Objetivo Apo E is a polymorphic protein, existing in humans in three common isoforms. It is mainly distributed in serum lipoproteins and both the apo E concentration and the apo E phenotype play an important role in lipid metabolism. The (4 allele is a risk factor for cardiovascular diseases and neurodegenerative disorders. Various methods have been developed for measurement of apo E level in serum and substantial differences exist in the reported values in healthy subjects. One of the reasons of these discrepancies could be the variety of methods and calibrators used for its measurements. The use of a common reference material could improve the coherency and accuracy of the results.The overall objective was to study the feasibility of the preparation of an apo E reference material and to test the ability for it to become a reference material. Particular objectives were: a) establish an appropriate method by which to obtain purified recombinant apo E, b) assess the properties of this purified protein, c) prepare and characterize a limited batch of lyophilized apo E material, d) test the ability of this material to be used as a reference material through an interlaboratory studyWe have succeeded in the development of an optimized procedure for the production and the purification of recombinant human apo E. Using a bacterial expression system in E. Coli and a one step affinity chromatography, large quantities of recombinant apo E isoforms can be obtained and purified (> 95% purity). The physicochemical, biological and immunological properties of this protein were close to that of the human plasma protein. Using this purified protein we were able to prepare a limited batch of lyophilized material with an acceptable homogeneity and stability. The transferability study between various methods and laboratories ( n=13) using this material showed that it exhibits a precision, linearity and immunoreactivity similar to those of human serum samples with most of the used methods. Moreover, its use as a common calibrator greatly improved the comparability and accuracy of apo E measurement in blood serum (or plasma). In conclusion, after some additional checks, the preparation of a recombinant human apo E reference material is feasible and will be proposed in a near future.To meet these objectives, the project was divided into six stages: a) evaluation of existing methods for the measurement of apo E, b) optimization of the production and the purification of recombinant human apo E, c) characterization of its physicochemical, biological and immunological properties, d) preparation of a limited batch of lyophilized recombinant apo E material, e) characterization of this batch of material (homogeneity, stability, and commutability) and f) interlaboratory trials using this material in order to assess its suitability to be used as a reference material. Ámbito científico natural sciencesbiological sciencesgeneticsDNAnatural sciencesbiological sciencesbiochemistrybiomoleculesproteinsnatural sciencesbiological sciencesbiochemistrybiomoleculeslipidsmedical and health sciencesclinical medicinecardiologycardiovascular diseases Programa(s) FP3-MAT - Specific research and technological development programme (EEC) in the field of measurements and testing, 1990-1994 Tema(s) Data not available Convocatoria de propuestas Data not available Régimen de financiación CSC - Cost-sharing contracts Coordinador UNIVERSITE HENRI POINCARE NANCY 1 Aportación de la UE Sin datos Dirección Rue Lionnois 24-30 NANCY Francia Ver en el mapa Enlaces Sitio web Opens in new window Coste total Sin datos Participantes (2) Ordenar alfabéticamente Ordenar por aportación de la UE Ampliar todo Contraer todo Algemeen Ziekenhuis St.Jan Bélgica Aportación de la UE Sin datos Dirección Ruddershove 8000 Brugge Ver en el mapa Coste total Sin datos Centre de Médecine Préventive Nancy Francia Aportación de la UE Sin datos Dirección 2 avenue du Doyen J. Parisot 54501 Nandoeuvre-les-Nancy Ver en el mapa Coste total Sin datos