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Biomedical subjects

A M Díaz

Publications and source records attributed to A M Díaz.

At least 19 recordsLinked to original sources
PubMed2002

Production of culture filtrates of Sporothrix schenckii in diverse culture media.

Mycelial-form Sporothrix schenckii was studied to determine if growth in complex (Sabouraud-dextrose) or defined culture media (minimal medium, Gibco Medium 199 with and without Hepes buffer) with differing initial pH levels would affect expression of antigen components. Cultures were evaluated by continuous monitoring and serial sampling for various parameters. Great variation was seen in the protein and antigenic patterns induced by the different media. The expression of a 55 kDa component accompanied by significant acidification of the culture medium at the beginning of the stationary growth phase was notable in Sabouraud medium. In the chemically defined media, a 90 kDa component was expressed that reacted with sera from patients with sporotrichosis. The pH in these media showed little change during the different growth phases of the fungus. Among the media studied, minimal medium favored the expression of the greatest number of antigenic components. In all of the assays, the stationary growth phase appeared to be optimal for content of antigenic components. Cross reactions were not observed with any of the culture filtrates using sera from patients with other mycoses.

Culture Media↗
PubMed2000

Simultaneous spectrofluorimetric determination of (acetyl)salicylic acid, codeine and pyridoxine in pharmaceutical preparations using partial least-squares multivariate calibration.

A partial least-squares calibration (PLS) method for the simultaneous spectrofluorimetric determination of salicylic acid (SA), codeine (CO) and pyridoxine (PY) is proposed. The determination of SA, CO, and PY has been carried out in mixtures of up to three components by recording the emission fluorescence spectra between 300 and 500 nm (lambda(exc) = 220 nm). Due to the fact of the strong spectral overlap among the excitation and also among the emission spectra of these compounds, a previous separation should be carried out in order to determine them by conventional spectrofluorimetric methodologies. Here, a full-spectrum multivariate calibration PLS method is developed. The experimental calibration matrix was constructed with 14 samples. The concentration ranges considered were 0.1-2.0 (SA), 0.25-3.0 (CO) and 0.10-2.0 (PY) mg x l(-1). The optimum number of factors was selected by using the cross-validation method. The method also allows the simultaneous determination of acetylsalicylic acid (ASA), CO and PY by previous alkaline hydrolysis of ASA to SA. To check the accuracy of the proposed method, it was applied to the determination of these compounds in synthetic mixtures and in pharmaceuticals.

Analgesics, Opioid↗
PubMed2000

In vitro anti-inflammatory activity of iridoids and triterpenoid compounds isolated from Phillyrea latifolia L.

Two iridoids, oleuropeoside and ligustroside, and two triterpenoid compounds, oleanolic acid and ursolic acid, have been isolated from the leaves of Phillyrea latifolia L. (Oleaceae). These compounds were tested for interactions with the cyclooxygenase (COX) and 5-lipoxygenase (5-LOX) pathways of arachidonate metabolism in calcium ionophore-stimulated mouse peritoneal macrophages and human platelets, and for their effect on cell viability. Structure-activity relationships obtained for in vitro screening results were discussed. These compounds are capable of exerting inhibitory actions on enzymes of the arachidonate cascade. All compounds assayed showed a significant effect on prostaglandin E2 (PGE2)-release, with inhibition percentages similar to the reference drug indomethacin (IC50 = 0.95 microM). The IC50 values of the active compounds are: oleuropeoside 47 microM, ligustroside 48.53 microM, oleanolic acid 23.51 microM and ursolic acid 60.91 microM. In the leukotriene C4 (LTC4)-assay, only oleanolic acid showed a significant effect (IC50 = 16.79 microM). We also investigated the action of compounds on thromboxane B2 (TXB2)-release induced by calcium ionophore in human platelets. Of all the tested compounds, only ligustroside (IC50 = 122.63 microM) and ursolic acid (IC50 = 50.21 microM) showed a significant effect, although with less potency than the reference drug ibuprofen (IC50 = 1.27 microM). Thus, our compounds possess an array of potentially beneficial anti-inflammatory properties which may, alongside other constituents, contribute to the claimed therapeutic properties of the plant from which they are derived.

Animals↗
PubMed1999

A rapid and selective solid-phase UV spectrophotometric method for determination of ascorbic acid in pharmaceutical preparations and urine.

