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S A Eremin

Publications and source records attributed to S A Eremin.

At least 19 recordsLinked to original sources
PubMed2006

Long-wavelength fluorimetry as an indirect detection system in immunoaffinity chromatography: application to environmental analysis.

The potential of long-wavelength fluorimetry when used as the detection system in immunoaffinity chromatography is assessed for the first time by applying this approach to the analysis of water and sludge samples. Nile blue (NB) was used to synthesize a long-wavelength fluorescent tracer for linear alkylbenzenesulfonates (LASs) using the carbodiimide method, in which the amino group of NB is covalently coupled to the activated carboxylic acid group of a LAS mimic with N-hydroxysuccinimide and dicyclohexylcarbodiimide. The method consists of the injection of a pre-incubated mixture containing linear sodium 4-dodecylbenzenesulfonate (LDS; used as the LAS model), anti-LAS antibodies, and the long-wavelength tracer into a commercial Protein G column. Free and bound tracer fractions are separated in the column, and the peak height of the immunochromatogram (corresponding to the free tracer) is directly measured at 626 nm (lambda (ex)) and 674 nm (lambda (em)), and then correlated to the analyte concentration. It is not necessary to perform an elution step immediately after every sample application. The dynamic range of the method is 0.05-2.5 microg ml(-1) LDS, and the detection limit is 15 ng ml(-1). The precision, expressed as the relative standard deviation, is 4.8-6.4%. Other surfactants (sodium dodecylsulfate and Triton X-100) do not cause interference. The recoveries obtained by applying the method to the analysis of water (ground- and wastewater) and sludge (primary and activated) samples ranged from 86.0 to 111.3%. Water sample analysis included an initial solid-phase extraction step, which cleaned up the samples and improved the detection limit fivefold.

Benzenesulfonates↗
PubMed2006

Amperometric immunosensor for nonylphenol determination based on peroxidase indicating reaction.

Novel immunosensor for nonylphenol (NP) determination has been developed by immobilization of specific antibodies together with horseradish peroxidase on the surface of carbon screen-printed electrode. The signal of the immunosensor is generated by the involvement of NP accumulated in the peroxidase oxidation of mediator (Methylene Blue, hydroquinone or iodide). This results in the increase of the signal recorded by linear-sweep voltammetry. The sensitivity of the detection depends on the nature of mediator, its concentration and incubation period. Cross-selectivity of the response toward readily oxidized phenolic compounds has been determined. The immunosensor developed makes it possible to detect from 20 microgL(-1) to 44 mgL(-1) of NP with detection limit 10 microgL(-1) of NP.

Animals↗
PubMed2004

[Detection of sulfamethoxazole by a piezoquarz immunosensor].

A mass susceptible immunosensor for FIA of sulfamethoxazole residues in liquid products was designed. The immunosensor is based on piezoelectric transducer. Hapten-protein conjugate (SMX-Diazo-BSA) immobilized on the preliminarily silanized electrode surface of piezoelectric quartz crystal was used as the bioreceptor coating. Optimization of the FIA conditions permitted to develop a simple and express procedure for one-step detection of sulfamethoxazole in a sample and further regeneration of the bioreceptor layer. The measuring ranges are 1 to 50 ng/ml and the detection limit is 0.15 ng/ml. The detection results were compared with the HPLC data. The advantages of the new procedure are its simplicity and rapid provision of the analysis results, possible direct detection of the analyte without additional label and repeated use of the bioreceptor layer. The new immunosensor was applied to testing of various milk specimens. It was shown that the quantity of sulfamethoxazole in all the specimens was lower than the recommended Euroresidue standards (100 ng/ml).

Biosensing Techniques↗
PubMed2001

Antibody-based methods for surfactant screening.

