造血干细胞自动磁分选仪的研制及初步应用
|
摘要
造血干细胞自动磁分选仪采用微型计算机和现代控制技术,通过磁性纳米粒子与CD34单克隆抗体结合,构建磁性纳米粒子生物分子复合物,制成细胞磁分选溶液,进行造血干细胞分离。对分离过程和管道清洗进行全程自动控制,可以批量、安全、精确、快速地对干细胞进行分离,用于从外周血或脐带血中分离出造血干细胞。采用该分离系统后,可以使造血干细胞分离成本成倍地降低。用于临床后可为众多的肿瘤和免疫疾病的患者带来治疗和治愈的机会,用该分选系统分选后,在异体PBSCT中它可去除T细胞、减少和减轻GVHD发生,这种大量细胞磁分选仪采用微型计算机和现代控制技术,对细胞分离和洗涤过程进行全程自动控制,可以批量、安全、高效,快速地对干细胞进行分离。
本论文主要取得了以下结果:
(1)造血干细胞自动磁分选仪采用微型计算机和现代控制技术,对细胞分离和洗涤过程进行全程自动控制。根据仪器的功能及应用,设计该仪器的硬件系统包括磁分选系统,样品传送系统、电脑控制系统和外壳。硬件设计主要完成了分选柱研制、管道设计、外壳设计等工作。造血干细胞自动磁分选仪的硬件系统设计全面、实用,作为一台医疗研究仪器除了具有性能优越,精确耐用等硬性指标外,更具备美观大方,便于操作的特色。
(2)造血干细胞自动磁分选仪是通过电脑编程实现全自动化控制的,电脑控制系统可分为:电磁阀控制、磁块电机控制、蠕动泵控制,每个控制系统又细化为几个部分。控制系统软件编程时要求对仪器运行的每一步骤有精确的描述和表达。仪器运行分为两个大的步骤,一为系统测试和管道润洗步骤,该步骤有34步命令,历时2分钟;一为细胞自动分选步骤,该步骤有89步命令,历时20分钟。整个仪器运行状态时间为25分钟左右。造血干细胞自动磁分选仪的分选从冲洗系统前滤器开始,然后开始标记细胞的分选。
(3)造血干细胞自动磁分选仪采用微型计算机和现代控制技术。实验通过磁性纳米粒子与CD34单克隆抗体结合,构建磁性纳米粒子—生物分子复合物,制成细胞磁分选溶液。磁性纳米粒子大部分粒径都在50-80nm,微粒的分布窄,是混合型包裹,里面包裹了好几个磁性纳米粒子,分散性也比较好,团聚现象不是很严重,具有良好的超顺磁性。在造血干细胞自动磁分选仪上进行造血干细胞分离富集,得到了较好的试验结果,经过流式细胞仪检测获得分离纯度50%以上,分离效率40%以上,体外扩增效果良好,具备较强的实用性。
Hematopoietic stem cell automatic magnetic separation instrument, which we design and development, uses microcomputer and modern control technology. Through a magnetic nanometer granule and the CD34+ monoclonal antibody union to construct the magnetic nanoparticles biological molecular complexes, it makes the magnetic cell sorting solution for hematopoietic stem cell separation. Carrying on the full automatic control to the separation process and the pipe clean, may separate the stem cell batch, safely, precisely and fast. in order to isolate stem cells for the blood or cord blood from the isolated hematopoietic stem cells. Using this separation system, may cause to make the cost of blood stem cell separation reduce multiplied. Can be used in clinical as many tumors and autoimmune diseases in patients brought treatable and curable, use of the election system after the election, in which it allogeneic T cells can be removed to reduce GVHD occurred. Such large magnetic cell sorting adopts micro computer control and modern technology, on the cell separation and washing entire process control, batch, safe, efficient and rapid method to isolate cells.
This paper mainly includes two aspects as follows:
(1) Hematopoietic Stem Cell Sorter automatically using magnetic micro-computersand modern control technology, cell separation and washing the whole process automatic. According to the function and application of equipment, design of the hardware system of the apparatus including magnetic separation system, the sample delivery systems, computer control system and shell. Hardware Design of the main separation column completed development, pipeline design, casing design and so on. Hematopoietic Stem Cell Sorter automatically magnetic hardware system design a comprehensive, practical, as a medical research equipment In addition to superior performance, durability and other hard targets accurately, but also have the elegant, user-friendly features.
(2) Hematopoietic Stem Cell Sorter automatically magnetic computer programmingthrough the control of fully automated, computer-controlled system can be divided into: control solenoid valve, magnetic motor control block, control of peristaltic pump, each control system for parts and refinement. Control system software to run when the requested equipment for each step and describe the precise expression. Instrument operation is divided into two major steps, one for system testing and pipe run washing steps, the step command steps to 34, spend 2 minutes. Second, cellular automata for sorting steps, the step command steps to 89, spent 20 minutes. Machines running the entire time is 25 minutes. Hematopoietic Stem Cell Sorter automatic magnetic separation wash system from the beginning of pre-filter, and then began sorting the labeled cells.
(3) Hematopoietic Stem Cell Sorter automatically using magnetic micro-computers and modern control technology. Experiment through the magnetic nanoparticles with CD34 monoclonal antibody with magnetic nanoparticles to build - complex biological molecules, made of magnetic cell sorting solution. Most of magnetic nanoparticles in the size 50-80nm, narrow distribution of particles is a hybrid package, which packages a number of magnetic nanoparticles, dispersion also better reunion phenomenon is not very serious, a good super-paramagnetic. Hematopoietic stem cells in the automatic magnetic separation apparatus for separation and enrichment of hematopoietic stem cells, have been a better test results, after the separation by flow cytometry to obtain more than 50%purity, more than 40% separation efficiency, good effect in vitro amplification with more practical.
