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B Pokrić

Publications and source records attributed to B Pokrić.

At least 19 recordsLinked to original sources
PubMed2006

Immune complex-based vaccine for pig protection against parvovirus.

The insoluble immune complexes (ICs) were prepared under the conditions of double immunodiffusion in gel, using the suspension of the ultrasound treated PK-15 cell-line infected with porcine parvovirus (PPV) containing both viral particles and viral proteins, as well as pig or rabbit anti-PPV polyclonal immune sera. The immunodiffusion performed in an agarose gel allows only viral subunits with a molecular mass equal to or less than 1000 kDa, rather than the viral particles, to diffuse through the gel and reach the point where the immunoprecipitate is to be formed. The immunoprecipitation under the conditions of the diffusion ensures the optimal, i.e. equimolar ratio of both immunoprecipitating components, antibody/antigen in the IC. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the Western blot analyses showed the ICs were composed of two proteins, a protein in which molecular mass corresponded to the VP2 of the PPV and a protein with a molecular mass of the IgG. This suggests that the ICs are mainly composed of the VP2 antigen and IgG class antibodies. The potency of the IC-vaccines prepared in the form of a water-in-oil-in-water emulsion was compared with that of a commercially available, inactivated oil vaccine. The vaccination of gilts, 6 weeks before mating, with the IC containing allogeneic pig antibodies, resulted in the development of high and long-lasting anti-PPV antibody titres, similar to those generated by the licenced vaccine (P > 0.01). The content of the virus material administered by the IC was twice lower than that in the licenced vaccine. Neither systemic nor local reactions were observed in the gilts during the period of the trial with the IC vaccine. The number of viable piglets per litter varied between 9 and 12 and no signs of the PPV infection were detected. Rabbits were used as one of the alternative laboratory animal models accepted for the testing of the vaccine against the PPV. The rabbit humoral immune response generated by the IC containing the allogeneic antibodies were higher than that generated by the ICs containing the xenogeneic pig antibodies. It was similar to that generated by two-times higher content of the virus material administered by a commercially available vaccine. The IC-based vaccines belong to non-replicating, subunit vaccines, which are both ecologically convenient and the safest vaccines of all.

Animals↗
PubMed2000

Integration of macromolecular diffraction data using radial basis function networks.

This paper presents a novel approach for intensity calculation of X-ray diffraction spots based on a two-stage radial basis function (RBF) network. The first stage uses pre-determined reference profiles from a database as basis functions in order to locate the diffraction spots and identify any overlapping regions. The second-stage RBF network employs narrow basis functions capable of local modifications of the reference profiles leading to a more accurate observed diffraction spot approximation and therefore accurate determination of spot positions and integrated intensities.

Journal Article↗
PubMed1998

Protection of broiler breeders by an inactivated combined water-in-oil-in-water viral vaccine.

A four-component vaccine, prepared by combining the single vaccines, contains subunits of Newcastle disease and infectious bronchitis viruses, as well as whole inactivated infectious bursal disease and egg drop syndrome viruses. The vaccine is prepared in the form of a low-viscosity water-in-oil-in-water emulsion with low mineral oil content. Heavy breeders were vaccinated at the age of 20 weeks by intramuscular administration of 0.5 ml vaccine/bird in an experiment carried out under field conditions, involving 5000 female and 450 male parents. The birds had previously been vaccinated with live vaccines according to an obligatory field vaccination programme. Vaccination with the WOWE vaccine near the point of lay elicited serological responses protecting both the parents and their progeny. Each of the antigens administered in the four-component vaccine was as effective as the respective single component vaccine. The mortality, recorded during the 31-week experimental period, was 6.2%. Mortality and morbidity were not triggered by viruses against which vaccination was carried out. Egg production was not affected by the vaccination and was 170.2 eggs per hen during the 28-week production period.

Adenoviridae Infections↗
PubMed1996

Analysis of the in vitro secretory activity of human pituitary adenomas: modification of corticotropin release from adenoma tissue explant cultures by addition of a human plasma ultrafiltrate bioactive fraction.