This brief overview summarises the immunoassay-based results obtained in the course of two years of the European INCO-Copernicus project BIOTOOLS. The project is aimed at simplifying the procedures for detection of surface active compounds (SAC) using, among others, antibody-based methods, i.e., microtiter plate-based enzyme-linked immunosorbent assays (ELISA), polarisation fluoro immunoassays (PFIA), and enzyme flow injection immunoassays (FIIA). Thirty-three rabbits were immunised with five different sulphophenyl moieties and three p-hydroxyphenyl moieties conjugated to protein immunogens to produce analytical antibodies against linear alkylbenzene sulphonates (LAS) and nonylphenol (NP). Although most of the antibodies exhibited binding reaction in indirect ELISA, only a few showed the required assay sensitivity. The best antibodies for LAS exhibited a 50% binding inhibition at IC50 19.8 microg L(-1) in indirect ELISA. Similar inhibition was observed for direct ELISA using peroxidase tracers. Antibodies against NP allowed the establishment of an indirect assay operating in the mg L(-1) range. A rapid and simple protocol for the screening of NP and LAS using homogeneous PFIA is described. The assay time for 10 samples was 7 minutes, thus allowing fast detection of the selected SAC at the mg L(-1) level. A generic competitive FIIA system, using a protein G column for separation of free and antibody-bound beta-galactosidase (beta-Gal) tracer, was developed for the screening of LAS, NP, and nonylphenol decaethoxylate (NPEO10). The FIIA had a sample throughput (STP) of 5-10 samples per hour, with limits of detection (LOD) for LAS, NP, and NPEO10 of 19.5, 52, and 2.4 microg L(-1), respectively. The developed FIIAs were applied to spiked rain and surface water.

Animals↗
PubMed2001

Fluorescence polarization studies of different forms of angiotensin-converting enzyme.

The interaction of three forms of bovine angiotensin-converting enzyme (ACE) with the competitive peptide inhibitor lisinopril with a fluorescent label was studied using fluorescence polarization. The dissociation constants Kd of the enzyme-inhibitor complexes in 50 mM Hepes-buffer (pH 7.5) containing 150 mM NaCl and 1 microM ZnCl2 at 37 degrees C were (2.3 +/- 0.4).10(-8), (2.1 +/- 0.3).10(-8), and (2.1 +/- 0.2).10(-8) M for two-domain somatic ACE, single-domain testicular ACE, and for the N-domain of the enzyme, respectively. The interaction of the enzyme with the inhibitor strongly depended on the presence of chloride in the medium, and the apparent dissociation constant of the ACE-chloride complex was (1.3 +/- 0.2).10(-3) M for the somatic enzyme. The dissociation kinetics of the complex of the inhibitor with somatic ACE did not fit the kinetics of a first-order reaction, but it was approximated by a model of simultaneous dissociation of two complexes with the dissociation rate constants (0.13 +/- 0.01) sec(-1) and (0.026 +/- 0.001) sec(-1) that were present at approximately equal initial concentrations. The dissociation kinetics of the single-domain ACE complexes with the inhibitor were apparently first-order, and the dissociation rate constants were similar: (0.055 +/- 0.001) and (0.041 +/- 0.001) sec(-1) for the N-domain and for testicular ACE, respectively.

Animals↗
PubMed1999

[Contribution of determinants, located in Bacillus anthracis chromosomes, in realizing the pathogenic properties of the pathogen].

Comparative study of virulence of B. anthracis strains harbouring pXO1 and pXO2 plasmids in mice and guinea pigs showed that among six B. anthracis strains, three were 100-1000 times less virulent for guinea pigs. Genetic construction of B. anthracis strains using transduction and conjugation transfer of resident plasmids permitted us to rule out the effects of modified pXO1 and pXO2 replicons and to prove the existence of nonidentified chromosome locuses responsible for the development of an infectious process in anthrax, along with plasmid determinants of virulence.

Animals↗
PubMed1999

[Correlation of the virulence of Bacillus anthracis with expression of signs, coded for by chromosomal genes].