本论文主要取得了以下结果:
(1)造血干细胞自动磁分选仪采用微型计算机和现代控制技术,对细胞分离和洗涤过程进行全程自动控制。根据仪器的功能及应用,设计该仪器的硬件系统包括磁分选系统,样品传送系统、电脑控制系统和外壳。硬件设计主要完成了分选柱研制、管道设计、外壳设计等工作。造血干细胞自动磁分选仪的硬件系统设计全面、实用,作为一台医疗研究仪器除了具有性能优越,精确耐用等硬性指标外,更具备美观大方,便于操作的特色。
(2)造血干细胞自动磁分选仪是通过电脑编程实现全自动化控制的,电脑控制系统可分为:电磁阀控制、磁块电机控制、蠕动泵控制,每个控制系统又细化为几个部分。控制系统软件编程时要求对仪器运行的每一步骤有精确的描述和表达。仪器运行分为两个大的步骤,一为系统测试和管道润洗步骤,该步骤有34步命令,历时2分钟;一为细胞自动分选步骤,该步骤有89步命令,历时20分钟。整个仪器运行状态时间为25分钟左右。造血干细胞自动磁分选仪的分选从冲洗系统前滤器开始,然后开始标记细胞的分选。
(3)造血干细胞自动磁分选仪采用微型计算机和现代控制技术。实验通过磁性纳米粒子与CD34单克隆抗体结合,构建磁性纳米粒子—生物分子复合物,制成细胞磁分选溶液。磁性纳米粒子大部分粒径都在50-80nm,微粒的分布窄,是混合型包裹,里面包裹了好几个磁性纳米粒子,分散性也比较好,团聚现象不是很严重,具有良好的超顺磁性。在造血干细胞自动磁分选仪上进行造血干细胞分离富集,得到了较好的试验结果,经过流式细胞仪检测获得分离纯度50%以上,分离效率40%以上,体外扩增效果良好,具备较强的实用性。
Hematopoietic stem cell automatic magnetic separation instrument, which we design and development, uses microcomputer and modern control technology. Through a magnetic nanometer granule and the CD34+ monoclonal antibody union to construct the magnetic nanoparticles biological molecular complexes, it makes the magnetic cell sorting solution for hematopoietic stem cell separation. Carrying on the full automatic control to the separation process and the pipe clean, may separate the stem cell batch, safely, precisely and fast. in order to isolate stem cells for the blood or cord blood from the isolated hematopoietic stem cells. Using this separation system, may cause to make the cost of blood stem cell separation reduce multiplied. Can be used in clinical as many tumors and autoimmune diseases in patients brought treatable and curable, use of the election system after the election, in which it allogeneic T cells can be removed to reduce GVHD occurred. Such large magnetic cell sorting adopts micro computer control and modern technology, on the cell separation and washing entire process control, batch, safe, efficient and rapid method to isolate cells.
This paper mainly includes two aspects as follows:
(1) Hematopoietic Stem Cell Sorter automatically using magnetic micro-computersand modern control technology, cell separation and washing the whole process automatic. According to the function and application of equipment, design of the hardware system of the apparatus including magnetic separation system, the sample delivery systems, computer control system and shell. Hardware Design of the main separation column completed development, pipeline design, casing design and so on. Hematopoietic Stem Cell Sorter automatically magnetic hardware system design a comprehensive, practical, as a medical research equipment In addition to superior performance, durability and other hard targets accurately, but also have the elegant, user-friendly features.
(2) Hematopoietic Stem Cell Sorter automatically magnetic computer programmingthrough the control of fully automated, computer-controlled system can be divided into: control solenoid valve, magnetic motor control block, control of peristaltic pump, each control system for parts and refinement. Control system software to run when the requested equipment for each step and describe the precise expression. Instrument operation is divided into two major steps, one for system testing and pipe run washing steps, the step command steps to 34, spend 2 minutes. Second, cellular automata for sorting steps, the step command steps to 89, spent 20 minutes. Machines running the entire time is 25 minutes. Hematopoietic Stem Cell Sorter automatic magnetic separation wash system from the beginning of pre-filter, and then began sorting the labeled cells.
(3) Hematopoietic Stem Cell Sorter automatically using magnetic micro-computers and modern control technology. Experiment through the magnetic nanoparticles with CD34 monoclonal antibody with magnetic nanoparticles to build - complex biological molecules, made of magnetic cell sorting solution. Most of magnetic nanoparticles in the size 50-80nm, narrow distribution of particles is a hybrid package, which packages a number of magnetic nanoparticles, dispersion also better reunion phenomenon is not very serious, a good super-paramagnetic. Hematopoietic stem cells in the automatic magnetic separation apparatus for separation and enrichment of hematopoietic stem cells, have been a better test results, after the separation by flow cytometry to obtain more than 50%purity, more than 40% separation efficiency, good effect in vitro amplification with more practical.
引文
[1]刘虹,曹水,任秀宝,张澎,安秀梅,郝希山.免疫磁性分选系统大量分选CD34+细胞的实验研究[J].中国实验血液学杂志,2004.798-802
[2]胡振红.磁性细胞分离技术的原理和研究进展[J].华南国防医学杂志,2001.28-32.
[3]JooH.Kang,Je-Kyun Park.Technical Paper on Microfluidic Devices-Cell Separation Technology[J].APBN.2005
[4]Sixin LI,Jiancheng ZHANG,Yue SHEN,BO NI,Jingang ZHANG..Synthesis and Properties of Magnetic Nanoparticles[J].J.M ater.Sci.Technol,2006.659-664
[5]洪缨.外周血造血干细胞移植概况[M].国外医学输血及血液学分册,2005.422-427
[6]Wei Chen,Hebai Shen,Xingyu Li,,Nengqin Jia,Jianming Xu.Synthesis of immunomagnetic nanoparticles and their application in the separation and purification of CD34+ hematopoietic stem cells[J].Applied Surface Science(2006)
[7]张阳德等.纳米生物材料学[M].第二版.北京.化学工业出版社,2005.2-50
[8]杨文胜 高明远 白玉白等.纳米材料与生物技术[M].北京.化学工业出版社,2005.48-52
[9]张立德 解思深等.纳米材料和纳米结构[M].北京.化学工业出版社,2005.147-169
[10]施利毅等,纳米材料[M].上海,华东理工大学出版社,2007.59-61
[11]柯扬船(美)皮特·斯壮,聚合物-无机纳米复合材料[M].北京,化学工业出版社,2003.4-18
[12]张立德,牟季美著,纳米材料和纳米结构[M].北京,科学出版社,2001.78-95
[13]徐祖顺,易昌凤,聚合物纳米粒子[M].北京,化学工业出版社,2006.11-16
[14]孙彦,生物分离工程[M],北京,化学工业出版社,1998,1-19
[15]Tartai P,Morales M P,Verdaguer S,Carreno T,Sema C J.The preparation of magnetic nanoparticles for applications in biomedicine.[J]Phys.D:Appl.Phys.2003,36:182-197.
