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폴리락티드 - 글리콜리드 마이크로피어에 봉입된 단백질의 항원성 평가
송세현(Seh Hyon Song),조성완(Seong Wan Cho),신택환(Taek Hwan Shin),윤미경(Mi Kyoung Yoon),최영욱(Young Wook Choi) 한국약제학회 2001 Journal of Pharmaceutical Investigation Vol.31 No.3
N/A Biodegradable polymeric microspheres were studied for their usefulness as carriers for the delivery of vaccine antigens. However, protein antigen could be denatured during microencapsulation processes due to the exposure to the organic phase and stress condition of cavitation and shear force. Therefore this study was carried out to re-evaluate the degree of protein denaturation during microencapsulation with poly(lactide-co-glycolide) (PLGA) copolymer. PLGA microspheres containing ovalbumin (OVA), prepared by W/O/W multiple emulsification method, were suspended in pH 7.4 PBS and incubated with shaking at 37.5℃. Drug released medium was collected periodically and analyzed for protein contents by micro-BCA protein assay. In order to evaluate the protein integrity, release medium was subjected to the analyses of SDSPAGE and size exclusion chromatography (SEC). And enzyme-linked immunosorbent assay (ELISA) was introduced to measure the immunoreactivity of entrapped OVA and to get an insight into the three-dimensional structure of epitope. The structures of entrapped protein were not affected significantly by the results of SDS-PAGE and SEC. However, immunoreactivity of released antigen was varied, revealing the possibility of protein denaturation in some microspheres when it was evaluate by ELISA method. Therefore, in order to express the degree of protein denaturation, antigenicity ratio (AR) was obtained as follows: amount of immunoreactivity of OVA/total amount of OVA released × 100(%). ELISA method was an efficient tool to detect a protein denaturation during microencapsulation and the comparison of AR values resulted in more accurate evaluation for immunoreactivity of entrapped protein.