[Production and properties evaluation of monoclonal antibodies against Pseudomonas aeruginosa exotoxin A].

AIM: Production, study of properties and evaluation of a possibility to use monoclonal antibodies against Pseudomonas aeruginosa exotoxin A (ETA) for the detection of molecules of recombinant exotoxin A and anatoxin. MATERIALS AND METHODS: Producer hybridomas for monoclonal antibodies against ETA were generated. Recombinant exotoxin A, its atoxic forms, P. aeruginosa anatoxin and P. aeruginosa PA-103 live culture were used for immunization of mice. RESULTS: The experiment of fusion of malignant cell line with immune lymphocytes obtained from a mouse immunized with recombinant ETA turned out to be the most productive. Intensity of interaction of monoclonal antibodies with recombinant atoxic forms of ETA was evaluated in enzyme immunoassay. Protective (toxin-neutralizing) activity of antibodies in cell culture and the ability to detect exotoxin A in latex-agglutination reaction were studied. Antibodies of hybrid culture No. 21 are able to detect molecules of recombinant ETA and anatoxin in the latex-agglutination reaction. CONCLUSION: Use of the antibodies produced for testing toxigenicity of P. aeruginosa strains as well as detection of anatoxin in the technological process of production of prophylaxis preparations based on exotoxin A is expected.

[Immunochromatographic and latex-agglutination systems for diphtheria toxin detection].

The use of two monoclonal antibody types specific to different epitopes of diphtheria toxin systems have been developed to reveal diphtheria corynebacteria toxigenicity rapidly based on immunochromatographic and latex-agglutination detection of the diphtheria toxin. The methods have been tested on a sample of 36 clinical isolates. The possibility of significant detection of the toxigenic properties of the Corynebacterium strain, grown for 1 day, has been demonstrated. The developed methods allow for the detection of diphtheria toxin in concentrations of 3-4 ng/ml. The developed test systems are a perspective tool for diphtheria diagnostics because of significant time shortening as compared to traditional microbiological methods.

[Development of monoclonal latex diagnostic kit for diphtheria toxin and toxoid detection].

Latex diagnostic kit with high specificity and sensitivity of diphtheria toxin and toxoid detection has been developed on the basis of protective monoclonal antibodies to diphtheria toxin and polyacrolein microspheres. Diagnostic kit was stable during 1 year of shelf-life (time of the study).

[Use of monoclonal antibodies against horse immunoglobulin in an enzyme immunoassay of bacterial toxins and anatoxins].

Immunization of BALB/c mice by horse antiserum against diphtheria made it possible to obtain IgG1 monoclonal antibodies (MoAbs) 2B7E4 specific for light chains of horse immunoglobulin (Ig). Unlike commercial preparations of anti-horse immunoglobulin antibodies, which are specific for the whole Ig molecule or its Fc-fragment, the peroxidase (HRP) conjugate of the MoAb, 2B7E4-HRP did not interact with human, mouse, rabbit, and sheep Igs, or horse albumin. The conjugate obtained was used with MoAbs against bacterial toxins and commercial horse anatoxins, as a universal reagent in sandwich enzyme immunoassay (ELISA) for bacterial toxins and anatoxins. The detection sensitivity of diphtheria toxin/anatoxin equaled 0.0005 Lf/ml; tetanus toxin and anatoxin were detected with sensitivities of 20 LD50/ml and 0.005 UI/ml, respectively. A similar sandwich ELISA for botulinum anatoxins (group measurement) allowed types A, B, and E to be detected at 0.02, 0.002, and 0.001 UI/ml, respectively; selective measurement was only possible in the case of type E anatoxin (0.001 UI/ml).

[Peroxidase refolding in the presence of specific antibodies]. / Izuchenie refoldinga peroksidazy v prisutstvii spetsificheskikh antitel.

A panel of eight monoclonal antibodies raised against horseradish root peroxidase has been assembled and characterized. Affinity constants were determined for all antibodies, and their specificity for various structural forms of the enzyme (native peroxidase, apoperoxidase, and denatured peroxidase) were assessed by competitive enzyme immunoassay. The effects of the antibodies on the process of refolding of peroxidase after its denaturing with 6.5 M guanidine hydrochloride were studied spectrophotometrically, by the restoration of the enzymatic activity in the reaction of 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonate). The yield of the active enzyme in the course of the refolding was increased 1.5 to 1.7 times in the presence of antibody H1. Effects of the antibodies constituting the panel on the activity of native peroxidase and the stability of its dilute solutions were analyzed.