[16]王强斌.功能高分子磁性微球的制备及其生物学应用研究[D].上海:华东理 工大学,2001
[17]高虹.纳米高分子聚合物在肿瘤靶向化疗中的初步应用研究[D].上海:复旦大学,2004
[18]吴伟等,磁性纳米粒子在生物传感器中的应用研究进展,[J]化学通报,在线预览版
[19]张银燕 尹衍升 张金升 马来鹏.纳米Fe_3O_4磁性液体稳定性的研究[J].化学物理学报,2004,17(1):83-87
[20]Massart,R.Preparation of aqueous magnetic liquids in alkaline and acidic media.[J]Magnetics,IEEE Transactions on.1981,17(2):1247-1248
[21]M.P.Morales,S.Veintemillas,M.I.Montero,C.J.Serna,A.Roig,LI.Casas,B.Martinez and F.Sandiumenge.Surface and internal spin canting in γ-Fe2O3nanoparticles.[J]Chem Mater.1999,11(8):3058-3064.
[22]L.Wang,J.Luo,M.M.Maye,Q.Fan,Q.Rendeng,M.H.Engelhard,C.Wang,Y.Lin and C.-J.Zhong.Iron oxide-gold core-shell nanoparticles and thin film assembly [J]Mater Chem.2005,15(18):1821-1832
[23]Huachang Lu,Guangshun Yi,Shuying Zhao,Depu Chen,Liang-Hong Guo and Jing Cheng.Synthesis and characterization of multi-functional nanoparticles possessing magnetic,up-conversion fluorescence and bio-affinity properties
[24]Kyoungja Woo,Jangwon Hong,Sungmoon Choi,Hae-Weon Lee,Jae-Pyoung Ahn,Chul Sung Kim,and Sang Won Lee.Easy Synthesis and Magnetic Properties of Iron Oxide Nanoparticles[J]Chem.Mater.;2004,16(14):2814-2818;
[25]Yu,W.W.,Falkner,J.C.,Yavuz,C.T.,and Colvin,V.L.Synthesis of monodisperse iron oxide nanocrystals by thermal decomposition of iron carboxylate salts.[J]Chem.Commun.2004,20(14):2306-2307
[26]Ulf Wiedwald,Dr,Kai Fauth,Dr,Markus Heβler,Hans-Gerd Boyen,Dr,etal.From Colloidal Co/CoO Core/Shell Nanoparticles to Arrays of Metallic Nanomagnets:Surface Modification and Magnetic Properties[J]Chem Phys,2005,6(12):2522-2526.
[27]Zhen Li,Qiao Sun,Mingyuan Gao Preparation of Water-soluble Magnetite Nanocrystals by Refluxing Ferric Hydrated Salts in 2-Pyrrolidone:Mechanism Leading to Fe304[J]Angew Chem.Int Ed,2005,44(3):123-126
[28]邱广明,邱广亮,磁性明胶复合磁性微球的制备和性质,[J]食品科学1998,19(11):7-10
[29]刘春艳,金志兰,栗淑媛,邱广亮.磁性琼脂糖复合微球的制备和性质[J]精 细化工1999,16(1):38-42
[30]安小宁,苏致兴.高磁性壳聚糖微粒的制备与应用[J]兰州大学学报,2001,37(2):100-104.
[31]B aharT,CelibiS S.J Appl Polym Sci,1999,72:805
[32]DekkerR F M.Appl Biochem,1989,22:28
[33]CuyperM D,JonianM.Langmuir,1990,7:64 7
[34]Gupta P K,H ongC T,L amC F,etal.Int J Pharm,1988,43:161
[35]王强斌,占宏晨,朱以华,等.羧基磁性高分子微球的制备和表征[J].化学世界,2001,8(1):410-413.
[36]刘学涌,丁小斌,郑朝晖,等.分散聚合制备聚苯乙烯/聚氧乙烯两亲磁性高分子微球..[J]合成化学,2001,9(4):281-284.
[37]邱广明,章贤明,孙宗华.磁性聚苯乙烯微球的合成和特性.[J]高分子材料科学与工程,1993(2):38-43.
[38]阳承利.磁性PMMA微球的制备与表面修饰[J]中国科学(B辑,化学),2004,34(4):265-269.
[39]何虹,杨俊,王亭杰,金涌.悬浮聚合法制备磁性微球的粒度分布特性.[J]高校化学工程学报.2001,15(3):277-281
[40]Tsuruta M.Manufacture of magnetic microspore.[J]P Pat.1987,09(5):27
[41]D eslandesY.[J]App PoLym Sci,1987,(34):2249
[42]万惠文,张强,饶微.聚合物粒子的制备[J].武汉工业大学学报,1999,21(1):22-24.
[43]Sole nee Hajna J USP 4421660,1983
[44]邱广明,杨春雁,孙宗华,等.单分散亚微米级磁性微球的合成[J].功能高分子学报.1996,9(4):565-571
[45]邱广明,章贤明,孙宗华.磁性聚苯乙烯微球的合成和特性.高分子材料科学与工程,1993(2):38-43.
[46]Dreseo P A.Prepartion and Properties of Magnetite and Polymer Magnetite Nanopartieles[J]Langmuir,1999,15(6):1945-1951
[47]Danniel J C,Schuppiser J L.Magnetic Polymer Latex and Preparation Process[P].US4358388,1982-11-09
[48]Sauzedde F,Elaissari A,Piehot C.Hydrophille Magnetic Polymer Latexes I: Adsorption of Magneti Iron Oxide Nanopartieles onto Various Cationic Latexes[J]Colloid and Polymer Science, 1999,277(10): 846-855
[49]Rabelo D, Lima E C D, Tavares Filho N., etal. synthesis of Manganese Ferrite Nanopartieles in Maeroporous Styrenedivinylbenzene Copolymer [J] Journal of Magnetism and Magnetie Materials, 2004, 11(10):272-276
[50] Wang P H. Pan C Y. Polymer Metal Composite Mierospheres Preparation and Characterization of poly(StcoAN)Ni Mierospheres [J]European Polymer Journal, 2000, 36(10): 2297-2300
[51]Koneracka M ,Kopcansky P,Timko M ,Ramchand C N ,Sequeim A D ,Trevan M.Direct binding procedure of proteins and enzymest of immagnetic particles. [J] Mol.Catal.B Enzym.2002,18(1): 13-18.