Clonal analysis of Bacillus anthracis strains showed heterogeneity of the majority of populations by the proteolytic, hemolytic, and pigment-adsorbing activities. Phenotypes isolated within each population were subdivided into 4 subpopulations. The appearance of sub-populations of virulent and vaccine strains (B. anthracis Sterne, Zenkowskii) was due to the ability of clones adsorbing Congo red and possessing high activities of proteolytic and hemolytic enzymes to dissociate with the formation of phenotypes characterized by different expression of proteolysis, hemolysis, pigment adsorption, pigment production, and sporulation signs. A characteristic feature of vaccine strains of B. anthracis CTIl, Wright, and Pasteur is a relative homogeneity of their populations consisting mainly of cells with low activities of proteolytic, hemolytic, and pigment-adsorbing enzymes. Study of the heterogeneity of di-, mono-, and plasmid-free derivatives of B. anthracis showed that the process of phenotype formation did not depend on the plasmids proper. Evaluation of the virulence of clones isolated from highly virulent B. anthracis strain 81/1 showed that the processes associated with loss of sporulation capacity notably decrease the virulence for laboratory animals.

Animals↗
PubMed1998

High sample throughput flow immunoassay utilising restricted access columns for the separation of bound and free label.

A flow immunodetection system with high sample throughput capacity is described for the screening of various analytes. The immunochemical detection principle is based on the chromatographic separation of the formed immunocomplex (AbAg or AbAg*) and the free antigen (Ag) by a restricted access (RA) column, utilising size-exclusion and reversed-phase mechanism. A fluorescein labelled analyte (Ag*) was used in the competitive assay format with fluorescence detection. The speed and simplicity of the assay were the greatest advantages, allowing measurement of the analyte to be carried out in less than 1 min. The biocompatibility and capacity of the restricted access material allowed multiple injections of up to 5000, without any breakthrough of the fluorescent tracer molecule and thus need for regeneration. The flow immunoassay was developed using the well-known atrazine herbicide and some transformation products as model compounds, due to their human toxicity and widespread use. The sample throughput was 80 samples per hour and the detection limits were 1.4 nM (300 pg/ml) for atrazine (Ab I) and 2.3 nM (500 pg/ml) for the sum of triazines (Ab II-III). Different sample matrices, PBS buffer, creek water, and urine were successfully applied in the flow system without the need for any sample handling step. For plasma samples an additional clean-up step using solid-phase extraction had to be included. The resulting detection limits for atrazine in plasma and water samples using this clean-up and trace enrichment procedure were found to be 2 ng/ml and 20 pg/ml, respectively. The analysis could be performed at a sample throughput rate of 400 per 6-h working shift.

Atrazine↗
PubMed1998

A flow immunoassay for studies of human exposure and toxicity in biological samples.

This paper describes a heterogeneous competitive flow immunoassay with a high sample throughput which can be used for the screening of smaller analytes in various samples. The method is based on off-line incubation of the analyte (Ag), a fluorescent labelled tracer (Ag*) and the corresponding antibody (Ab). The separation of bound (Ab-Ag*) and free tracer (Ag*) is based on a size exclusion and reversed phase mechanism utilizing a restricted access (RA) column. The column traps the free unbound tracer (Ag*) in its hydrophobic (C18) inner cavity but excludes the large Ab-Ag* complex, which is passed on and measured by the fluorescence detector. The flow immunoassay was developed using the triazine herbicide atrazine as a model compound owing to its human toxicity and widespread use. A sample throughput of 80 samples per hour and a detection limit of 300 pg ml-1 in water were obtained. Urine samples were successfully applied for direct injections into the flow system, while for human plasma samples an additional clean-up step using solid phase extraction was efficiently included where pure extract is obtained with the highly stable and biocompatible extracting column material. The resulting detection limits for atrazine in plasma and water samples using this clean-up and trace enrichment procedure were found to be 2 ng ml-1 and 20 pg ml-1 respectively.

Animals↗
PubMed1997

Fluorescence polarization immunoassay of progesterone.

