Development of a real-time PCR method for rapid sexing of human preimplantation embryos

Genes on the X chromosome are known to be responsible for more than 200 hereditary diseases. After IVF, the simple selection of embryo sex before uterine transfer can prevent the occurrence of affected offspring among couples at risk for these genetic disorders. The aim of this investigation was to develop a rapid method of preimplantation genetic diagnosis (PGD) using real-time polymerase chain reaction (PCR) for the sexing of human embryos, and to compare it to the fluorescence in-situ hybridization technique, considered to be the gold standard. After biopsies were obtained from 40 surplus non-viable embryos for transfer, a total of 98 blastomeres were analysed. It was possible to analyse 24 embryos (60%) by both techniques, generating a total of 70 blastomeres (35 per technique), white 28 blastomeres from 16 embryos (40%) were analysed only by real-time PCR. A rapid and safe method was developed in the present study for the sexual diagnosis of a single human cell (blastomere and buccal cell) using the emerging technology of real-time PCR. (C) 2009, Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

Typing class I HLA-A gene using a nested PCR-RFLP procedure

In order to detect several new HLA-A class I alleles that have been described since 1998, the original PCR-RFLP method developed to identify the 78 alleles recognized at that time at high resolution level was adapted by us for low and medium resolution levels using a nested PCR-RFLP approach. The results obtained from blood samples of 23 subjects using both the PCR-RFLP method and a commercial kit (MicroSSP1A®, One Lambda Inc.) showed an agreement higher than 95%. The PCR-RFLP adapted method was effective in low and medium resolution histocompatibility evaluations.

Quantitative real-time RT-PCR and chromogenic in situ hybridization: precise methods to detect HER-2 status in breast carcinoma

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

The heminested RT-PCR for the study of rabies virus pathogenesis

The aim of the present trial was to evaluate the heminested RT-PCR for the study of rabies virus distribution in mice inoculated experimentally. Inoculation was by the intramuscular route in 150 mice, using the dog street rabies virus. Groups of five animals were killed at different times. Fragments of different organs were collected and the material was tested by Fluorescent Antibody Test (FAT) and heminested RT-PCR (hn RT-PCR). Positive results were obtained beginning on the 10th day after inoculation in the brain, spinal cord, salivary gland, limbs, lungs, liver, spleen, urinary bladder, tongue and right kidney. Hn RT-PCR was shown to be more efficient for the study of rabies virus distribution in different tissues and organs. (C) 2004 Elsevier B.V.. All rights reserved.

Cloning, sequencing and expression analysis of the equine hepcidin gene by real-time PCR

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Direct Detection of Mycobacterium bovis in Bovine Lymph Nodes by PCR

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Species-specific nested PCR as a diagnostic tool for Brucella ovis infection in rams

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

First identification of Sarcocystis tenella (Railliet, 1886) Moule, 1886 (Protozoa: Apicomplexa) by PCR in naturally infected sheep from Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Comparison of agar gel immunodiffusion test, rapid slide agglutination test, microbiological culture and PCR for the diagnosis of canine brucellosis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Comparison of a PCR assay in whole blood and serum specimens for canine brucellosis diagnosis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Performance of IFAT, ELISA, direct parasitological examination and PCR on lymph node aspirates for canine visceral leishmaniasis diagnosis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Correlação dos achados clínicos e hematológicos com diagnóstico definitivo de erliquiose canina por meio de PCR

The Ehrlichiosis is a worldwide diseases of great importance in a veterinary medicine is an important infectious diseases whose prevalence has increased significant in the last year in the Brazilian states. Due to the fact that this study was designed to correlate the findings hematological clinical signs and PCR, being the most sensitive. This study evaluated twenty dogs seen at veterinary hospital UNESP - Botucatu campus during the 03 from august to September 28 2009. Animal cited 65% were positive in the PCR test. Among the most prominent clinical findings 76.92% (10/13) with anorexia, 53.84% (7/13) with hepatoesplenomegaly, 46.15% (6/13) with apathy and 38.46% (5/13) epistaxis. The thirteen animals positive PCR 92.30% (12/13) showed thrombocytopenia (<150.000 platelets) e 61.53 (8/13) anemic (<5.50 x10). Thus, we conclude that the PCR was a good method for detection differential canine ehrlichiosis may be adopted together with the clinical and hematological findings for the accurate diagnosis of the disease.

Expressão gênica dos REα, REβ e PR em tumores mamários de cadelas por meio do q-PCR

A expressão de receptores de estrógeno (ER) e progesterona (PR) por meio da técnica de q-PCR foi avaliada em 26 cadelas portadoras de neoplasias mamárias e cinco cadelas sem afecções mamárias (grupo controle). Os resultados mostraram que os três grupos de animais estudados - com tumor maligno ou benigno e controle - expressaram receptores de estrógeno alfa, beta e progesterona. A quantificação relativa mostrou tendência para uma expressão maior de receptores no grupo controle e menor no grupo de animais com neoplasias malignas. Além disso, observou-se expressão maior de ERα em relação ao ERβ, e as neoplasias malignas de origem mista apresentaram maiores concentrações dos receptores PR, ERα e ERβ que as neoplasias de origem epitelial.

Detection of bovine Herpesvirus type 5 in formalin-fixed, paraffin-embedded bovine brain by PCR: a useful adjunct to conventional tissue-based diagnostic test of bovine encephalitis

The aim of this study was to evaluate the application of PCR technique for the detection of BoHV-5 in routinely formalin-fixed, paraffin-embedded brain tissues in 20 naturally infected calves affected by fatal meningoencephalitis. Brains were divided into two halves, one kept fresh for virus isolation and PCR assay, targeting the glycoprotein C gene from BoHV-5 genome. The other half brain, corresponding to posterior cortex region, was submitted to formalin fixation and embedded into paraffin blocks for microscopic evaluation and total DNA isolation. Most of the slides showed severe multifocal non-supurative encephalitis with neuronal degeneration, neurophagia, and no acidophilic intranuclear inclusions could be found in neurons and glial. The 20 fresh samples were confirmed, by virus isolation and PCR assay, as having the BoHV-5 virus and, respective glicoprotein C sequence, while 15 of 20 formalin-fixed, paraffin-embedded samples were considered positive for the same analysis. The results revealed the first description of PCR efficiency, applied to formalin-fixed, paraffin-embedded brain collected from naturally infected calves, improving the detection of BoHV-5 from archival samples in South America. (c) 2007 Published by Elsevier B.V.