The lack of control of tumour behaviour is manifested in different ways, depending primarily on the type of tumour. This results in numerous problems of tumour diagnosis and therapy. In the case of "benign" tumours, like pituitary adenomas, in vitro studies are often used for evaluation of the tumour. The use of tissue explant cultures of human pituitary adenomas and the comparison of the feature of cultured tumours with their behaviour in vivo showed that corticotropin is released not only from the tumours associated with Cushing's disease, but also from clinically non-functioning tumours. Hence, it was supposed that the release of corticotropin in vivo from non-secreting tumours is probably under the influence of certain neuroendocrine and/or systemic humoral factors. To test this possibility, samples of 22 tumours were cultured in plain culture medium or in the presence of the "human plasma ultrafiltrate bioactive fraction" (tentatively termed as TBP) prepared by anion-exchange chromatography. In the presence of TBP the release of corticotropin was strongly inhibited in adenomas showing relatively high spontaneous secreting activity in vitro (> 200 ng/l in 24 hours), while immunohistochemistry of these tumours indicated accumulation of corticotropin inside the cells. In contrast, TBP stimulated corticotropin release from tumours that showed relatively low basic corticotropin release (< 200 ng/l in 24 hours), with no obvious change in cellular corticotropin immunoreactivity. Such a dual activity of TBP was not observed for 8 samples of adenomas cultured in the presence of surrounding pituitary tissue, probably because TBP did not affect corticotropin secretion by the normal pituitary cells (as indicated by immunohistochemistry). From these results, it appears that TBP could be one of the humoral factors involved in the regulation of corticotropin release from pituitary adenoma tissue. Its possible involvement in the regulation of corticotropin release from normal pituitary tissue, however, is uncertain.

Adenoma↗
PubMed1993

Determination of size of antigenic fragments after treatment of enveloped viruses with non-ionic detergents.

The two-dimensional double immunodiffusion, the so-called 'two-cross' method, is introduced for determination of the size of antigenic subunits released after solubilization of enveloped viruses with non-ionic detergents. The method enables the determination of diffusion coefficients and calculation of relative molecular masses of immunoreacting components without using any standard and without prior isolation, purification or labelling of material to be analysed. The molecular masses of surface antigenic fragments of the Newcastle disease virus (NDV) disrupted by non-ionic detergent Triton X-100 were determined. Two antigenic fragments, the larger having a molecular mass between 350 and 187 kDa and the smaller from 140 to 86 kDa, were released by the action of 0.1-1.0% detergent at 20 degrees C. One fragment, whose molecular mass varied from 210 to 187 kDa, was obtained at 37 degrees C after treatment of NDV with 0.2-1.0% detergent, respectively The detergent disruption of purified NDV is studied for comparison. No difference was found whether purified or NDV in allantoic fluid was subjected to the same detergent extraction.

Allantois↗
PubMed1993

Quantitative and qualitative characterization of virus envelope proteins and specific polyclonal antibodies.

A method for simultaneous qualitative and quantitative characterization of virus envelope proteins and virus-neutralizing antibodies is described. The quantitative estimate is based solely on the precipitation of the antibody/antigen (Ab/Ag) complexes at equivalence under equilibrium conditions. The qualitative analysis is performed by determining the diffusion coefficients, i.e., molecular masses of antigen and antibodies. Isolation, purification, labelling of antigen and antibodies and use of any standards are not required. The analyses are carried out directly in crude biological fluids in which antigen and antibodies naturally occur. The results obtained are independent on the avidity of the Ab/Ag system to be analysed. The method was tested by using antigenic subunits of Newcastle disease virus (NDV) and four chicken anti-NDV immune sera of different avidities.

Animals↗
PubMed1993

Application of the immune complex for immune protection against viral disease.

An immune complex (IC), composed of antigenic subunits of the Newcastle disease virus (NDV) and specific polyclonal allogeneic antibodies, was used to protect chickens against NDV. Antibodies in the IC were chicken immunoglobulin G. The antibody:antigen ratio in IC was 2.03. The IC was prepared at equivalence by direct mixing of NDV-infected allantoic fluid, treated with Triton X-100, and chicken anti-NDV serum. In order to bind NDV antigenic subunits to specific antibodies, previous isolation and purification of antigen is not required. Chickens were immunized with 1 mg IC, containing 0.3788 mg of viral antigens. The IC, prepared in the form of an oil-emulsion, was administered intramuscularly. The IC generated high levels of anti-NDV antibodies and successfully protected chickens against live virus challenge. Therefore, the IC could be recommended as a safe and environmentally convenient vaccine.

