stable chelate wherein several molecules of beta-diketones are properly oriented and chemically fixed. The molecular mechanic calculation of three dimensional structure of different Eu chelates should be used in optimizing the strategy of chemical synthesis of new compounds. New chelates can be efficiently used for the development of a new approach to measuring distances equal to over 10 nm in the biological systems. Pesticides also represent challenging targets because the levels of compounds are to be detects in minor concentrations, thus the time-resolution fluorescence assay may have some advantages due to its high sensitivity. DNA-based microbiochips are the most promising area in the practical use of a new chelate. A novel europium chelate is a key compound for the development of new methods for studying biological and chemical objects, as well as new diagnostic systems based on a basically new instrumental base that is characterized by extremely high miniaturization.

Chelating Agents↗
PubMed1998

[Synthesis of phthalocyanine conjugates with monoclonal antibodies in AOT/n-octane reversed micelles and in water-organic solvent mixtures].

Conjugates of cobalt and aluminum phthalocyanines with monoclonal antibodies were synthesized using an AOT/n-octane reversed micellar system or water-organic mixtures with a low content of organic solvent as media. The effect of the degree of hydration of the micelles and the concentration of phthalocyanines on the composition of conjugates was studied. The immune activity of the resulting conjugates in comparison to that of native antibodies was evaluated. The catalytic activity of free cobalt phthalocyanines and their antibody conjugates was studied in the reaction of ascorbic acid oxidation.

Aluminum↗
PubMed1990

[Photosensitized formation of singlet oxygen in water solutions of covalent porphyrin-antibody conjugates].

The effective photogeneration of singlet molecular oxygen (1O2) by porphyrins (coproporphyrin I; 2,4-bi (alpha-methoxyethyl) deuteroporphyrin IX and cyclopanten-coproporphyrin I) conjugated with antibodies (mouse monoclonal IgG and IgM and human gamma-globulin) have been observed with the direct luminescence method of 1O2 detection. Absolute quantum yields of 1O2 formation by the conjugates have been determined. The data suggest that porphyrin-antibody conjugates are promising for the use as drugs in photodynamic tumor treatment.

Animals↗
PubMedDate not supplied

[Key amino acid residues responsible for the color of green and yellow fluorescent proteins from the coral polyp Zoanthus].

Site-directed mutagenesis was used to study the structural basis of color diversity of fluorescent proteins by the example of two closely related proteins from one organism (coral polyp Zoanthus sp.), one of which produces green and the other, yellow fluorescence. As a result, the following conversions of emission colors were performed: from yellow to green, from yellow to a dual color (yellow and green), and from green to yellow. The saltatory character of the spectral transitions and the manifestation of the dual-color fluorescence suggest that chemically different fluorophores are responsible for the green and yellow fluorescence. The simultaneous presence of three residues, Gly63, Lys65, and Asp68, is necessary for the efficient formation of the yellow rather than green fluorophore. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2002, vol. 28, no. 4; see also http://www.maik.ru.

Amino Acid Sequence↗
PubMedDate not supplied

[Lanthanide fluorescent immune analysis for determination of hormone thyroxine in dry blood spots].

The determination of thyroxin (T4) is a basic confirmative test for congenital hypothyroidism. 70% cases of this period inborn disease are taking asymptomatic course. We developed immunofluorescent T4-assay in dried blood spots with anti-T4 monoclonal antibody and europium chelate (dianhydride of diethylentriaminipentaacetic acid (DA-DTPA)). This method requires DELFIA Plate Fluorometer 1232 (Wallac, Finland) or its sub modifications. Panel of monoclonal antibodies for T4 has been obtained. Type/subtype (IgG2b, IgG1), affinity constants (10(7)-10(8) M-1), cross reactivity to homologous structures (0-2%) were determined. Stable clones with high affinity and viability were selected for the development of the assay. Conjugates of thyroxin and europium chelate were synthesized. Inclusion of sodium salicylate (2 mg/ml) and EDTA (2 mM) into the buffer reduced the nonspecific signal. Limit for T4 detection (T4 standarts) was 10 nM with not more than 15% variation coefficient. Accuracy was estimated by Bio-Rad Lipochek Immunoassay Plus Control Kit. Obtained results were within control confidence interval.

Antibodies, Monoclonal↗
PubMedDate not supplied

[Fluoroimmunometric method of determining digitoxin].

Digitoxin at concentrations up to 5 x 10(-10) M (therapeutic concentrations are 2 x 10(-8) M) can be reliable measured by a fluoroimmunometric method with coproporphyrin as a tracer. The use of nylon filters with immobilized digitoxin to remove an excess of labelled antibodies increases reliability and reduces the time of measurements, and thus simplifies the assay.

Chromatography, Ion Exchange↗
PubMedDate not supplied

[Influenza viruses studied by fluorescent immunoassay with temporary resolution].