[Specific features of tuberculin sensitivity in children with allergic dermatoses]. / Osobennosti tuberkulinovoi chuvstvitel'nosti u detei s allergodermatozami.

The recent rise in the incidence of allergic diseases, including allergic dermatoses, requires a comprehensive approach to solving a number of problems associated with this disease. Analyzing the case reports of 62 children with allergic dermatoses has indicated that this is a particular contingent of children, which needs a more careful approach to different preventive measures, to interpreting tuberculin reactions and hence to substantiating the use chemotherapeutical prevention. A graded skin tuberculin test and an immunological study should be performed in critical cases.

[Comparative immunochemical characteristics of polyclonal and monoclonal antibodies to aflatoxin B1]. / Sravnitel'naia immunokhimicheskaia kharakteristika poliklonal'nykh i monoklonal'nykh antitel k aflatoksinu B1.

Four hybrid clones (MM-(AB1)-1, MM-(AB1)-2, MM-(AB1)-3, and MM-(AB1)-4) were obtained by hybridoma technology with immunization of BALB/c mice with a BSA conjugate of aflatoxin B1 carboxymethyloxime derivative. Antibodies produced by these clones varied in the ability to recognize the aflatoxin B1 analogues. The sensitivity of enzyme immunoassay based on all monoclonal antibodies was higher compared to analysis based on polyclonal rabbit antibodies (0.1 and 0.4 ng/ml, respectively).

[The estimation of the amount of diphtheria toxin and its activity by various tests in the dynamic cultivation of Corynebacterium diphtheriae PW8]. / Otsenka kolichestva difteriinogo toksina i ego aktivnosti raznymi testami v dinamike kul'tivirovaniia Corynebacterium diphtheriae PW8.

Direct correlation between the results of tests for the biological activity of diphtheria toxin, carried out in vivo guinea pigs and in vitro in the microcytotoxicity test in CHO cell culture, has been established, which makes it possible to use the latter as one of the methods for the rapid, reproducible and economic evaluation of diphtheria toxin. The qualitative and quantitative evaluation of diphtheria toxin in the enzyme immunoassay with the use of monoclonal antibodies and in the microcytotoxicity test demonstrates that these two tests, when used for controlling cultivation processes, have essential advantages over the flocculation test as regards their specificity and information content.

[Preparation and characteristics of antibodies to a sulfur-containing hapten, mimicking the transition state in lactamization reactions]. / Poluchenie i kharakteristika antitel k serosoderzhashchemu gaptenu, imitiruiushchemu perekhodnoe sostoianie v reaktsiiakh laktamizatsii.

A sulfur-containing hapten was proposed to be an analogue of the tetrahedral transition state of lactamization reactions. The dynamics of the immune response to this hapten upon using different immunization schemes was studied. Sixty-one clones of mouse anti-hapten antibodies were obtained. Three clones efficiently bound not only the transition state analogue (the hapten) but also potential substrates of lactamization reactions. Antibody-induced changes in the optical absorption spectra of substrate solutions were studied for a substrate with stabilized active conformation (2-aminomethylbenzoate) and an unmodified substrate (gamma-aminobutyric acid), which demonstrated that the catalytic antibodies obtained significantly accelerated the lactam ring formation.

[Effects of the antioxidant ionol on humoral immune response in experimental influenza]. / Vliianie antioksidanta ionola na gumoral'noe zveno immunogo otveta pri éksperimental'nom grippe.

The effect of the antioxidant ionol on a humoral immune response was studied in experimental influenza. With preventive introduction of ionol, the affinity of anti-influenza G antibodies circulating in the blood stream was shown to be decreased as compared to that in control animals. The changes in the affinity during influenza infection were cyclic and less marked in the group of animals receiving ionol. The administration of the antioxidant substantially reduced the detection rate of specific blood immune complexes and contributed to the production of antibody-forming cells. The findings suggest that ionol has a favourable effect on a humoral immune response in experimental influenza and alleviates the course of an infectious process.

[Glucose utilization in influenza of varying severity in intact or stressed animals]. / Uvoenie gliukozy na fone grippoznoi infektsii raznoi tiazhesti u intaktnykh ili stressirovannykh zhivotnykh.

Animals preexposed to stress developed manifest hyperglycemia in the first 2 days of influenza infection, which was replaced by prolonged hypoglycemia. The status of carbohydrate metabolism normalized no sooner than 3 to 4 weeks after the disease onset, although the acute phase of infection was over by the end of week 2. The changes were evident in the pattern of "sugar curve" after glucose loading. Glucose utilization failed to normalize even 2 h after the loading in animals with stress, influenza infection, or stress-associated complications of this infection.

