Biomedical subjects

Stella Maria Barrouin-Melo

Publications and source records attributed to Stella Maria Barrouin-Melo.

7 recordsLinked to original sources

PubMed2006

A standardized cytological and immunochemical method for the analysis of fine-needle spleen aspirates: assessment of leukocyte population changes in canine visceral leishmaniosis.

A method for the evaluation of splenic cellularity using samples collected by fine-needle aspirative biopsy was standardized in this work. The procedure includes erythrocyte lysing, preparation of cytospin films and staining by histochemical and immunocytochemical techniques. The cellular profiles of spleen preparations were compared with those observed in peripheral blood samples subjected to the same procedure. Two groups were compared, one consisting of 14 healthy uninfected and the other of 15 polysymptomatic Leishmania chagasi/infantum-infected dogs, from an endemic area for visceral leishmaniosis. Cell populations were identified by conventional hematoxilin-eosin and Wright' stainings, and by immunocytochemistry using monoclonal antibodies against canine CD45RA and CD45RB, phagocytes and a pan-leukocyte antigen. Larger neutrophil (P < 0.0001) and monocyte/macrophage (P = 0.0036) relative counts and lower lymphocyte relative counts (P < 0.0001) were found in the spleen, and not in the blood, of the animals with leishmaniosis than in those of the healthy animals. The proportions of CD45RB+ cells were higher, and of CD45RA+ cells were lower, both in the spleen and in the blood of animals with leishmaniosis than in those of healthy dogs (P < 0.05). Additionally, hematoxilin-eosin-stained cytospins of spleen aspirates from Leishmania-infected animals permitted the easy visualization of amastigote forms inside phagocytes, under light microscopy.

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PubMed2006

Can spleen aspirations be safely used for the parasitological diagnosis of canine visceral leishmaniosis? A study on assymptomatic and polysymptomatic animals.

The purpose of this study was to evaluate the safety of spleen aspiration as a sampling technique for the parasitological detection by culture and microscopy of Leishmania (chagasi) infantum. Two hundred and nine domiciled dogs from an endemic area for visceral leishmaniasis in Bahia State, Brazil, were studied. Most dogs (87%) were seropositive for anti-L. chagasi antibodies by ELISA. Clinical signs of disease were recorded and the animals monitored during and after spleen puncture in order to detect possible complications associated with the procedure. From a total of 257 splenic punctures in the 209 animals, only three minor events occurred, with no significant consequence for the animals and no association with risk factors. Leishmania was isolated from 149/180 (83%) seropositive dogs, and from 6/26 (23%) seronegative animals. The procedure did not cause adverse side effects or unnecessary suffering and confirmed the diagnosis in a large percentage of dogs. We conclude that spleen aspiration can be considered an effective and safe procedure for the definitive diagnosis of canine visceral leishmaniosis.

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PubMed2005

A monoclonal antibody against a canine CD45 homologue: analysis of tissue distribution, biochemical properties and in vitro immunological activity.

This report describes the characterisation of a monoclonal antibody (mAb), AB6, which recognises specifically a cluster of canine leukocyte surface molecules. The immunogen used for obtaining the AB6 mAb was a lysate of canine peripheral blood mononuclear cells (PBMC). This novel mAb belongs to the IgG2a isotype, and reacted in Western blot with four different canine leukocyte glycoproteins with apparent molecular weights of 180, 190, 205 and 220 kDa. The AB6 mAb recognised the majority of canine peripheral blood leukocytes as determined by flow cytometry (97%). It also exhibited a broad reactivity pattern against lymphoid and myeloid cells, inhibited the proliferation of mitogen-stimulated canine PBMC and did not recognise human PBMC and murine splenocytes. The biochemical properties, cell and tissue specificity, and in vitro biological activity of the AB6 mAb indicate that it recognises a canine CD45 homologue. The mAb could become a valuable diagnostic and research tool for the evaluation of immune functions in dogs.

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PubMed2004

Comparison between splenic and lymph node aspirations as sampling methods for the parasitological detection of Leishmania chagasi infection in dogs.

The sensitivities of spleen and lymph node cultures for the diagnosis of canine visceral leishmaniasis were compared in 64 anti-Leishmania antibody positive dogs from an endemic area in Brazil. The sensitivity of spleen cultures for Leishmania detection was 97.9%; in lymph node cultures it was 25%. Positive spleen culture was more frequent (p = 0.048, Fisher's exact probability test) in symptomatic (28 out of 33 animals) than in asymptomatic animals (19 out of 31 animals). These results support the use of spleen instead of lymph node aspiration as the choice method for the parasitological diagnosis of the infection.

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PubMed2004

Recombinant single-chain canine interleukin 12 induces interferon gamma mRNA expression in peripheral blood mononuclear cells of dogs with visceral leishmaniasis.

Canine visceral leishmaniasis poses important concerns for public health and veterinary medicine in many areas of the world. Resistance to it seems to be associated with cellular specific immune responses of the so-called Th1 type. Interleukin-12 (IL-12) is one of the most potent inducers of Th1 type of immune responses to co-administered antigens. Herein, the cloning of canine IL-12, as a single-chain fusion protein (sccaIL-12), and its expression in biologically active form in COS-7 cells is reported. Supernatants from these cells stimulated the expression of comparable amounts of interferon gamma mRNA in peripheral blood mononuclear cells from dogs with natural visceral leishmaniasis. In addition, after stimulation with sccaIL-12, there was no difference between interferon gamma mRNA expressions in peripheral blood mononuclear cells of dogs with visceral leishmaniasis and from normal healthy control animals.

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PubMed2004

A novel monoclonal antibody against canine monocytes/macrophages.

The production and partial characterization of a monoclonal antibody, the IgG1 IH1, which recognizes an antigen distributed in canine monocytes/macrophages, is reported here. The distribution and apparent molecular weight of the antigen recognized by the IH1 MAb was determined in peripheral blood leukocytes, peripheral blood monocyte-derived macrophages and tissue sections of spleen, liver and skin, using Western blotting, immunocytochemistry, immunohistochemistry and flow cytometry. The IH1 MAb-recognized antigen was detected in Western blotting under non-reducing conditions spread out as a large band covering the position corresponding to the migration of molecules with molecular weights from 55 to 73 kDa. The IH1 MAb labeled blood monocytes, tissue macrophages in lymph nodes, and in the mantle zone of the spleen, and Kupffer cells in the liver. It did not react with human cells. In flow cytometric analysis, the IH1 MAb reacted with a subpopulation of monocytes. The MAb described herein may become a valuable tool for diagnosis and research on canine diseases.

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PubMed2004

Production of monoclonal antibodies against canine leukocytes.

A panel of anti-canine leukocyte monoclonal antibodies (MAbs) was produced by immunizing BALB/c mice with canine peripheral blood mononuclear cells (PBMC), either resting or stimulated with concanavalin A (ConA). Three out of 28 clones-IH1, AB6, and HG6-screened by ELISA and producing antibody with the highest specificity for canine cell immunostaining, were subjected to three subsequent subcloning steps by limiting dilution, and selected for further characterization. These MAbs belonged to IgG1 (HG6 and IH1) and IgG2a (AB6) isotypes. The distribution of cell populations expressing the antigen recognized by the antibodies was identified by indirect immunoflorescence on canine PBMC and on tissue sections of lymph node, spleen, liver and skin. The possible crossreactivity with human PBMC was also examined in immunocytochemistry. One of the antibodies specifically recognized macrophages. The MAbs presented here can be foreseen as possible valuable diagnostic and research tools to study immune functions in dogs.

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