[52] Huang S H, Liao M H, Chen D H. Direct binding and characterization of lipase onto magnetic nanoparticles. [J]Biotechnol.Prog.2003,19(7):1095-1100.
[53]Koneracka M, Kopcansky P, Antalik M, Timko M, Ramchand C N, Lobo D, Mehta R V,Upadhyay R V .Immobilization of proteins and enzymest of inemagnetic particles. [J]Magn.Magn .Mater. 1999,201(14):427-430.
[54] Garcia III A, Oh S, Engler C R. Celhdase immobilization on Fe304 and characterization. [J]Biotechnol Bioeng 1989,33(8):321-326.
[55]AkgolS, Kacar,Y ,DenizliA ,AricaM Y.Hydrolysis of sucrosebyinvertase immobilized onto novel magnetic polyvinyl alcohol microspheres. [J]Food Chem.2001,74(9):281-288.
[56]Bilkova Z, Slovakova M, Lycka A, Horak D, Lenfeld J, Turkova J, Churacek J. Oriented immobilization of galactose oxidase to bead and magnetic bead cellulose and poly(HE M A- co- ED MA) and magnetic poly( HEMA-co-EDMA) mierospheres [J]Chromatogr. B 2002,770(3):25-34.
[57]DyalA ,Loos K ,Noto M ,etal .Activity of and idarugosalipase immobilizedon γFe203 magnetic nanopartieles [J] Am.Chem.Soc.2003,125(11):1684-1685.
[58]Guo Z, Bai S, Sun Y. Preparation and characterization of immobilized lipase on magnetic hydrophobic mierospheres [J] EnzymeMicrob. Technol. 2003,32(5): 776-782.
[59] Chen JP,Su D H.Latex particles with thermo-flocculation and magnetic properties for immobilization of achymotrypsin.[J]Biotechnol.Prog.2001,17(4):369-375
[60] Betancor L, Fuentes M, Dellamora Ortiz G, Lopez-Gallego F, Hidalgo A, Alonso-Morales N,Mateo C ,Guisan JM ,Femandez-Lafuente R .Dextran aldehyde coating of glucose oxidase immobilized on magnetic nanoparticles prevent sitsin activation gasbubbles. [J]Mol.Catal.B-Enzym.2005,32(4):97-101.
[61]Bahar T, Celebi S S .Immobilization of glucoamylaseon magnetic poly(styrene)particles. [J]Appl. Polym.Sci.1999,72(11):69-73.
[62]Kondo A,Fukuda H.Preparation of thermosensitive magnetic hydrogel microspheres and application to enzyme immobilization. [J]Ferment.Bioeng.1997,4(10): 337-341.
[63] Varlan A R, Sansen W. Covalent enzyme immobilization on paramagnetic polyacrolein beads.Biosens. [J]Bioelectron.1996,11(2):443-448.
[64]Ritich B, Spanova A, Ohlashennyy Y .Characterization of deoxyribonuceasel immobilizedon magnetic hydrophilic polymer particles. [J] Chromatogr.B 2002,774(6):25-31.
[65] Safarik I, In scientific and clinical applications of magnetic carriers. New York and London: Plenum Press, 1997. p323.
[66]Sestier C, Roger J, Pons J N. Electrophoresis, 1998,19:1220.
[67] Gupta A K, Curits A S G. Biomaterials, 2004, 25: 3029.
[68]Liao M H, Chen D H .Immobilization of yeast alcoholde hydrogenaseon magnetic nanoparticles for improving its stability. [J]Biotechnol. Let. 2001, 23 (4):1723-1727.
[69] Lei H, Wang W, Chen L L, Li X C, Yi B, Deng L. The preparation and catalytically active characterization of papain immobilizedon magnetic composite microspheres. [J]Enzyme Microb.Technol.2004,35(7):15-21.
[70] Xu X L, Friedman G, Hunfeld K D, Majetich S A, Asher S A. Superparamagnetic photonic crystals.[J]Adv. Mater.2001,13(8):1681-1684.
[71]Rana S, White P, Bradley M. Synthesis of magnetic beads for solid phase synthesisan dreaction scavenging.[J]Tetrahedron Let.1999,40(8):8137-8140.
[72] Safarikova M, Sefarik 1. Magnetic solid-phase extraction. [J]Magn Magn Mater 1999,194(1):108-112
[73] Yang M, Li H L. Determination of trace hydrazine by differential pulse voltammetry using magnetic microspheres. [J]Talanta 2001,55(11):479-484.
[74]Takafuji M,lde S, Ihara H, Xu Z H. Preparation of poly(1-vinylimidazole) grafted magnetic nanoparticles and their application for removal of mealions. [J]Chem.Mater.2004,16(9):1977-1983.
[75]Cherepy N J,Lison D B,Lovejoy J A.Ultrafast studies of photoexcited electron dynamics in γ 2and α 2Fe2O3 semiconductor nanoparticles[J]Phys Chem B,1998,102(10):770-776.
[76]P.Tartaj,M.P.Morales,T.Gonza lez-Carreno,S.Veintemillas-Verdaguer,C.J.Serna,Advances in magnetic nanoparticles for biotechnology applications,[J]Journal of Magnetism and Magnetic Materials.2005,28(5):290-291
[77]Murakami Y,Ito M,Takasu Y.Synthesis of nobel metal oxide ultrafine particles by sol gel method.[J]Hyomen,1994,32(7):486-494.
[78]Ichinose N.Application of ultrafine powder ceramics[J]Hyomen Kagaku,1992,13(4):190-197.
[79]张效岩,王英,张亚非;磁性纳米粒子的制备及应用[J]磁性材料及器件.2004,06(1):11-14
[80]Boutonnet M,etal.the preparation of monodisperse colloidal metal particles from microemulsions[J].Colloids and surfaces,1982,5(18)209 -225.
[81]Barnickel P.,etal.Size tailoring of silver colloids by reduction in W/O microemulsions[J].Journal of colloid and interface science,1992,148(7):80-90
[82]覃兴华,卢迪芬,反胶团微乳液法制备超微细颗粒的研究进展[J],化工新型材料,1998,12(2):27-29
[83]崔正刚,殷福珊,微乳化技术及应用[M],北京:中国轻工业出版社,1999.
