Design of Experiment-Based Optimization of Human Chondrocyte Cell Culture Using Four Different Bioreactor Systems
- 주제(키워드) Cell culture
- 발행기관 고려대학교 대학원
- 지도교수 김익환
- 발행년도 2018
- 학위수여년월 2018. 2
- 학위구분 석사
- 학과 대학원 생명공학과
- 원문페이지 109 p
- 실제URI http://www.dcollection.net/handler/korea/000000081038
- 본문언어 영어
초록/요약
Human life expectancy is increasing in part because of scientific advances. However, aging is related to many diseases and disorders in the body. One of the most common aging-related diseases isosteoarthritis (OA). If cell therapyrather than conventional drugs can be used to treat OA, treatment may be improved. However, a large number of chondrocyte cells are needed forOA therapy. Additionally, the cells must be high-quality and subjected to strict regulation. Traditional cell culture processesfor chondrocytes are costly and labor-intensive; thus,this study used Design Of Experiment (DOE) to test 4 different types of bioreactors to determine optimal methodsfor chondrocyte cell culture.This bioreactor system increased cell growth byapproximately 1833%.
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List of Figures…………………………………………………i
List of Tables………………………………………………vi
List of Abbreviations………………………………………vii
Abstract………………………………………………………viii
Introduction……………………………………………………1
Materials and Methods………………………………………5
1.Cellisolation&MasterCellBanking(MCB)………………5
2.Microcarrier preparation…………………………………7
3.Design of experiment(DOE)…………………………9
4. Microcarrier selection&feedingstrategyselection ………13
5. Serum-free media (SFM)selection………………………15
6. Biostat b bioreactor…………………………………………17
7. PBS bioreactor………………………………………………19
8.Wave bioreactor………………………………………………21
9. Slanted roller bottle (SRB)…………………………………23
10. Four-way slanted roller bottle (4-way SRB) …………25
11. Helix structure slanted roller bottle (HSSRB)…………27
12. Measurement of metabolite………………………………30
Results and Discussion………………………………………29
1. Result ofdesign of experiment (DOE) ……………………31
1.1. Cell kinetics of design of experiment (DOE)…………31
1.2. Analysisofdesign of experiment (DOE)……………… 34
2. Result of microcarrier selection &feeding strategy selection ………………………………………………36
2.1. Cell growth test of microcarrier selection………………36
2.2.Detach test of microcarrier selection ……………………41
2.3. Feeding strategy selection ………………………………44
3. Serum-free media (SFM) selection………………………47
4. Result of human articular chondrocyte (HAC) culture in Biostat b bioreactor……………………………………………49
4.1. Human articular chondrocyte (HAC) growth kinetics and viability in Biostat b bioreactor…………………………49
4.2. Human articular chondrocyte (HAC) metabolite kinetics in Biostat b bioreactor.…………………………………………51
4.3. Human articular chondrocyte (HAC) osmolality kinetics in Biostat b bioreactor …………………………………………53
4.4. Using serum-free media (SFM) to evaluate human articular chondrocyte (HAC) culture growth kinetics in Biostat b bioreactor.……………………………………………54
5. Human articular chondrocyte (HAC) growth kinetics and viability in PBS bioreactor……………………………………55
5.1. Human articular chondrocyte (HAC) growth kinetics and viability in PBS bioreactor………………………………56
5.2. Human articular chondrocyte (HAC) metabolite kinetics in PBS bioreactor.………………………………………………58
5.3. Human articular chondrocyte (HAC) osmolality kinetics in PBS bioreactor ………………………………………………60
5.4.Using serum-free media (SFM) to evaluate human articular chondrocyte (HAC) culture growth kinetics in PBS bioreactor…………………………………………………………62
6. Human articular chondrocyte (HAC) growth kinetics and viability in Wave bioreactor……………………………………65
6.1. Human articular chondrocyte (HAC) growth kinetics and viability in Wave bioreactor………………………………65
6.2. Human articular chondrocyte (HAC) metabolite kinetics in Wave bioreactor………………………………………………69
6.3. Human articular chondrocyte (HAC) osmolality kinetics in Wave bioreactor………………………………………………71
6.4.Using serum-free media (SFM) to evaluatehuman articular chondrocyte (HAC) culture growth kinetics in Wave bioreactor…………………………………………………………73
7. Human articular chondrocyte (HAC) growth kinetics and viability in slanted roller bottle system………………………75
7.1.Human articular chondrocyte (HAC) growth kinetics and viability in slanted roller bottle…………………………………75
7.2. Human articular chondrocyte (HAC) growth kinetics and viability in four-way slanted roller bottle (four-way SRB)……………………………………………………78
7.3.Human articular chondrocyte (HAC) metabolite kinetics in helix structure slanted roller bottle (HSSRB)……………81
7.3.Human articular chondrocyte (HAC) osmolality kinetics in helix structure slanted roller bottle (HSSRB)……………84
7.4. Using serum-free media (SFM) to evaluate human articular chondrocyte (HAC) culture growth in helix structure slanted roller bottle (HSSRB)………………………88
Conclusion………………………………………………………90
References………………………………………………………92
Abstract in Korean………………………………………………94
