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1、微米晶片在細胞生物學上的應用,細胞研究: 細胞研究的方向。 細胞生長環境。 細胞微環境的因素。 生物科技的劣勢。 演講者:陳俊男 07.27.2005,生物技術,DNA: agarose 電泳(McDonell, 1977), Southern blot, clone, PCR(1971). RNA: RT-PCR(1988), Northern blot(1994). Protein: SDS page(1960), western blot(1979) Proteomics: 2D Electrophoresis(1970s).,Proteomics Processes,生物科技的缺點:
2、1.耗時 2.費人力 3.高成本 4.無法在微米尺度細操控細胞,細菌 5.無法及時觀徹細胞生理狀況(real time),微米級晶片的新工具- 微流道,微流道晶片(lab-On-a Chip) 陣列式晶片 晶片的應用,55 C.,72 C,95 C.,c BLOOD-ON-A-CHIP,C. right pole :Mitotracker Green FM ; left pole :Mitotracker Red CM-H2XRos. entire cell: DNA-binding dye Hoechst 33342. D. 2.5 hrs later E. Blue region: lat
3、runculin = cause the disruption of actin filament F. PhalloidinAlexa 594-labelled , and 10 min latter , treatment with latrunculin A.,Figure 1 Chemotaxis.,Figure 2 Elastomeric Membranes, (Bovine adrenal capillary endothelial cell),Figure 3. (a) A cell spread on a square 30 um 30 um island; the cell
4、was stimulated with platelet-derived growth factor (PDGF) and stained for F-actin with fluorophore-labeled phalloidin. (b) Cells confined to various shapes (all with areas of 900 um2) were stimulated with PDGF and stained with phalloidin and 4 -6-diamidino-2-phenylindole to visualize F-actin (green)
5、 and nuclei (blue). Images obtained by fluorescence microscopy.,18 min of a living fibroblast after stimulation with PDGF. fibroblast, coated with fibronectin, stained with fluoresceinated phalloidin (C) Endothelial cell, coated with fibronectin, stained with fluoresceinated phalloidin (D) skeletal
6、C2C12 myoblast, coated thrombospondin-1, stained with anti-fascin antibodies . A, B) 30 30 m island; C, D) 40 40 m islands.,Figure 4. Elastomeric silicone microposts - a substrate to cellular contractions. (a) cells adhere to the tips of an array of closely spaced. (b) SEM showing a cell attached to
7、 the entire micropost substrate (c) illustration showing the process for microprinting adhesive proteins on the microposts. (d) Differential interference contrast (top) and immunofluorescence (bottom) micrographs :2 x 2 array of posts printed with fibronectin. (e), cells only attach on the tips. Sca
8、le bars indicate 10 m.,Figure 5. Bovine capillary endothelial cells were initially on rectangular patterns using (SAMs) monolayers surrounded by ethylene-glycolterminated (EG-terminated) SAMs. Application of a cathodic voltage pulse (1.2 V for 30 s) released the cells from the constraints of the mic
9、roislands by desorbing the EG-terminated SAMs and enabling proteins to adsorb from the culture medium that allowed the attachment and movement of cells. The numbers indicate the time elapsed (in minutes) after the voltage pulse.,Figure 6. (a) Agarose is wicked into channels formed by a poly(dimethyl
10、siloxane) stamp sealed against a glass coverslip and allowed to gel before the stamp is peeled off. (b) When cells are seeded onto these substrates containing bowtie-shaped wells, cells attach and culture as either single cells or pairs. (c) The single cell-to-cell contact formed in these pairs can be blocked by fabricating substrates in which the agarose forms a thin wall.,PEG (chemical fusion),Electrical fusion,40 x phase Hela S3,Alexa Fluor 488 phalloidin,Fibroblast,Thank You For Your Attention,感謝 中研院應用科學中心 陳培凌老師 徐昭業學長,
