亚微米级免疫磁球及其在细菌分离中的应用

以亚微米级的单分散磁性微球为基础,制备出了表面包被有沙门氏菌抗体的免疫磁球. 利用表面电位、荧光和ELISA等方法研究了抗体在磁性微球表面的吸附行为. 在沙门氏菌磁分离实验中,通过调节投料抗体的浓度,研究了免疫磁球表面抗体浓度和磁分离效率的相关性,与微米级商品化免疫磁球的对比中,亚微米级的免疫磁球表现出了更高的磁分离效率.     

疏水性磁性微球的制备及对盐藻的吸附研究

采用疏水性材料制备微米级磁性微球,并应用于对盐藻的吸附分离研究,考察了pH值,吸附作用时间,NaCl浓度以及磁性微球添加量对盐藻吸附的影响.结果显示磁性微球对盐藻的吸附受溶液的离子强度影响较大;pH值在一定范围内对盐藻的吸附有较大影响;延长吸附作用时间和加大磁性微球的添加量可以增加对盐藻的吸附量.通过调节pH和施加机械搅拌,磁性微球上吸附的盐藻可以很好被洗脱下来,磁性微球可以多次重复使用.     

具有高选择性吸附的表面分子印迹磁性纤维素微球的制备与性能

以纤维素和纳米Fe₃O₄为原料制得磁性纤维素微球, 在纤维素微球表面选择合适的模板分子, 以甲基丙烯酸、 丙烯酰胺和N,N'-亚甲基双丙烯酰胺为功能单体, 采用水溶液聚合法制得表面分子印迹磁性纤维素微球. 采用傅里叶变换红外光谱(FTIR)、 X射线衍射(XRD)和振动样品磁强计(VSM)等表征了分子印迹聚合物微球的结构. 以罗丹明B(RhB)为模板分子, 通过吸附动力学与吸附热力学实验研究了表面分子印迹磁性纤维素微球对RhB的吸附性能, 结果表明, 制备的表面分子印迹磁性纤维素微球对罗丹明B具有特异性识别作用, 饱和吸附量达到0.542 mg/mg, 吸附平衡时间为10 h左右. 表面分子印迹磁性纤维素微球大大降低了对吸附环境的依赖, 并可重复利用.     

磁性壳聚糖微球的制备及对Cr(Ⅵ)的吸附性能研究

用壳聚糖包埋磁流体,用戊二醛交联制成磁性壳聚糖微球,并用红外光谱表征其结构。用制备的磁性壳聚糖微球吸附Cr(Ⅵ)离子,考察了其对Cr(Ⅵ)离子的吸附性能;探讨了吸附时间、溶液pH值、吸附剂用量、温度、Cr(Ⅵ)起始浓度以及其他离子存在对Cr(Ⅵ)离子去除率的影响。实验结果表明,磁性壳聚糖微球吸附Cr(Ⅵ)离子的最佳条件为:吸附平衡时间40 min,最佳吸附pH值6左右,磁性壳聚糖微球用量10 mg,温度升高有利于提高磁性壳聚糖微球的吸附效率,Cr(Ⅵ)离子起始质量浓度为12μg/mL,无机盐的存在引起磁性壳聚糖微球的吸附性能降低。并且考察了吸附剂的再生性能,实验结果表明磁性壳聚糖微球具有良好的重复使用性。     

磁性壳聚糖复合微球的制备及其Cu~(2+)吸附性能

以球磨后的粉煤灰磁珠(MS)颗粒为磁核,通过溶胶凝胶法和反相微乳液法依次包覆SiO_2和壳聚糖(CS),制备了MS@SiO_2@CS磁性微球。利用扫描电镜及能量色散谱仪、热重分析仪、红外光谱仪、X射线衍射仪、振动样品磁强计对所得样品的结构和磁性进行了系统表征。结果表明,磁珠颗粒表面实现了逐层包覆,较均匀的分散于壳聚糖基体中,MS@SiO_2@CS微球的比饱和磁化强度可达7.04 emu·g~(-1)。Cu~(2+)离子吸附实验表明,所得磁性壳聚糖微球对Cu~(2+)具有良好的吸附能力,最大吸附量可达11.08 mg·g~(-1);而且可通过磁选法高效固液分离。吸附动力学研究表明,MS@SiO_2@CS微球对Cu~(2+)离子的吸附符合准二级动力学模型,以化学吸附为主。     

改性果胶磁性微球的制备及吸附性能研究

为了提高果胶磁性微球的分散性及成球性,采用油酸对磁性微球进行表面改性后,制备改性果胶磁性微球。通过红外光谱、扫描电镜、XRD和磁性分析等对样品进行表征。油酸和果胶在纳米四氧化三铁表面形成了良好的修饰层,且油酸的加入提高了果胶磁性微球的分散性及成球性。研究了改性果胶磁性微球用量和溶液pH值等对其吸附Cu(Ⅱ)性能的影响,考察了其吸附动力学和吸附等温线。25℃时,改性果胶磁性微球对Cu(Ⅱ)的吸附达平衡需要2 h,饱合吸附容量为52.36 mg/g,Freundlich模型和Langmuir模型可以较好的拟合实验结果,最大吸附量为119.05 mg·g~(-1)。     