Monoclonal antibodies, epidemic strains of influenza A and B viruses, conjugates were studied by fluorescence immunoassay with temporary resolution using equipment of different companies. Differences and specificity of monoclonal antibodies to influenza A and B viruses were demonstrated. The highest sensitivity of the method with the test system used was determined, being 20 ng/ml of viral protein. The method was shown to be useful for influenza virus detection both in solutions containing purified virions and in virus-containing allantoic preparations. It was established that virological studies could be carried out using conjugates and knock-down microplates of the national make alongside with foreign ones.

Antibodies, Monoclonal↗
PubMedDate not supplied

[Chromatography of water-soluble porphyrins and their conjugates with proteins].

The chromatographic behaviour of tetra- and octacarboxylic porphyrins and tetra-(p-sulphophenyl)-porphin was being studied by HPLC using Protein Pak and TSK-type gel permeation columns. The mobility of the porphyrins on the carriers depended on the ionic strength of the eluating buffer Tris-HCl. The conditions for separation of the porphyrins and their protein conjugates were optimized. The separation was performed under mild conditions with minimum denaturation of conjugates. The purification technique can be used to separate porphyrin conjugates with various proteins. The technique can be also used in conventional column chromatography on Toyopearl supports.

Chromatography, High Pressure Liquid↗
PubMedDate not supplied

[Fluorescent immunoanalysis. Synthesis, spectro-fluorescent and immunologic properties of conjugates of coproporphyrin I with antibodies].

The synthesis of protein conjugates with the new high-efficient fluorescent labile coproporphyrin-I was optimized. A number of conjugates of monoclonal antibodies with different coproporphyrin-I content were synthesized, and their spectral properties were studied in water and micellar solutions, i.e. adsorption, excitation and emission spectra, fluorescence quantum yields, fluorescence pH-dependences. The binding constants of coproporphyrin-I and its protein conjugates with serum albumin were determined. The antibodies labelled with coproporphyrin-I retain the functional activity and photochemically stable in water solutions. The sensitivity of fluorometric detection of coproporphyrin-I and its conjugates with proteins is more than 10 times greater than in case of FITC.

Animals↗
PubMedDate not supplied

[A method of fluorescence immunoassay with temporal resolution and the use of europium label and polymeric complexon].

Recently developed solid-phase immunofluorescent assay with temporal distinction involved a newly produced procedure for binding of of the boron group label with antibodies via modified polymer, which is able to bind with antibodies across the avidin bridge. The assay developed is more simple than the widely used procedures, it exhibits high sensitivity being superior as compared with a corresponding immunoenzyme assay by a decimal order.

Animals↗
PubMedDate not supplied

[Fluorescent immunoassay of thyroxine in blood serum].

A method of solid-phase fluoroimmunoassay of thyroxine (T4) on polystyrene 96-microtiter plates using coproporphyrin III-labeled antibodies was developed; 0.02 ml of blood serum without preliminary treatment was required. Thimerosal was added to lessen the T4-binding effect of the serum on the results of T4-binding effect of the serum on the results of the immunoassay. A calibration curve was obtained to determine T4 covering the entire physiological range of concentrations (from 3 to 21 micrograms of thyroxine in 100 ml of the serum). A variation coefficient within this range did not exceed 15%. For 14 unknown blood serum samples of patients the coefficient of correlation with the results obtained using a commercial kit for T4 determination (Abbott, USA) was 0.960. The time of the assay was 2-2.5 h. This method is simple and easy for use on a large scale.

Calibration↗
PubMedDate not supplied

[Obtaining immune sera to high-titre thyroxine for immunoassay].

A method of synthesis of L-thyroxine-protein conjugates was described. It included an additional step of acylation of thyroxine by succinic anhydride. The acylated derivative was activated by carbodiimide to produce a compound which could react with the protein amino-groups. Protein conjugates with a high thyroxine content (over 20 thyroxine residues per molecule) were synthesized with minimal protein cross-linking using this method. The excess of acylated thyroxine was easily dialized against alkaline buffer. The rabbits immunized monthly with such conjugates gave antisera with high titers (1:2000-1:4000 by the ELISA method) in 9-13 weeks after the first injection. The method can be applied to other thyroid hormones and some-molecular-weight antigens.

Acylation↗
PubMedDate not supplied

[Isolation and study of the properties of IgG from antisera to influenza virus proteins].

IgG to internal (NP, M) and external (HA, NA) proteins of influenza virus were isolated from immune rabbit sera using caprylic acid. The IgG retained their specificity and activity, worked in HI, lectin test, and enzyme-immunoassay. IgG migrated towards cathode in electrophoresis on acetate cellulose. Electrophoresis in polyacrylamide gel revealed a heavy chain (55,000) and a light chain (25,000) in IgG molecule. Different methods revealed changes in IgG structure after lyophilization. The possibility of recovering IgG from rat sera using caprylic acid was demonstrated. A comparative analysis of IgG recovered by means of caprylic acid as well as by polyethylene glycol or ammonium sulphate showed the IgG recovered with caprylic acid to contain less admixtures and to be suitable for use in immunological tests without additional purification.

Animals↗