ABSTRACT
Introducción: la hepatitis B (HB) sigue siendo un importante problema de Salud Pública debido a las consecuencias que acarrea en los pacientes infectados. Entre los grupos de alto riesgo de adquirir este virus se encuentran los pacientes con enfermedad renal crónica (ERC) en tratamiento hemodialítico, por lo que la vacunación constituye un importante método de prevención y protección contra el virus de la hepatitis B (VHB). Objetivos: determinar el nivel de antiHBsAg y hallar la prevalencia de respuesta inadecuada a la vacuna contra la HB. Detectar los factores de riesgo asociados a la falta de respuesta inmunológica a la vacuna contra HB. Metodología: estudio observacional, descriptivo, prospectivo, de corte transverso, con componente analítico. Se incluyeron pacientes adultos con ERC en tratamiento hemodialítico trisemanal del Hospital Nacional de Itauguá durante el 2015. A todos se les determinó antiHBsAg. Resultados: se incluyeron 89 sujetos, de los cuales 47% tuvieron una respuesta inadecuada a la vacuna contra el VHB, con un leve predominio del sexo masculino. Conclusiones: la frecuencia de respuesta inadecuada a la vacuna es 47%. La edad, el hábito tabáquico, las comorbilidades y el estado de nutrición por IMC resultaron como factores de riesgo no significativos en la respuesta inadecuada a la vacuna contra el VHB, mientras que los años de hemodiálsis y la uremia se relacionaron de manera significativa a esta mala respuesta.
Introduction: Hepatitis B (HB) is still an important problem of Public Health due to the consequences in the infected patients. Among the groups with high risk of acquiring this virus, there are the patients with chronic renal disease (CRD) receiving hemodialysis treatment. Vaccination is an important prevention and protection method against the hepatitis B virus (HBV). Objectives:To determine the level of anti-HBsAg, find the prevalence of inadequate response to the HB vaccine and detect the risk factors associated with the lack of immunological response to HB vaccine. Methodology: This was a cross-sectional prospective descriptive observational study with analytical component. Adult patients with CRD receiving hemodialysis treatment three times a week in the National Hospital of Itauguá during 2015 were included. Anti-HBsAg was determined in all of them. Results: Eighty nine patients were included and 47% of them had inadequate response to HB vaccine with a slight predominance of men. Conclusions: The frequency of inadequate response to the vaccine was 47%. Age, smoking habits, co-morbidities and nutritional state by BMI were non-significant risk factors in the inadequate response to HB vaccine while years of hemodialysis and uremia were significantly related to this bad response.
ABSTRACT
Objective To study the expression in high density fermentation of anti HBsAg Fab fragment in Pichia pastoris , and the purification and activity detection of expressed target protein. Method The high density fermentation of genetically engineered Pichia pastoris was proceeded in a 5L bioreactor using fed batch fermentation. The fermentation temperature was set at 28-30℃,the pH was 5 0-5 5, and the DO was kept over 20%. When the absorbance (OD 600 ) of the broth reached 400-450 (the first time of fermentation), 200-250 (the second time), and 300-350 (the third time), the induced phase was initiated, and the methanol concentration was 0 5%-1%. The fermentation ended after 96h's induction, the target protein was purified by affinity chromatography, and its activity was assessed by ELISA. Results It showed that the optimum initial cell density during methanol induced phase should be 300-350, which was good for control of the fermentation process and the expression of recombinant Fab. At the end of the fed batch phase, a yield of about 245mg/l of Fab was reached, and 98% purity could be reached as demonstrated by affinity chromatography. The results of ELISA showed that the supernatant of fermentation and the purified recombinant Fab could bind to HBsAg specifically. Conclusion The success of high density fermentation lays a sound foundation of mass production and clinical applications of recombinant humanized anti HBsAg Fab fragment
ABSTRACT
Objective The aim of the study was to develop a purification procedure on Pichia pastoris GS115/Fab expressing human anti-HBsAg Fab fragment. Methods Purity and yield ratio and conjugated activity of purified Fab fragment were analyzed with three purified ways of goat anti human Fab affinity chromatography and 14F7 monoclonal antibody affinity column, as well as Ion exchange Size exclusion column. Results 98% purity was reached through 14F7 monoclonal antibody column, and 95% purity was gained after goat anti human Fab fragment column. But yield ratio of the two affinity columns was low, being 35% and 55%, respectively. ForIon exchange Size exclusion column, purity and yield ratio of Fab fragment were very good, being 93.8% and 80% or more, respectively. Results of ELISA analysis showed that purified Fab fragment through three columns could bind to HBsAg specifically. Conclusion The purification process of recombinant anti-HBsAg Fab fragment was established. It lays a foundation for industrialization and clinical research of human anti-HBsAg Fab fragment.