Animals↗
PubMed1992

Microgravimetric determination of precipitable antigens and antibodies in native biological fluids.

A method for the exact and simultaneous measurements of immunoprecipitable antigens and antibodies in native biological fluids is described. It is based solely on the microgravimetric analysis of at least two immunoprecipitates of different stoichiometric compositions. The volume ratios of antigen and antibody solutions for the preparation of the immunoprecipitates at equivalence are determined by the two-cross immunodiffusion technique. The proposed procedure has advantages over other methods for quantitative determination of proteins, since the separation, purification and labelling of immune components, as well as the use of standards, are not required. The method was tested by using human serum albumin (HSA) and three rabbit antiHSA immune sera of various avidities. Immunoprecipitates of different stoichiometric compositions were prepared at pH 5.0, 5.5, 7.0 and 8.6. The minimal precision of the method was +/- 0.2 micrograms of protein. The coefficients of variation were 0.14 to 0.38% for HSA and 0.03 to 0.08% for antiHSA antibody.

Animals↗
PubMed1992

Humoral immune response to the antigen administered as an immune complex.

Antigen (HSA) bound in immune complexes at equivalence with syngeneic anti-HSA antibodies elicit much stronger humoral immune response then soluble HSA. On the other hand, administration of immune complexes formed with xenogeneic (rabbit) anti-HSA antibodies suppressed humoral immune response against HSA, but not against rabbit IgG in mice. We suggest that immunization with antigen bound in immune complex might represent a powerful tool in enhancing humoral immune responses.

Animals↗
PubMed1992

Cellular immune response to the antigen administered as an immune complex in vivo.

Recently, it was shown that 10(2)- to 10(3)-fold lower doses of human serum albumin (HSA) are sufficient for the same T-cell response in vitro, if HSA is administered to the cultures bound in the immune complex rather than in the soluble form. In the present study, we analysed the capacity of HSA in the form of immune complexes to elicit specific cellular immune response in vivo. We found that antigen bound in the immune complex with murine, syngeneic polyclonal antibodies elicited the same T-cell response as fivefold higher doses of free antigen. On the other hand, HSA bound in the immune complexes with xenogeneic, rabbit polyclonal antibodies did not enhance anti-HSA cellular immune response. Our results indicate that binding of antigen in the immune complex could play an important role in enhancing an antigen-specific cellular immune response in vivo.

Animals↗
PubMed1991

Cellular immune response to the antigen administered as an immune complex.

Immune complexes are specifically bound to the Fc receptors on the surface of various types of cells capable of presenting antigens. It was therefore determined if human serum albumin (HSA), bound in an immune complex, is presented more efficiently to the HSA-specific T cells than HSA alone. Primed, polyclonal murine T cells were stimulated in vitro with either HSA alone or HSA bound at equivalence to syngeneic, polyclonal anti-HSA antibodies of IgG class. The stoichiometric composition of the insoluble immune complex was AgAb2.91. The present study shows that 10(2)-10(3)-fold lower doses of HSA are sufficient for the same T-cell response in vitro, if HSA is administered to the cultures in the form of the immune complex rather than in the soluble form. The enhancement of T-cell proliferation in the presence of immune complex was blocked with monoclonal antibody (mAb) against Fc receptor. Our results indicate that binding of antigen in the immune complex could play an important role in enhancing an antigen-specific cellular immune response.

Animals↗
PubMed1985

Effects of polyethylene glycol and dextrans on immunoprecipitations: a two-cross immunodiffusion study.