[84]Jurgons,R.,Seliger,C.Hilpert,A.Trahms,L.Odenbach,S.Alexiou,C.[J]Phys.Condens.Matter.2006,18(4):2893
[85]Gu,H.W.;Xu,K.M.;Xu,C.J.;Xu,B.[J]Chem.Commun.,2006,19(1):941-949
[86]Aspden,T.J.;Mason,J.D.Jones,N.S.[J]Pharm.Sci,1997,86(2):509
[87]李强,高镰,栾伟玲,等.纳米ZnO制备工艺中ζ电位与分散性的关系[J]无机材料学报,1999,14(5)813-817
[88]黄小龙,张黎明.壳聚糖基载药纳米微粒制备研究进展[J].功能高分子学报,2003,16(4):593-598
[89]Yang,C.C.;Chen,D.H.Preparation and adsorption properties of monodisperse chitosan-bound Fe3O4 magnetic nanoparticles for removal of Cu(Ⅱ)ions[J].Colloid Interf.Sci.,2005,283(4):446-451
[90]Crsi,K.;Chellat,F.;Yahia,L Hocine;Fernandes J.C.Mesenchymal stem cells, MG63 and HEK293 transfection using chitosan-DNA nanoparticles.Biomaterials.2003;24(5):1255-1264.
[91]贾芳,孟昭兴,蒋燕,王华,刘伯里.手性5[(1S)-内型-冰片氧基]-2(5H)-呋喃酮与硫酚(醇)的不对称Michael加成反应北京师范大学学报(自然科学版),2000,36(2):225-260
[92]A.S.Dilmen,Z.Mustafaeva,M.Demchenko,T.Kaneko,Y.Osada,M.Mustafaev Water-soluble covalent conjugates of bovine serum albumin with anionic poly(N-isopropyl-acrylamide) and their immunogenicity[J]Biomaterials 2001,14(22):2383-2392,
[93]Merril CR,Switzer RC,Van Keuren ML.Trace polypeptides in cellular extracts and human body fluids detected by two-dimensional electrophoresis and a highly sensitive silver stain.Proc Natl Acad Sci U S A.1979 Sep;76(9):4335-4339
[94]Chu,M.Q.;Song,X.;Cheng,D.;Liu,S.P.;Zhu,Preparation of quantum dot-coated magnetic polystyrene nanospheres for cancer cell labelling and separation.[J].Nanotechnology.2006,17:3268
[95]Tanima,B.;Susmita M.;Singh,K.Preparation,characterization and biodistribution of ultrafine chitosan nanoparticles[J].Int Pharm,2002,243(12):93-
[96]蒋挺大.甲壳素.北京:北京工业出版社,2003:146-156
[97]罗志敏 马秀玲,陈盛,钱伟.磁性壳聚糖-聚丙烯酸微球的制备及表征[J].化学通报,2005,68(7):551-554.
[98]Peng,X.H.;Zhang,L.N.Surface fabrication of hollow microspheres from N-methylated chitosan cross-linked with gultaraldehyde[J]Langmuir,2005,21(9):1091-1095
[99]Zhi,J.;Wang,Y.J.;Lu,Y.C.;Ma,J.Y.;Luo,G.S.React.Funct.Polym.,2006,66:1552
[100]Peter,T.;Jr.Adv.Protein Chem.,1985,37:161-245
[101]Lu,C.H.;Luo,C.Q.;Cao,W.X.Colloid and Surface B,2002,25:19
[102]Eftink,M.R.;Zajicek,J.L.;Ghiron,C.A.Biochim.Biophys.Acta.,1977,491:473
[103]魏晓芳 丁西明 刘会洲.pH诱导牛血清白蛋白芳香氨基酸残基微环境变化的光谱分析[J]光谱学及光谱分析,2000,20(4):556-558
[104]Peng.Z.G,Hidajat.K,Uddin.M.S..Adsorpton.of bovine serum albumin on nanosized magnetic particles.[J]Journal of Colloid and Interface Science,2004, 271:277-283.
[105]Terry E.Thomas,Cindy L.Miller,and Connie J.Eaves.Purification of Hematopoietic Stem Cells for Further Biological Study.A Companion to Methods in Enzymology 17,202-218(1999)
[106]唐佩弦.造血干细胞研究发展历史引发的思考.中国实验血液学杂志2005;13(5):723-732
[101]Halens S,Kohn DB.Gene therapy using hematopoietic stem cells:sisyphus approaches the cerst[J].Human Gone Therapy.2000.11(9):1259-1267
[107]徐勇,霍梅.免疫磁珠分离及流式细胞仪分选纯化外周血CD34+/CD90+干细胞.临床检验杂志2004年第22卷第4期
[108]Motoki Kuhara,Haruko Takeyama,Tsuyoshi Tanaka,et al.Magnetic Cell Separation.Using Antibody Binding with Protein A Expressed on Bacterial Magnetic Particles.Anal.chem.2004,76,6207-6213
[109]L.P.Zhu,X.Q.Chen.(editors) The experimental methods of immunology.People' s Military Medical Press,2003,P.70.
[110]Kimitaka Kato and Andreas Radbruch.Isolation and characterization of CD34+hematopietic stem cells from human peripheral blood by high-gradient magnetic cell sorting.Cytometry14:384-392(1993)
[111]熊玉宁,滕爱萍,王景文,沈悌.不同来源人CD34+细胞体外集落培养的研究及意义.中华内科杂志,2001年7月第40卷第7期
[2]胡振红.磁性细胞分离技术的原理和研究进展[J].华南国防医学杂志,2001.28-32.
