ABSTRACT Global advances in reproductive biotechnology have allowed for the transfer of embryos from donor females with high genetic merit to recipients using the cryopreservation technique, which preserves an embryo of excellent quality and viability, thereby achieving a feasible pregnancy rate. The objective of this study was to evaluate the quality and viability of Holstein embryos that have been cryopreserved for more than 40 years under glycerol freezing. The embryos were transferred to the recipient heifers using a non-surgical method. Two 17-month-old Holstein heifers (360 kg live weights) which were clinically healthy and reproductively active were used as the recipients. Two bovine embryos of Grade 1 quality were thawed and evaluated for their morphology. Of the two embryo transfers, one pregnancy was achieved, resulting in the birth of a calf. Therefore, embryos frozen in liquid nitrogen and glycerol as a cryopreservative for more than 40 years maintained their quality and viability to produce a live calf.

RESUMO Os avanços globais em biotecnologia reprodutiva permitiram a transferência de embriões de fêmeas doadoras com alto mérito genético para receptoras, usando-se a técnica de criopreservação, que preserva um embrião de excelente qualidade e viabilidade, alcançando, assim, uma taxa de gravidez viável. O objetivo deste estudo foi avaliar a qualidade e a viabilidade de embriões Holstein, criopreservados por mais de 40 anos, sob congelamento de glicerol. Os embriões foram transferidos para as novilhas receptoras, usando-se um método não cirúrgico. Duas novilhas Holstein de 17 meses de idade (360kg de peso vivo), que eram clinicamente saudáveis e reprodutivamente ativas, foram utilizadas como receptoras. Dois embriões bovinos de qualidade Grau 1 foram descongelados e avaliados quanto à sua morfologia. Das duas transferências embrionárias, uma gravidez foi obtida, resultando no nascimento de um bezerro. Portanto, os embriões congelados em nitrogênio líquido e glicerol como criopreservante por mais de 40 anos mantiveram sua qualidade e viabilidade para produzir um bezerro vivo.

The aim of the present study was to select phage clones from a combinatorial library of filamentous phage that mimic the N protein of Porcine Reproductive and Respiratory Syndrome Virus (PRRS) using a monoclonal antibody (SDOW17). An combinatorial library of random peptide 7-mer cysteine-constrained fused to a minor coat protein (pIII) of the M13 phage was used. To determine the effect of enrichment after each round of selection, the eluted phage were titrated, bacteriophages increased from 4,8 x 10³ plaque-forming units (pfu) in the first round to 3,2 x 10(5) pfu in the third round. After three rounds of panning 32 phage clones were randomly picked, each individual clone was amplified, purified and titred. The binding reactivity of phage clones with anti-PRRS antibodies was determined using a phage ELISA. Alignments of the selected clones with the published sequences of the N protein of PRRS virus were located at the amino terminal region and middle portion of the protein. The consensus amino acid residues of the 32 mimotopes sequenced was NYRYQ. Mimotope sequences were used to calculate the epitopes of the nucleocapsid protein by the web tool of Peptiope server with PepSurf algorithm. Two conformational epitopes against monoclonal antibody anti-protein N were identified, located in a different position and appeared mainly as an a-helix. The selected mimotopes and conformational epitopes could be regarded as informative for diagnosis of disease.

El objetivo del estudio fue seleccionar de una biblioteca combinatoria de fagos filamentosos clonas que mimeticen la proteína N del virus del Síndrome Respiratorio y Reproductivo Porcino (PRRS) utilizando un anticuerpo monoclonal (SDOW17). Se utilizó una biblioteca combinatoria de fagos filamentosos (M13) que expone aleatoriamente péptidos de 7 aminoácidos en la región N-terminal fusionado con la proteína III. Para determinar el efecto del enriquecimiento después de cada ronda de selección, los fagos eluidos fueron titulados; estos se incrementaron de 4,8 x 10³ unidades formadoras de colonias (ufc) en la primera ronda a 3,2 x 10(5) ufc en la tercera. Después de tres rondas de selección, 32 clonas fueron picadas al azar, cada clona individual fue amplificada, purificada y titulada. La reactividad de la unión de las clonas a los anticuerpos anti-PRRS fue determinado utilizando un ELISA de fagos. El alineamiento de las clonas secuenciadas con la obtenida del GenBank, ubican a estas en la región aminoterminal y porción media de la proteína N del vPRRS. La secuencia consenso de las 32 clonas seleccionadas fue NYRYQ. Las secuencias de los mimotopos fueron utilizadas para calcular los epitopos de la proteína N, mediante la herramienta bioinformática del servidor Peptiope con el algoritmo PepSurf. Dos epitopos conformacionales contra el anticuerpo monoclonal antiproteína N fueron identificados, localizados en diferentes posiciones y presentados principalmente como hélices a. Los mimotopos seleccionados y epitopos conformacionales podrían ser considerados informativos para el diagnóstico de la enfermedad.