Title Page
I. 초록
Contents
II. Introduction 13
III. Result 16
PART I 16
1. Method 16
1.1. Synthesis of Liposome and Ecto-Liposome 16
1.2. Physicochemical Characterization 18
1.3. Bio-TEM Sample Preparation 18
1.4. Encapsulation of Triam to Liposome and Ecto-Liposomes 18
1.5. Encapsulation of DOX to Liposome and Ecto-Liposomes 19
1.6. Cell culture 19
1.7. SDS-PAGE 19
1.8. Fluorescence resonance energy transfer (FRET) Assay 21
2. Results 23
2.1. Optimization of Ecto-Liposomes 23
2.2. Fabrication and Characterization of Ecto-Liposome 26
2.3. FRET Assay of Ecto-Liposome 31
2.4. SDS-PAGE of Ecto-Liposome 33
2.5. Uptake of Ecto-Liposome 35
2.6. Fabrication and Characterization of Ecto-Lipo-Triam 37
2.7. Fabrication and Characterization of Ecto-Lipo-DOX 39
2.8. Cell Viability Assay of Ecto-Lipo-DOX 41
PART II 43
1. Method 43
1.1. Synthesis of AuS-TA 43
1.2. Conjugation of Anti-CD90-Triam to AuS 43
1.3. Conjugation of ADMSC AuS-CD90-Triam 44
1.4. Physicochemical Characterization 44
1.5. TEM Sample Preparation 44
1.6. Cell Culture 45
1.7. Confocal Analysis 45
2. Results 47
2.1. Physicochemical Analysis of AuS-TA 47
2.2. Triam conjugation analysis of AuS-TA 51
2.3. Physicochemical analysis of ADMSC conjugated AuS-TA 55
2.4. Confocal Analysis of ADMSC AuS-CD90-TA 61
IV. Discussion 63
V. References 65
Abstract 68
Table 1. Conjugation ratio analysis of AuS-Triam 54
Table 2. Conjugation ratio analysis of ADMSC AuS-CD90-Triam 60
Figure 1. Schematic illustration of method for synthesis of Ecto-Liposome. 17
Figure 2. Liposome Synthesis Jig 22
Figure 3. Lipid chemical structure and Ecto-Liposome Optimization parameters 25
Figure 4. UV-vis of Liposomes, Ectosomes and Ecto-Liposome 28
Figure 5. Physicochemical properties of Liposomes, Ectosomes and Ecto-Liposome. Transmission electron microscopy (TEM) images (scale bar;... 29
Figure 6. NTA analysis of Ecto-Liposome. The number and size of Ecto-Liposome (J774-derived ectosomes and liposomes were fused) determined... 30
Figure 7. FRET Assay of Ecto-Liposome. The illustration of FRET effect of Ecto-Liposome (upper). Two graph shows that fluorescence intensity at... 32
Figure 8. SDS-PAGE analysis of Liposome, Ectosome, and Ecto-Liposome (E-L). 34
Figure 9. CLSM images of the cellular uptake of fluorescence-labeled Ecto-Liposome (DiO, green, Liposome). 36
Figure 10. Physicochemical properties of Ecto-Lipo-Triam. (a) The schematic illustration of Ecto-Lipo-Triam. (b) The UV-vis (c) The size (d)... 38
Figure 11. Physicochemical properties of Ecto-Lipo-DOX. (a) The schematic illustration of Ecto-Lipo-DOX. (b) The UV-vis (c) The size (d)... 40
Figure 12. Ecto-Lipo-DOX cell viability after treatment doxorubicin hydrochloride (DOX), Ecto-Liposome, Liposome-DOX, and Ecto-... 42
Figure 13. Method of AuS-TA 48
Figure 14. UV-vis analysis of AuS-Triam. UV-Vis results of the Triam peak at 242nm 49
Figure 15. The size and zeta potential of AuS-PEG and AuS-Triam (PEGylated AuS-Triam). 50
Figure 16. TEM images of AuS-PEG and AuS-Triam showing the distributions of Au (yellow), S (an element in PEG; orange), and F (an... 52
Figure 17. Mass of conjugated AuS and Triam after synthesis of AuS-Triam. The final ratio of AuS and Triam. The graph shows the loss percentage of Au. 53
Figure 18. Method of ADMSC AuS-CD90-Triam 56
Figure 19. UV-vis of AuS-PEG, Triam, AuS-Triam, ADMSC, and ADMSC AuS-CD90-TA. 57
Figure 20. Size and Zeta potential of AuSTAR-Triam and AuSTAR-CD90-Triam 58
Figure 21. Calibration curve of ADMSC AuS-CD90-Triam 59
Figure 22. Representative confocal image shows conjugation of AuS-Triam (green) on the MSC (red) trough Anti-CD90 (Yellow). All bars 62