A homogeneous fluorescence polarization immunoassay (FPIA) was developed to measure levels of progesterone in urine using a TDx analyzer in photocheck mode (Abbott Labs). Two tracers of ethylenediamine fluorescein thiocarbamyl (EDF) were employed; one was synthesized from 11 alpha-hydroxyhemisuccinate progesterone (Prog-11OH-HS) and the other was synthesized from 3-(o-carboxymethyl)oxime progesterone (Prog-3CMO). Each derivative of progesterone was conjugated with bovine serum albumin and used as an immunogen which produced monoclonal antibody clone 15A (MAb 15A, anti-Prog-11OH-HS) and clone 2B7 (MAb 2B7, anti-Prog-3CMO), respectively. Different combinations of tracers and antibodies were investigated in the FPIA system. Similar sensitivity was observed when using the pair, MAb 2B7 and its homologous tracer, Prog-3CMO-EDF, or MAb 15A and its homologous tracer, Prog-11OH-HS-EDF. In this immunoassay, no separation step was required and the total time for an assay of 10 samples was approximately 7 min. The progesterone detection limit in a 10 microliters sample was 3 ng/ml. The cross-reactivity results indicate that the A-, B- and D-ring of a steroid are buried in the binding pocket of MAb 15A, while the C-ring faced outward, resulting in cross-reactivity with 11-alpha hydroxy progesterone. The A-, B- and C-ring of a steroid of MAb 2B7, in contrast, are buried deep in the pocket leaving the D-ring facing outward, resulting in some different degrees of cross-reactivity with C17 position substituted steroids.

Fluorescence Polarization Immunoassay↗
PubMed1996

[Polarization fluoroimmunoassay of propazine in reversed micelles of aerosol OT in octane].

The interaction between fluorescein-labeled propazine and antibodies against this hapten was studied in the reversed micelles of Aerosol OT in n-octane by a polarization fluoroassay. The effect of the hydration degree of micelles W0 (W0=[H2O]/[Surf]), which determines their size and surfactant concentration, on the binding of the antigen with antibodies was studied. A high hydration degree of the reversed micelles (W0 = 15-30) and low concentration of the surfactant (less than 50 microM) are optimal for binding. The binding efficacy depends upon the structure of the fluorescein-labeled hapten, particularly upon the length of the bridge binding fluorescein with propazine. It was shown that the polarization fluoroimmunoassay of propazine may be carried out in a reversed micellar system in nonpolar organic solvent (octane) with a detection limit of about 100 nM (20 microns/l). This is an order of magnitude higher than that achievable upon analysis in aqueous medium. The proposed polarization fluoroimmunoassay in a reversed micellar system makes it possible to detect haptens that are poorly soluble in water directly in organic extracts, e.g. in chloroform solutions.

Dioctyl Sulfosuccinic Acid↗
PubMed1995

Localization of the epitope in methamphetamine and its antibody use for the detection of methamphetamine and benzphetamine by polarization fluoroimmunoassay.

An antibody was prepared, using a four carbon-bridged methamphetamine molecule as an immunogen in order to develop a polarization fluoroimmunoassay for urine screening of methamphetamine and benzphetamine. Also, its binding characteristics were investigated to locate epitope sites of methamphetamine. The study showed that the antibody was highly capable of eliciting a polarization fluoroimmunoassay response. However, the detection limit was much greater for benzphetamine (0.05 ppm) than for methamphetamine (0.2 ppm) and weakly antibody binding was found with methamphetamine. This difference in sensitivity may reflect the similarity of benzphetamine to the immunogen used to produce the antibody. Both benzphetamine and the immunogen have a tertiary amine attached to a carbon bridges whereas methamphetamine has only a secondary amine and amphetamine has a primary amine group. The difference of cross-reactivity data between phenylethylamine drugs and beta-hydroxyl phenylethylamine drugs indicates that the beta-carbon position have a major influence on the antibody interaction. Thus, the substitution of hydroxyl group on beta-carbon resulted in virtually no antibody affinity, even if a tertiary amine or secondary amine group was present in the molecule. This suggests that the beta-carbon chain plays a primary role as the epitope site with cooperative binding site of tertiary amine or secondary amine in alpha-carbon position. A hydroxyl group at the beta-carbon position plays an important inhibitory role to the antibody binding.