Precipitating titers and immunochemical titers obtained in a wide range of antigen-to-antibody concentration ratios by the two-cross immunodiffusion technique are compared with the corresponding laser light scatter precipitin curves. The two-cross immunodiffusion technique has also been applied to investigate whether polyethylene glycol of molecular mass 6000 and dextrans of molecular masses from 10,000 to 2,000,000 enhance the immunoprecipitation processes of the system human serum IgG-rabbit immune serum at pH 5.5 and 8.1 at 20 degrees C. It was found that the significant increase of precipitating titers of both precipitating components in the presence of polyethylene glycol is a consequence of a strong decrease of solubility of the primary antigen-antibody complex. The decrease of solubility does not affect the immunochemical titer of the immune serum, indicating stoichiometrical invariance of the precipitate at the equivalence. The apparent strong decrease of diffusion coefficients of both antigen and antibody in 20- and 40-g/liter polyethylene glycol solution is attributed to increase of viscosity of the solutions and to a partial self-association of protein molecules due to steric exclusion. In 40-g/liter polyethylene glycol solutions at pH 5.5 every fourth molecular entity of antigen and every third molecular entity of antibody are present in the form of a two-molecular self-associate, whereas in 20-g/liter polyethylene glycol solutions only 1% of antigen molecules and 8% of antibody molecules are associated. With the increase of pH to 8.1 the self-association of protein molecules is strongly further enhanced. Dextrans in 20-g/liter solutions, without regard to their relative molecular masses, do not influence precipitating titers and solubility of the antigen-antibody system at equivalence and do not enhance self-association of protein molecules. The strong decrease of diffusion coefficients of immunoglobulin G antigen and antibodies in dextran solutions is solely attributed to the increase of viscosity of the dextran solutions; hence there was no evidence of interaction of dextrans with serum IgG proteins.

Animals↗
PubMed1979

Precipitation of calcium phosphates under conditions of double diffusion in collagen and gels of gelatin and agar.

One-dimensional double diffusion was applied to determine critical concentrations at which the precipitation of calcium phosphates occurs in reconstituted connective tissue collagen and agar gels at 37 degrees C and in gelatin gels at 25 degrees C. Experiments were performed in the presence of unbuffered 0.15 mol dm-3 NaCl, or 0.15 mol dm-3 NaCl-veronal adjusted to pH 7.4. It was found that critical concentrations of precipitation of both precipitating components, CaCl2 and phosphate buffer (pH 7.4), were equimolar and independent of the ratios of initial concentrations of the components. Critical concentrations of precipitation were not affected by the concentrations and kinds of gels used. The first-formed precipitates showed amorphous structure by X-ray diffraction analyses. Infrared (IR) spectra of the precipitates indicated CaHPO4 . H2O to be their predominant species. The molar Ca/P ratio obtained by chemical analyses was 1.08. This precipitate transformed in time into octacalcium phosphate. In all experiments, two very thin membranes of precipitate were formed in the gel column at the onset of precipitation simultaneously on both sides of the actual disc of precipitate. IR spectra and chemical analyses showed that both membranes were identical to the actual precipitation discs.

Agar↗
PubMedDate not supplied

Cytoprotective effects of met-enkephalin and alpha-MSH on ethanol induced gastric lesions in rats.

We used the rat model of ethanol induced gastric lesions to measure cytoprotective effects of neuropeptides met-enkephalin and alpha-melanocyte stimulating hormone (alpha-MSH). The lesions were induced with i.g. application of 1 ml 96% ethanol. The peptides were given i.p. 1 h before the ethanol. Sacrifice was made 1 h after ethanol application and hemorrhagic gastric area was assessed in mm(2). alpha-MSH and met-enkephalin exhibited significant and additive cytoprotective effects. The protective effects of alpha-MSH were significantly stronger than of met-enkephalin. Almost total absence of lesions was obtained with met-enkephalin and alpha-MSH mixture 10:1 (10 mg/kg met-enkephalin and 1 mg/kg alpha-MSH). The addition of indomethacin (5 mg/kg s.c.) almost completely abolished the effect of met-enkaphalin, while alpha-MSH mediated cytoprotection was weakened but still present. Interestingly, indomethacin also blocked almost completely the cytoprotective effects of met-enkephalin and alpha-MSH mixture. The latter result may have a practical consequence for the clinical trials in which met-enkephalin and alpha-MSH could be used in combination with non-steroidal anti-inflammatory drugs.

Animals↗