Ascorbic acid was determined by a solid-phase UV spectrophotometry technique through the sorption of this on a dextran-type anion-exchange resin, Sephadex QAE A-25 and posterior direct measurement of its absorbance on the resin at 267 and 400 nm, packed in a 1-mm cell. The calibration graph was linear over the range 0.3-5.0 microg ml(-1). The sensitivity obtained is more than 50 times higher than that of the corresponding solution method. The detection limit was 0.05 microg ml(-1) and the relative standard deviation 0.74% (n = 10). This method is very rapid and highly selective for determining ascorbic acid in the presence of other species absorbing in the ultraviolet region and which are normally encountered with it. The one-step method proposed was successfully applied in the determination of ascorbic acid in pharmaceutical preparations and urine and the results were of comparable accuracy as indicated by a statistical analysis of the data, using both t- and F-tests.

Ascorbic Acid↗
PubMed1992

Production of monoclonal antibodies to human immunodeficiency virus type-2.

The ROD strain of the human immunodeficiency virus type 2 (HIV-2) was used to produce monoclonal antibodies. Virus grown in CEM cells was partially purified by ultracentrifugation and solubilized in a buffer containing Triton X-100. BALB/c mice were inoculated intraperitoneally with 50 micrograms of solubilized virus preparations mixed 1:1 with complete Freund's adjuvant. Animals were boosted on day 28 and sacrificed on day 31. Spleen cells from the immunized animals were fused with SP20/Ag 14 myeloma cells and cultured in HAT medium. Following selection of the hybrids of interest by an HIV-2 ELISA procedure, hybridomas were cloned twice by limiting dilution. Six clones were found to produce antibodies that reacted with HIV-2 antigens as judged by ELISA. These antibodies were concentrated by ammonium sulfate precipitation, and analyzed by the Western blot procedure. Monoclonal antibodies specifically reactive to an HIV protein of 68 KD were obtained. These antibodies did not react with an HIV-2 band of 55 KD. These data showed that the monoclonal antibodies recognized the carboxy terminal region (the RNAse H domain) of the HIV-2 retrotranscriptase enzyme.

Animals↗
PubMed1990

Monoclonal antibodies to Nocardia asteroides and Nocardia brasiliensis antigens.

Nocardia asteroides and Nocardia brasiliensis whole-cell extracts were used as antigens to generate monoclonal antibodies (MAbs). Six stable hybrid cell lines secreting anti-Nocardia spp. MAbs were obtained. These were characterized by enzyme-linked immunosorbent assay, Western blot (immunoblot), and immunofluorescence assay. Although all the MAbs exhibited different degrees of cross-reactivity with N. asteroides and N. brasiliensis antigens as well as with culture-filtrate antigens from Mycobacteria spp., they have the potential for use as reagents in the purification of Nocardia antigens.

Antibodies, Bacterial↗
PubMed1989

[Antirabies vaccine from the brain of the suckling rat: antigen composition and immunogenic capacity].

Suckling mouse brain (SMB) rabies vaccine is the preparation most widely used in the countries of Latin American and the Caribbean. This vaccine, prepared according to the Fuenzalida and Palacios method, consists of three fixed rabies virus strains (CVS, 51, and 91). However, the World Health Organization recommends that rabies vaccines for human use be prepared using only a single strain of this virus. In order to determine whether any one of the antigens of the SMB vaccine could be eliminated from the preparation, the immunogenic capacity of the standard trivalent SMB vaccine was compared with that of experimental bivalent (CVS-51, CVS-91, and 51-91) and monovalent (CVS, 51, and 91) SMB vaccines. The study was conducted using adult and suckling albino mice provided by the laboratory at the Pan American Zoonoses Center, Buenos Aires, Argentina, and different strains of fixed and street rabies virus. The experimental vaccines were prepared using the Fuenzalida-Palacios method. Potency and cross-immunity tests were conducted. The results showed that the trivalent vaccine was the most effective in protecting the mice against both fixed and street rabies virus infections and also in inducing rapid development of neutralizing antibody at high titers.

Animals↗
PubMed1988

Production of monoclonal antibodies to Sp2/O-Ag 14 myeloma cells.

The hybridoma technology developed by Kohler and Milstein allows the obtention of antibodies of a single specificity (i.e. monoclonal antibodies). This methodology was first introduced in the Immunology Laboratory of the Department of Microbiology at the Medical Sciences Campus (MSC), University of Puerto Rico (UPR) in 1986. The production of monoclonal antibodies to Sp2/O-Ag 14 reported herein describes the procedure used and is of importance because it constitutes the first work on monoclonal antibodies performed at our institution and in Puerto Rico.

Animals↗
PubMed1988

Analysis by fluorescence microscopy and flow cytometry of monoclonal antibodies produced against cell surface antigens.