Antibody Affinity↗
PubMed1994

A new visual enzyme immunoassay of methamphetamine using linear water-soluble polyelectrolytes.

A new visual enzyme immunoassay (EIA) technique has been developed. Oppositely charged synthetic linear water-soluble polyelectrolytes (poly-N-ethyl-4-vinyl-pyridine as polycation and polymethacrylate as polyanion) were used as carriers for reagent immobilization. The ability of these molecules to form an insoluble complex was applied for the separation of bound and free components of the immunoassay reaction mixture. This approach was realized in methamphetamine visual EIA. In the first stage of the assay two specific reactions took place during incubation of the analytical reagents with the probe to be analyzed: (1) competition between methamphetamine and hapten conjugated with peroxidase for the interaction with specific antibodies and (2) interaction of these antibodies with the protein A-polymethacrylate conjugate. As a result of these reactions the (polyanion-protein A)-antibody-(hapten-peroxidase) complex was formed. Then the reaction mixture was filtered through an Ultrabind membrane (0.45 microns) with adsorbed poly-N-ethyl-4-vinylpyridine, and the immunological complexes were immobilized to the membrane by electrostatic interaction. The level of peroxidase binding on the membrane was measured by diaminobenzidine substrate. The system described was optimized to achieve both high rapidity (20 min) and an appropriate sensitivity (0.4 micrograms/ml) for methamphetamine assay.

Animals↗
PubMed1993

[Determination of phenobarbital by polarized fluoroimmunoanalysis. The effect of immunogen structure and tracer on specificity and detection limit].

The influence of the immunogen's tracer's structure on properties of antibodies to phenobarbital was studied by the polarisation fluoroimmunoassay method. It was shown that in homogeneous method of polarisation fluoroimmunoassay the titer of antibodies to phenobarbital (1/1000-1/10,000) is affected by the position of linker binding the antigen with carrier-protein and by the structure of the linker. The affinity constant ((1-4) x 10(9) M-1), cross-reactivity, limit of the detection (0.6-2.6 micrograms/ml) phenobarbital with different combinations of antibodies and tracers were calculated. The antisera grew the more heterogeneous in affinity and specificity, the longer became the linker. The more heterogeneous antibodies are preferable in the class specific assay of barbiturates.

Animals↗
PubMed1992

Urinary cotinine fluoroimmunoassay for smoking status screening adapted to an automated analyser.

A polarization fluoroimmunoassay for cotinine, a major metabolite of nicotine, has been adapted for fully automated screening of urine samples on the Abbott TDx analyser. The method has sensitivity and specificity suitable for the discrimination of active smokers from non-smokers (including passive smokers) by application of a cut-off at 0.5 mg l-1 of total urinary cotinine. Most active smokers' urine gave results over 1 mg l-1, whereas apparent levels in non-smokers were 0.08 mg l-1 or lower. A result for one sample can be obtained in about 5 min and a throughput of 80 samples h-1 can be maintained for large-scale screening applications.

Adult↗
PubMed1992

[A comparative analysis of the immunochemical determination of gentamycin by fluorescence polarization and quenching].

A polarization fluorescence immunoassay (PFIA) for gentamicin with using a set of reagents made in this country was developed. One ml of fluorescein-labeled gentamicin (50 nM) and 100 microliters of antiserum are added to 50 microliters of the sample and the fluorescence polarization is measured. The time of the assay is 10 to 15 minutes, the range of the measurable concentrations is 0 to 800 ng/ml, the sensitivity of the method is 5 ng/ml and the accuracy is 5.8-10.3 per cent. A fluorescence quenching immunoassay (FQIA) for gentamicin was also developed. Determination of gentamicin by the FQIA does not require the use of a specific polarization fluorimeter. Its linear calibrating dependence is more convenient. However, its accuracy and sensitivity are 3 times lower than those of the PFIA.

Drug Monitoring↗