一种螯合树脂的合成及对金属离子的吸附研究

采用表面引发原子转移自由基聚合(SI-ATRP)技术,以自制的单分散磁性聚合物微球(Fe3O4/PGMA/EDMA)为ATRP大分子引发剂,Cu Br/2,2'-联吡啶(Bpy)为催化剂,将乙烯基四唑聚合在磁性聚合物微球表面,制备了磁性螯合吸附树脂,用元素分析对其进行了表征,并测定了该树脂对几种金属离子的吸附性能,结果发现新型螯合树脂对贵金属离子吸附能力强,具有良好的应用前景,工业推广价值较高。     

用于检测重金属离子的磁性荧光复合微球

采用静电逐层自组装的方法,首先将PSS和PAH聚电解质交替沉积在CaCO3中空微球表面,然后将Fe3O4磁性纳米粒子与CdSe量子点负载在中空微球表面不同的聚电解质层中,制备出具有磁性和荧光双重功能的复合微球,并将其作为荧光离子探针,研究了其对Cu2+和Pb2+离子检测的灵敏度、选择性及可行性。结果表明,复合微球显示出良好的磁性和荧光性能,对Cu2+和Pb2+离子的检测具有较高的灵敏度和选择性。尤为重要的是,可通过磁分离的方法将微球快速地从待测液中回收,从而能够避免量子点对环境造成的二次污染。     

磁性微球的制备及应用

磁性微球在生物医学和分离工程等多个领域都有广泛的应用,如固定化酶、靶向药物载体、核磁共振成像、细胞标记及分离、核酸和蛋白质分离纯化、环境治理等.近年来随着纳米新材料及制备工艺的不断创新,磁性微球的制备及应用都取得了大量进展.磁性微球的制备方法主要可以分为4类:(1)包裹法,在微球生成的同时将磁性纳米颗粒包裹在微球基质内,又可以细分为异质聚合法、乳化-溶剂挥发法、膜乳化和微流控技术、反相悬液-交联法、Stober法、聚合诱导胶体聚集法等;(2)模板组装法,在模板微球内部或表面组装磁性纳米颗粒,包括溶胀法、吸附法和静电自组装法等;(3)模板原位生成法,在模板微球的内部或表面原位生成磁性纳米颗粒,包括共沉淀、沉淀-氧化法、有机前驱体热分解法等;(4)溶剂(水)热法,以有机溶剂或水为介质,密闭体系中的反应物在一定的温度和溶液的自生压力下反应得到磁性微球.本文按照制备方法对近几年来磁性微球领域的研究进展进行了总结,包括反应原理、磁性微球的性质、表面功能化和应用介绍.     

内含Fe_3O_4和金纳米粒子的碳化微球的制备及其催化性能

通过蒸馏-沉淀聚合制备了丙烯酸-二乙烯苯共聚物微球,经离子交换吸附Fe2+离子,然后通过空气中加热、氩气气氛中高温碳化得到了含Fe3O4纳米粒子的多孔磁性碳化微球。在水介质中多孔磁性碳化微球吸附氯金酸,然后还原得到内含金纳米粒子的磁性碳化微球。以硼氢化钠还原对硝基苯酚生成对氨基苯酚反应为例,研究了内含金纳米粒子的磁性碳化微球的催化作用。结果表明,内含金纳米粒子的磁性碳化微球对该反应有很好的催化作用。通过外磁场很容易将磁性微球从反应液相中分离出来,微球重复使用10次后其催化活性基本未变。     

The effective and specific isolation of antibodies from human serum by using thiophilic paramagnetic polymer beads

A rapid and effective method to specifically isolate immunoglobulin G (IgG) from human serum by thiophilic paramagnetic polymer beads was developed. The thiophilic paramagnetic beads were synthesized with vinyl acetate and divinylbenzene by microsuspension polymerization in the presence of magnetite nanoparticles. Divinylsulfone and 2-mercapto-benzothiazole were subsequently used to modify the surface of these beads, resulting in thiophilic particles that exhibited a high specificity to the antibodies in serum at low salt concentration. The adsorbed IgG was eluted by 0.8 M KCl and 72% of the IgG in the serum was recovered. The purity of the isolated IgG reached 98.4% and the bioactivity was fully maintained (>99%). The high efficiency, mild conditions and simplicity make this technology suitable for the economic purification of antibodies in a large scale.     

Purification of the specific immunoglobulin G1 by immobilized metal ion affinity chromatography using nickel complexes of chelating porous and nonporous polymeric sorbents based on poly(methacrylic esters). Effect of polymer structure

Ni2+ complexes of the chelating nonporous and porous bead sorbents based on methacrylic esters crosslinked with ethylene dimethacrylate were used in isolation of the horseradish peroxidase-specific immunoglobulin IgG1 from the crude mouse ascitic fluid by immobilized metal ion affinity chromatography (IMAC). Iminodiacetic and aspartic acids were attached to porous poly(glycidyl methacrylate) beads differing in size, morphology and chemical composition. Ethylenediaminetriacetic acid and quinolin-8-ol chelating groups were attached mainly to the surface hydroxyl groups in nonporous poly(diethylene glycol methacrylate) beads through spacers. The latter sorbents exhibited better kinetic characteristics than the former but a very low IgG1 sorption capacity. In a single-step IMAC procedure, the best efficiency in the specific IgG1 purification was obtained with porous sorbents (recovery 92%, purity 73%). Differences in IMAC separations are discussed from the point of view of morphology of polymer beads as well as of the type and concentration of chelating ligands.     