[3]JooH.Kang,Je-Kyun Park.Technical Paper on Microfluidic Devices-Cell Separation Technology[J].APBN.2005
[4]Sixin LI,Jiancheng ZHANG,Yue SHEN,BO NI,Jingang ZHANG..Synthesis and Properties of Magnetic Nanoparticles[J].J.M ater.Sci.Technol,2006.659-664
[5]洪缨.外周血造血干细胞移植概况[M].国外医学输血及血液学分册,2005.422-427
[6]Wei Chen,Hebai Shen,Xingyu Li,,Nengqin Jia,Jianming Xu.Synthesis of immunomagnetic nanoparticles and their application in the separation and purification of CD34+ hematopoietic stem cells[J].Applied Surface Science(2006)
[7]张阳德等.纳米生物材料学[M].第二版.北京.化学工业出版社,2005.2-50
[8]杨文胜 高明远 白玉白等.纳米材料与生物技术[M].北京.化学工业出版社,2005.48-52
[9]张立德 解思深等.纳米材料和纳米结构[M].北京.化学工业出版社,2005.147-169
[10]施利毅等,纳米材料[M].上海,华东理工大学出版社,2007.59-61
[11]柯扬船(美)皮特·斯壮,聚合物-无机纳米复合材料[M].北京,化学工业出版社,2003.4-18
[12]张立德,牟季美著,纳米材料和纳米结构[M].北京,科学出版社,2001.78-95
[13]徐祖顺,易昌凤,聚合物纳米粒子[M].北京,化学工业出版社,2006.11-16
[14]孙彦,生物分离工程[M],北京,化学工业出版社,1998,1-19
[15]Tartai P,Morales M P,Verdaguer S,Carreno T,Sema C J.The preparation of magnetic nanoparticles for applications in biomedicine.[J]Phys.D:Appl.Phys.2003,36:182-197.
[16]王强斌.功能高分子磁性微球的制备及其生物学应用研究[D].上海:华东理 工大学,2001
[17]高虹.纳米高分子聚合物在肿瘤靶向化疗中的初步应用研究[D].上海:复旦大学,2004
[18]吴伟等,磁性纳米粒子在生物传感器中的应用研究进展,[J]化学通报,在线预览版
[19]张银燕 尹衍升 张金升 马来鹏.纳米Fe_3O_4磁性液体稳定性的研究[J].化学物理学报,2004,17(1):83-87
[20]Massart,R.Preparation of aqueous magnetic liquids in alkaline and acidic media.[J]Magnetics,IEEE Transactions on.1981,17(2):1247-1248
[21]M.P.Morales,S.Veintemillas,M.I.Montero,C.J.Serna,A.Roig,LI.Casas,B.Martinez and F.Sandiumenge.Surface and internal spin canting in γ-Fe2O3nanoparticles.[J]Chem Mater.1999,11(8):3058-3064.
[22]L.Wang,J.Luo,M.M.Maye,Q.Fan,Q.Rendeng,M.H.Engelhard,C.Wang,Y.Lin and C.-J.Zhong.Iron oxide-gold core-shell nanoparticles and thin film assembly [J]Mater Chem.2005,15(18):1821-1832
[23]Huachang Lu,Guangshun Yi,Shuying Zhao,Depu Chen,Liang-Hong Guo and Jing Cheng.Synthesis and characterization of multi-functional nanoparticles possessing magnetic,up-conversion fluorescence and bio-affinity properties
[24]Kyoungja Woo,Jangwon Hong,Sungmoon Choi,Hae-Weon Lee,Jae-Pyoung Ahn,Chul Sung Kim,and Sang Won Lee.Easy Synthesis and Magnetic Properties of Iron Oxide Nanoparticles[J]Chem.Mater.;2004,16(14):2814-2818;
[25]Yu,W.W.,Falkner,J.C.,Yavuz,C.T.,and Colvin,V.L.Synthesis of monodisperse iron oxide nanocrystals by thermal decomposition of iron carboxylate salts.[J]Chem.Commun.2004,20(14):2306-2307
[26]Ulf Wiedwald,Dr,Kai Fauth,Dr,Markus Heβler,Hans-Gerd Boyen,Dr,etal.From Colloidal Co/CoO Core/Shell Nanoparticles to Arrays of Metallic Nanomagnets:Surface Modification and Magnetic Properties[J]Chem Phys,2005,6(12):2522-2526.
[27]Zhen Li,Qiao Sun,Mingyuan Gao Preparation of Water-soluble Magnetite Nanocrystals by Refluxing Ferric Hydrated Salts in 2-Pyrrolidone:Mechanism Leading to Fe304[J]Angew Chem.Int Ed,2005,44(3):123-126
[28]邱广明,邱广亮,磁性明胶复合磁性微球的制备和性质,[J]食品科学1998,19(11):7-10
[29]刘春艳,金志兰,栗淑媛,邱广亮.磁性琼脂糖复合微球的制备和性质[J]精 细化工1999,16(1):38-42
[30]安小宁,苏致兴.高磁性壳聚糖微粒的制备与应用[J]兰州大学学报,2001,37(2):100-104.
[31]B aharT,CelibiS S.J Appl Polym Sci,1999,72:805
[32]DekkerR F M.Appl Biochem,1989,22:28
[33]CuyperM D,JonianM.Langmuir,1990,7:64 7
[34]Gupta P K,H ongC T,L amC F,etal.Int J Pharm,1988,43:161
[35]王强斌,占宏晨,朱以华,等.羧基磁性高分子微球的制备和表征[J].化学世界,2001,8(1):410-413.
[36]刘学涌,丁小斌,郑朝晖,等.分散聚合制备聚苯乙烯/聚氧乙烯两亲磁性高分子微球..[J]合成化学,2001,9(4):281-284.
[37]邱广明,章贤明,孙宗华.磁性聚苯乙烯微球的合成和特性.[J]高分子材料科学与工程,1993(2):38-43.
[38]阳承利.磁性PMMA微球的制备与表面修饰[J]中国科学(B辑,化学),2004,34(4):265-269.
[39]何虹,杨俊,王亭杰,金涌.悬浮聚合法制备磁性微球的粒度分布特性.[J]高校化学工程学报.2001,15(3):277-281
[40]Tsuruta M.Manufacture of magnetic microspore.[J]P Pat.1987,09(5):27
[41]D eslandesY.[J]App PoLym Sci,1987,(34):2249
[42]万惠文,张强,饶微.聚合物粒子的制备[J].武汉工业大学学报,1999,21(1):22-24.
[43]Sole nee Hajna J USP 4421660,1983
[44]邱广明,杨春雁,孙宗华,等.单分散亚微米级磁性微球的合成[J].功能高分子学报.1996,9(4):565-571
[45]邱广明,章贤明,孙宗华.磁性聚苯乙烯微球的合成和特性.高分子材料科学与工程,1993(2):38-43.