The reactivity of seven monoclonal antibodies against the surface antigens of the murine myeloma cell line Sp2/O-Ag 14 was simultaneously analyzed by fluorescence microscopy and flow cytometry. To 1 X 10(6) Sp2/O-Ag 14 cells were added 200 microliters of the monoclonal antibody and the mixture was incubated at 4 degrees C for 30 min. After washing twice with PBS, the Sp2/O-Ag 14 cells were incubated at 4 degrees C for 30 min with a 1:400 dilution of fluoresceinated goat anti-mouse antibodies. Sp2/O-Ag 14 cells were ready for analysis after washing the cells 3 X with PBS. By fluorescence microscopic analysis different patterns of reactivity with monoclonal antibodies were detected. These patterns were identified as: smooth annular, dot-like annular, dot-like patches, diffuse and homogeneous. The observed patterns may represent different cell surface epitopes being recognized by the monoclonal antibodies. Flow cytometry analysis with the EPICS V system showed reactivity of seven monoclonal antibodies with Sp2/O-Ag 14 cell surface epitopes, which ranged from 79 to 90%. Compared to fluorescence microscopy, flow cytometry provides a faster, more sensitive and more accurate quantitative measurement of the reactivity of different monoclonal antibodies against Sp2/O-Ag 14 cell surface antigens.

Animals↗
PubMed1985

Serologic and immune responses to rabies virus during different human treatments with tissue culture and suckling mouse brain vaccines.

The antibody response to rabies virus was studied in twenty volunteers immunized with different schemes of suckling mouse brain and human diploid cell culture rabies vaccines. Throughout the study period, titers in serum neutralization and indirect fluorescent antibody tests, as well as the class of immunoglobulins with antirabies activity, varied in different individuals with the treatment scheme and the antigenic potency of the vaccine. The results suggest that measurement of the IgG class of antirabies antibodies, and possibly IgA as well, may be a more adequate criterion to assess the immunogenicity of rabies vaccines than the determination of SN titers alone.

Animals↗
PubMed1985

Cross protection of mice against different rabies virus isolates.

In an attempt to identify "atypical" strains which could account for vaccination failures, 10 street, one intermediate (DR19) and 4 fixed rabies virus isolates from men, cattle, dogs, cats, mongoose and vampire bats in five countries (Argentina, Brazil, Chile, Cuba, and France) were studied by cross-protection tests in mice. For the purpose of this study, any virus that killed more than 20% of the vaccinated mice challenged with that virus was considered "atypical". When the suckling mouse brain rabies vaccine was used, two "atypical" isolates were found: one, from a human case in Chile (91, 125 mouse passages) and the other, from a vampire bat in Brazil (DR19, 22 mouse passages). However, when mice immunized with a cell culture vaccine (PV-BHK) of a higher antigenic value than the brain vaccine, were challenged with those same isolates, mortality was below 20%. The fact that these two isolates killed enough vaccinated mice to be considered "atypical" could be related to antigenic differences between these viruses and those included in the vaccine. However, since this mortality was observed only in the mice immunized with the vaccine with a lower antigenic value, reasons are given why it could be attributed to other biological characteristics of those strains than antigenic differences. Causes for vaccines failures other than immunological differences of rabies virus strains are also analyzed and discussed.

Animals↗
PubMed1984

Evaluation of hyperimmune rabies sera by the counterimmunoelectrophoresis test.

This study demonstrates that the counterimmunoelectrophoresis test is a rapid and reliable method for the determination of antibody titres in hyperimmune antirabies sera. The application of this method to the selection of human sera for the production of antirabies globulin and for the final standardization of serum batches is discussed.

Animals↗
PubMed1983

Rabies neutralizing antibodies determination by the modified counterimmunoelectrophoresis test and the rapid fluorescent focus inhibition test.

The correlation between the modified counterimmunoelectrophoresis and the rapid fluorescent focus inhibition tests for detecting rabies virus antibodies in human sera was studied, using as reference the standard mouse neutralization test. Statistical analyses showed good correlation among the three serological tests. There were no statistically significant differences in titers obtained by serum neutralization and rapid fluorescent focus inhibition tests and the estimated titers based on the results of the modified counterimmunoelectrophoresis test.

Antibodies, Viral↗
PubMed1982

Pre-exposure rabies immunization of man with suckling mouse brain vaccine.

A case series study is reported which included serologic results obtained in 195 persons recruited from personnel and trainees of the Pan American Zoonoses Center in Buenos Aires, Argentina, and workers from other institutions. The individuals were vaccinated with three doses of suckling mouse brain vaccine administered on alternate days, and some of them were given a booster dose 30 or 365 days after the primary immunization. The sizeable percentage of reactors with a high neutralizing antibody response in a short period of time, the small number of persons requiring a booster dose 30 days after the first vaccine dose, the rapid and intense secondary immune response and the absence of serious post-vaccinal reactions justify encouraging the use of this abbreviated prophylactic course of suckling mouse brain vaccine.

Antibody Formation↗