[Analysis of interactions between influenza virus and murine serum proteins in non-infected and infected mice]. / Analiz vzaimodeistviia virusa grippa s belkami myshinoi syvorotki krovi neinfitsirovannykh i infitsirovannykh myshei.

The presence of proteins in the blood of intact mice of various lines which could form complexes of various stability with influenza virus and have antigen determinants common with mouse IgG was demonstrated. These proteins were different from mouse IgG and their molecular weight was 100-200 and 40-80 kD. The authors described quantitative changes in the studied proteins during simulated influenza as well as under the effect of ionol (antioxidant) administration. The study permitted the assumption that blood serum proteins of intact mice participate in the infection progression and are significant for its prognosis.

[Differentiated study of specific antibody populations as a possible way to increase the prognostic value of indicators of humoral immunity in influenza]. / Differentsirovannoe izuchenie populiatsii spetsificheskikh antitel kak vozmozhnyi pu't povysheniia prognosticheskoi znachimosti poka- zatelei gumora'lnogo immuniteta pri grippe.

The article overviews the literature and original research data on the ways to increase the informative value of assessed parameters of the humoral immune response to viral disease formation. The need to analyse the heterogeneity of specific IgG class antibodies by their affinity index and to determine the level of circulating specific immune complexes is discussed. It is demonstrated that the value of the humoral immunity studies in influenza-virus infection may be significantly increased by investigating not only the accumulation of circulating immunoglobulins of various classes but the antibody affinity within its class and by assessing the proportions of circulating and immune-complex-bound antibodies. This allows for considering the humoral immunity as a constituent of the interrelated and interdependent immunity mechanism complex.

[Thyroid development and morphofunctional state in the postnatal ontogeny of rats with cervical sympathetic ganglia removed]. / Razvitie i morfofunktsional'noe sostoianie shchitovidnoi zhelezy v postnatal'nom ontogeneze krys s udalennymi sheinymi simpaticheskimi gangliiami.

It has been demonstrated that bilateral extirpation of the cervical sympathetic ganglia in newborn rats results in the enlargement of perivascular connective tissue of the thyroid, as well as in degenerative changes in thyrocytes, desquamation of the latter in the follicular cavity and the increase in the number of large "light" cells. Unilateral extirpation of the cervical sympathetic ganglia usually is not accompanied by changes in the follicular structure of the gland. The enlargement of the connective tissue and changes in the blood vessels are more significant at the side of the operation, whereas the decreased in the height of the thyrocytes is similar in both lobes of the thyroid. Retardation in the development of the thyroid ant the decrease in its functional activity result, evidently, not only from direct denervation of the thyroid, but from changes in the level of regulation of the latter by higher autonomic centres of the brain as well.

[Hypothalamo-hypophyseal system and thyroid gland of Acipenser güldenstädti sturgeon fry during temperature changes]. / Gipotalamo-gipofizarnaia neirosekretornaia sistema i shchitovidnaia zheleza u mal'kov osetra Acipenser güldenstädti v usloviiakh smeny temperature.

In the parr of the sturgeon (body length varies from 6.1 to 14.0 cm) histometric studies have been made on the elements of hypothalamo-hypophyseal neurosecretory system, tireotropic cells (TC) of the hypophysis and the thyroid gland (TG) during the decrease of water temperature from 24--27 degrees to 10--14 degrees. It was shown that in all size groups of the fish an increase in the number, growth and differentiation of neurosecretory cells in the dorsal part of the preoptic nucleus are taking place. Correlation was found between the activity of neurosecretory cells of the dorsal part of the preoptic nucleus and hypophyseal TC. The response to the decrease in water temperature follows the pattern of stress reaction, being expressed to different extent in various size groups of the parr depending on the initial condition of the functional complex investigated.

[Electron microscopic characteristics of neurosecretory cells in the preoptic nucleus of sturgeon fry]. / Elektronnomikroskopicheskaia kharakteristika neirosekretornykh kletok preopticheskogo iadra mal'kov osetra

In the sturgeon fry nucleus preopticus (Npo), light and weakly differentiated neurosecretory cells (NSC) as well as differentiated (i-cells) were revealed. Among these types of cells, there were the transitional forms. No "picnomorphic" cells were found in any Npo zone. NSC contained different amounts of elementary secretory granules 110-270 nm in diameter. The distribution of organelles was evaluated statistically, their number increasing from i-cells to the NSC. These data suggest that growth and differentiation of the NSC take place during this stage of ontogeny. The results are discussed in terms of specific functioning and maturity of the NSC.