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Erythropoietin in vivo시험법의 in vitro 대체 시험법 확립
Replacement of the in vivo Bioassay for Erythropoietin with the In vitro Bioassay

  • 주관연구기관

    (주)녹십자 종합연구소

  • 연구책임자

    허재욱

  • 보고서유형

    최종보고서

  • 발행국가

    대한민국

  • 언어

    한국어

  • 발행년월

    2001-12

  • 주관부처

    식품의약품안전청

  • 사업 관리 기관

    식품의약품안전청
    Korea Food & Drug Administration

  • 등록번호

    TRKO201000015902

  • 키워드

    에리스로포이에틴.이성체.생물학적 활성.in vivo.in vitro.Erythropoietin.isoform.bioassay.

  • DB 구축일자

    2013-04-18

  • 초록 


    In vivo bioassays for biological medicines have long been considered final resort to unequivocally assess the biological activiti...

    In vivo bioassays for biological medicines have long been considered final resort to unequivocally assess the biological activities for them because there are some cases in which the biological activities obtained from in vivo bioassay and in vitro bioassay quite differ each other. Erythropoietin (EPO, epoetin) is a wellknown example among these. However, there has also been strong request to replace in vwo bioassays with in vitro bioassays or other physicochemical means in that in vivo bioassays give rise to many cumbersome problems such as high cost for the animals used, laborious procedures, significant fluctuation of data points leading to uncertainty of test results, and ethical matters involved in animal usage. EPO has been well characterized as to its biochemstry and structure function relationship and its manufacturing process is known to be stringently controlled for uniform production in many pharmaceutical companies even in Korea. Therefore, EPO could be a good candidate for the project which deals with replacement of in vivo bioassay. The in vivo biological activity of EPO depends on its sialic acid contents which confer microheterogeneity-isoforms to this protein. EPO isoforms with low sialic acid contents have lower in vivo biological · activity due to fast clearance in blood circulation. These isoforms can be detectable by isoelectric focusing technique. We have devise a method which consist of a in vitro bioassay using BaF3 cell line and a capillal-y zone electrophoresis(CZE) for the measurement of the EPO isoform distribution. The biological activity of EPO obtained using in vitro bioassay with BaF3 cell line showed good correlation (CV(%) 7.34, 5.85, 8,16, 8.08, 8.8) to EPO content measured either spectrophotometric assay (A280 0.1% =0.743) or radio immunoassay. The isoform distribution for EPO-BRP (1 :1 mixture of epoetin-a and epoetin-B, European Phannacopoeia) by CZE method resulted in isoform 2 through isoform 8. The major peaks in electrophoregram were composed of isoform 3 through 7. Oui recombinant EPO (epoetin-a) having equivalent in vivo biological activity showed the isoform distribution of isoforrn 3 through 9. The major peaks consisted of isoform 4 through 8. The peak area of isoforrn 4 was always smaller than that of isoform 5. The preparations of recombinant epoetin-a with lower in uiuo biological activity than EPO-BRP showed the isoform 2 through 8 in their electrophoregrams whose major peaks consisted of the isoform 3 through 7. The peak area of isoform 4 was larger than that of isoform 5.


    o 재조합 Erythropoietin의 Ba/F3 cell line을 이용하는 In vitro bioassay 법 확립
    o 재조합 Erythropoietin의 isoform 분석법 확립 o 재조합 Erythropoietin의 ...

    o 재조합 Erythropoietin의 Ba/F3 cell line을 이용하는 In vitro bioassay 법 확립
    o 재조합 Erythropoietin의 isoform 분석법 확립 o 재조합 Erythropoietin의 sialic acid 함량 및 당쇄 구조 분석법 확립
    o 재조합 Erythropoietin의 in vivo assay 결과와 상술한 연구 내용과의 상관 관계 (correlation) 확립


  • 목차(Contents) 

    1. 표지 ...1
    2. 제출문 ...3
    3. 국문요약문 ...4
    4. SUMMARY ...6
    5. 목차 ...8
    6. 제1장 서론 ...9
    7. 제2장 국내.외 기술개발 현황 ...10
    8. 제3장 연구개발수행 내용 및 결과 ...12
    9. 제1절 개요 ...12
    10. 제2절 ...
    1. 표지 ...1
    2. 제출문 ...3
    3. 국문요약문 ...4
    4. SUMMARY ...6
    5. 목차 ...8
    6. 제1장 서론 ...9
    7. 제2장 국내.외 기술개발 현황 ...10
    8. 제3장 연구개발수행 내용 및 결과 ...12
    9. 제1절 개요 ...12
    10. 제2절 연구내용 및 결과 ...12
    11. 제3절 연구 결과 ...19
    12. 제4장 연구개발목표 달성도 및 대외기여도 ...47
    13. 제1절 연구개발 목표 달성도 ...47
    14. 제2절 연구개발의 대외 기여도 ...47
    15. 제5장 연구개발결과의 활용성 및 계획 ...48
    16. 제6장 기타 중요변경사항 ...49
    17. 제7장 참고문헌 ...50
    18. 연구과제(세부과제) 요약서...52
  • 참고문헌

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