[46]Dreseo P A.Prepartion and Properties of Magnetite and Polymer Magnetite Nanopartieles[J]Langmuir,1999,15(6):1945-1951
[47]Danniel J C,Schuppiser J L.Magnetic Polymer Latex and Preparation Process[P].US4358388,1982-11-09
[48]Sauzedde F,Elaissari A,Piehot C.Hydrophille Magnetic Polymer Latexes I: Adsorption of Magneti Iron Oxide Nanopartieles onto Various Cationic Latexes[J]Colloid and Polymer Science, 1999,277(10): 846-855
[49]Rabelo D, Lima E C D, Tavares Filho N., etal. synthesis of Manganese Ferrite Nanopartieles in Maeroporous Styrenedivinylbenzene Copolymer [J] Journal of Magnetism and Magnetie Materials, 2004, 11(10):272-276
[50] Wang P H. Pan C Y. Polymer Metal Composite Mierospheres Preparation and Characterization of poly(StcoAN)Ni Mierospheres [J]European Polymer Journal, 2000, 36(10): 2297-2300
[51]Koneracka M ,Kopcansky P,Timko M ,Ramchand C N ,Sequeim A D ,Trevan M.Direct binding procedure of proteins and enzymest of immagnetic particles. [J] Mol.Catal.B Enzym.2002,18(1): 13-18.
[52] Huang S H, Liao M H, Chen D H. Direct binding and characterization of lipase onto magnetic nanoparticles. [J]Biotechnol.Prog.2003,19(7):1095-1100.
[53]Koneracka M, Kopcansky P, Antalik M, Timko M, Ramchand C N, Lobo D, Mehta R V,Upadhyay R V .Immobilization of proteins and enzymest of inemagnetic particles. [J]Magn.Magn .Mater. 1999,201(14):427-430.
[54] Garcia III A, Oh S, Engler C R. Celhdase immobilization on Fe304 and characterization. [J]Biotechnol Bioeng 1989,33(8):321-326.
[55]AkgolS, Kacar,Y ,DenizliA ,AricaM Y.Hydrolysis of sucrosebyinvertase immobilized onto novel magnetic polyvinyl alcohol microspheres. [J]Food Chem.2001,74(9):281-288.
[56]Bilkova Z, Slovakova M, Lycka A, Horak D, Lenfeld J, Turkova J, Churacek J. Oriented immobilization of galactose oxidase to bead and magnetic bead cellulose and poly(HE M A- co- ED MA) and magnetic poly( HEMA-co-EDMA) mierospheres [J]Chromatogr. B 2002,770(3):25-34.
[57]DyalA ,Loos K ,Noto M ,etal .Activity of and idarugosalipase immobilizedon γFe203 magnetic nanopartieles [J] Am.Chem.Soc.2003,125(11):1684-1685.
[58]Guo Z, Bai S, Sun Y. Preparation and characterization of immobilized lipase on magnetic hydrophobic mierospheres [J] EnzymeMicrob. Technol. 2003,32(5): 776-782.
[59] Chen JP,Su D H.Latex particles with thermo-flocculation and magnetic properties for immobilization of achymotrypsin.[J]Biotechnol.Prog.2001,17(4):369-375
[60] Betancor L, Fuentes M, Dellamora Ortiz G, Lopez-Gallego F, Hidalgo A, Alonso-Morales N,Mateo C ,Guisan JM ,Femandez-Lafuente R .Dextran aldehyde coating of glucose oxidase immobilized on magnetic nanoparticles prevent sitsin activation gasbubbles. [J]Mol.Catal.B-Enzym.2005,32(4):97-101.
[61]Bahar T, Celebi S S .Immobilization of glucoamylaseon magnetic poly(styrene)particles. [J]Appl. Polym.Sci.1999,72(11):69-73.
[62]Kondo A,Fukuda H.Preparation of thermosensitive magnetic hydrogel microspheres and application to enzyme immobilization. [J]Ferment.Bioeng.1997,4(10): 337-341.
[63] Varlan A R, Sansen W. Covalent enzyme immobilization on paramagnetic polyacrolein beads.Biosens. [J]Bioelectron.1996,11(2):443-448.
[64]Ritich B, Spanova A, Ohlashennyy Y .Characterization of deoxyribonuceasel immobilizedon magnetic hydrophilic polymer particles. [J] Chromatogr.B 2002,774(6):25-31.
[65] Safarik I, In scientific and clinical applications of magnetic carriers. New York and London: Plenum Press, 1997. p323.
[66]Sestier C, Roger J, Pons J N. Electrophoresis, 1998,19:1220.
[67] Gupta A K, Curits A S G. Biomaterials, 2004, 25: 3029.
[68]Liao M H, Chen D H .Immobilization of yeast alcoholde hydrogenaseon magnetic nanoparticles for improving its stability. [J]Biotechnol. Let. 2001, 23 (4):1723-1727.
[69] Lei H, Wang W, Chen L L, Li X C, Yi B, Deng L. The preparation and catalytically active characterization of papain immobilizedon magnetic composite microspheres. [J]Enzyme Microb.Technol.2004,35(7):15-21.
[70] Xu X L, Friedman G, Hunfeld K D, Majetich S A, Asher S A. Superparamagnetic photonic crystals.[J]Adv. Mater.2001,13(8):1681-1684.
[71]Rana S, White P, Bradley M. Synthesis of magnetic beads for solid phase synthesisan dreaction scavenging.[J]Tetrahedron Let.1999,40(8):8137-8140.
[72] Safarikova M, Sefarik 1. Magnetic solid-phase extraction. [J]Magn Magn Mater 1999,194(1):108-112
[73] Yang M, Li H L. Determination of trace hydrazine by differential pulse voltammetry using magnetic microspheres. [J]Talanta 2001,55(11):479-484.
[74]Takafuji M,lde S, Ihara H, Xu Z H. Preparation of poly(1-vinylimidazole) grafted magnetic nanoparticles and their application for removal of mealions. [J]Chem.Mater.2004,16(9):1977-1983.
[75]Cherepy N J,Lison D B,Lovejoy J A.Ultrafast studies of photoexcited electron dynamics in γ 2and α 2Fe2O3 semiconductor nanoparticles[J]Phys Chem B,1998,102(10):770-776.
[76]P.Tartaj,M.P.Morales,T.Gonza lez-Carreno,S.Veintemillas-Verdaguer,C.J.Serna,Advances in magnetic nanoparticles for biotechnology applications,[J]Journal of Magnetism and Magnetic Materials.2005,28(5):290-291
[77]Murakami Y,Ito M,Takasu Y.Synthesis of nobel metal oxide ultrafine particles by sol gel method.[J]Hyomen,1994,32(7):486-494.
[78]Ichinose N.Application of ultrafine powder ceramics[J]Hyomen Kagaku,1992,13(4):190-197.
[79]张效岩,王英,张亚非;磁性纳米粒子的制备及应用[J]磁性材料及器件.2004,06(1):11-14
[80]Boutonnet M,etal.the preparation of monodisperse colloidal metal particles from microemulsions[J].Colloids and surfaces,1982,5(18)209 -225.
[81]Barnickel P.,etal.Size tailoring of silver colloids by reduction in W/O microemulsions[J].Journal of colloid and interface science,1992,148(7):80-90
[82]覃兴华,卢迪芬,反胶团微乳液法制备超微细颗粒的研究进展[J],化工新型材料,1998,12(2):27-29
[83]崔正刚,殷福珊,微乳化技术及应用[M],北京:中国轻工业出版社,1999.
[84]Jurgons,R.,Seliger,C.Hilpert,A.Trahms,L.Odenbach,S.Alexiou,C.[J]Phys.Condens.Matter.2006,18(4):2893
[85]Gu,H.W.;Xu,K.M.;Xu,C.J.;Xu,B.[J]Chem.Commun.,2006,19(1):941-949
[86]Aspden,T.J.;Mason,J.D.Jones,N.S.[J]Pharm.Sci,1997,86(2):509
[87]李强,高镰,栾伟玲,等.纳米ZnO制备工艺中ζ电位与分散性的关系[J]无机材料学报,1999,14(5)813-817
[88]黄小龙,张黎明.壳聚糖基载药纳米微粒制备研究进展[J].功能高分子学报,2003,16(4):593-598
[89]Yang,C.C.;Chen,D.H.Preparation and adsorption properties of monodisperse chitosan-bound Fe3O4 magnetic nanoparticles for removal of Cu(Ⅱ)ions[J].Colloid Interf.Sci.,2005,283(4):446-451
[90]Crsi,K.;Chellat,F.;Yahia,L Hocine;Fernandes J.C.Mesenchymal stem cells, MG63 and HEK293 transfection using chitosan-DNA nanoparticles.Biomaterials.2003;24(5):1255-1264.
[91]贾芳,孟昭兴,蒋燕,王华,刘伯里.手性5[(1S)-内型-冰片氧基]-2(5H)-呋喃酮与硫酚(醇)的不对称Michael加成反应北京师范大学学报(自然科学版),2000,36(2):225-260
[92]A.S.Dilmen,Z.Mustafaeva,M.Demchenko,T.Kaneko,Y.Osada,M.Mustafaev Water-soluble covalent conjugates of bovine serum albumin with anionic poly(N-isopropyl-acrylamide) and their immunogenicity[J]Biomaterials 2001,14(22):2383-2392,
[93]Merril CR,Switzer RC,Van Keuren ML.Trace polypeptides in cellular extracts and human body fluids detected by two-dimensional electrophoresis and a highly sensitive silver stain.Proc Natl Acad Sci U S A.1979 Sep;76(9):4335-4339
[94]Chu,M.Q.;Song,X.;Cheng,D.;Liu,S.P.;Zhu,Preparation of quantum dot-coated magnetic polystyrene nanospheres for cancer cell labelling and separation.[J].Nanotechnology.2006,17:3268
[95]Tanima,B.;Susmita M.;Singh,K.Preparation,characterization and biodistribution of ultrafine chitosan nanoparticles[J].Int Pharm,2002,243(12):93-
[96]蒋挺大.甲壳素.北京:北京工业出版社,2003:146-156
[97]罗志敏 马秀玲,陈盛,钱伟.磁性壳聚糖-聚丙烯酸微球的制备及表征[J].化学通报,2005,68(7):551-554.
[98]Peng,X.H.;Zhang,L.N.Surface fabrication of hollow microspheres from N-methylated chitosan cross-linked with gultaraldehyde[J]Langmuir,2005,21(9):1091-1095
[99]Zhi,J.;Wang,Y.J.;Lu,Y.C.;Ma,J.Y.;Luo,G.S.React.Funct.Polym.,2006,66:1552
[100]Peter,T.;Jr.Adv.Protein Chem.,1985,37:161-245
[101]Lu,C.H.;Luo,C.Q.;Cao,W.X.Colloid and Surface B,2002,25:19
[102]Eftink,M.R.;Zajicek,J.L.;Ghiron,C.A.Biochim.Biophys.Acta.,1977,491:473
[103]魏晓芳 丁西明 刘会洲.pH诱导牛血清白蛋白芳香氨基酸残基微环境变化的光谱分析[J]光谱学及光谱分析,2000,20(4):556-558
[104]Peng.Z.G,Hidajat.K,Uddin.M.S..Adsorpton.of bovine serum albumin on nanosized magnetic particles.[J]Journal of Colloid and Interface Science,2004, 271:277-283.
[105]Terry E.Thomas,Cindy L.Miller,and Connie J.Eaves.Purification of Hematopoietic Stem Cells for Further Biological Study.A Companion to Methods in Enzymology 17,202-218(1999)
[106]唐佩弦.造血干细胞研究发展历史引发的思考.中国实验血液学杂志2005;13(5):723-732
[101]Halens S,Kohn DB.Gene therapy using hematopoietic stem cells:sisyphus approaches the cerst[J].Human Gone Therapy.2000.11(9):1259-1267
[107]徐勇,霍梅.免疫磁珠分离及流式细胞仪分选纯化外周血CD34+/CD90+干细胞.临床检验杂志2004年第22卷第4期
[108]Motoki Kuhara,Haruko Takeyama,Tsuyoshi Tanaka,et al.Magnetic Cell Separation.Using Antibody Binding with Protein A Expressed on Bacterial Magnetic Particles.Anal.chem.2004,76,6207-6213
[109]L.P.Zhu,X.Q.Chen.(editors) The experimental methods of immunology.People' s Military Medical Press,2003,P.70.
[110]Kimitaka Kato and Andreas Radbruch.Isolation and characterization of CD34+hematopietic stem cells from human peripheral blood by high-gradient magnetic cell sorting.Cytometry14:384-392(1993)
[111]熊玉宁,滕爱萍,王景文,沈悌.不同来源人CD34+细胞体外集落培养的研究及意义.中华内科杂志,2001年7月第40卷第7期