Preparation of Medium Cation Exchange Stationary Phase of Polymeric Matrix and Their Chromatographic Properties

Based on the monodisperse poly（glycidyl methacrylate-co-ethylenedimethacrylate） beads （PGMA/EDMA） with macropore as a medium, a new hydrophilic medium cation exchange （MCX） stationary phase for HPLC was synthesized by a new chemically modified method. The stationary phase was evaluated with the property of ion exchange, separability, reproducibility, hydrophilicity, effect of salt concentration, salt types, column loading and pH on the separation and retention of proteins in detail. It was found that it follows ion exchange chromatographic （IEC） retention mechanism. The measured bioactivity recovery for lysozyme was （96 ± 5）%. The dynamic protein loading capacity of the synthesized MCX packings was 21.8 mg/g. Five proteins were almost completely separated within 6.0 min at a flow rate of 4 mL/min using the synthesized MCX resin. The MCX resin was also used for the rapid separation and purification of lysozyme from egg white with only one step. The purity and specific bioactivity of the purified lysozyme was found more than 95% and 70345 U/mg, respectively.     

Preparation of thiophilic paramagnetic adsorbent for separation of antibodies

The micron-sized microspheres with superparamagnetic property were synthesized with vinyl acetate and divinylbenzene by microsuspension polymerization. After the complete alcoholysis, these hydroxyl-functionalized microspheres were activated by divinylfone and modified with mercaptoethanol to prepare the thiophilic magnetic adsorbent, which was used to specifically isolate immunoglobulin G （IgG） from human serum. This thiophilic magnetic adsorbent performed an evident salt-dependent adsorption behavior for IgG. Due to their salt-promoted adsorption towards IgG under high salt concentration, the absorbed antibodies could be extracted in low salt concentration with high purity.     

Synthesis of Monodisperse Poly（glycidylmethacrylate-co-ethylene dimethacrylate） Beads and Their Application in Separation of Biopolymers

Introduction Since 19541 the polymeric separation media has attracted much attention due to their chemical stability over the entire pH range. The rigid, highly cross-linked styrene copolymers were first used for chromatography by Moore.2 The macroporous copolymers currently available are not only chemically stable but also more resistant to mechanical forces prevailing in a column and therefore are comparable to the traditional packings based on silica gel. Most polymer separation media are …     

On-column capture of a specific protein in capillary electrophoresis using magnetic beads

A method for capturing specific molecules separated by CE has been explored. To demonstrate on-column capture of migrating analyte molecules, two detection windows were fabricated on a capillary. Magnetic beads containing immobilized molecules that react with the specific molecules under study were placed between the detection windows in the capillary using magnets. Molecules in a sample solution injected into the capillary were separated and detected at the first detection window. After passing through the first detection window, the separated molecules encountered the magnetic beads, where the specific analyte was captured. As a result, the peak area for those analyte molecules decreased or disappeared completely at the second detection window. Rabbit IgG and carbonic anhydrase were employed to demonstrate on-column capture of a specific molecule. For rabbit IgG, magnetic beads containing the immobilized antibody (anti-rabbit IgG) were used. Rabbit IgG molecules were captured on the magnetic beads during CE migration. Furthermore, the capture of carbonic anhydrase was demonstrated by the reaction between magnetic beads (containing immobilized anti-rabbit IgG) and anti-carbonic anhydrase (rabbit IgG), before the beads were packed in the capillary. After packing the magnetic beads in the capillary, a mixture of two proteins was injected into the capillary. Two proteins were detected at the first detection window, while the peak corresponding to carbonic anhydrase disappeared at the second detection window. The results show that using an appropriate antibody, the present technique would be applicable to any proteins.     

高效阳离子交换法分离纯化蛋清中的溶菌酶

 建立了用高效阳离子交换分离纯化蛋清中溶菌酶的新方法 ,讨论了纯化的工艺条件。蛋清样品匀浆后 ,用氯化钠初步纯化 ,然后用弱阳离子交换柱XIDACE WCX分离。结果表明 ,被纯化的溶菌酶和杂蛋白得到很好分离。经活性检测 ,溶菌酶过柱后的活性回收率为 10 7% ,蛋白的比活为 15 4 6 7U/mg ,纯化倍数提高了 5 6倍。用尺寸排阻 (SEC)鉴定 ,得到的溶菌酶呈均一性。该法较传统软基质低压离子交换分离速度快 ,纯化效率高。     

The combined use of high performance liquid chromatography and immuno-biochemical techniques for protein isolation: a new approach for identification of an individual protein from a pool of proteins

HPLC was used in combination with immuno-bead separation technique for identification of an individual protein from a pool of proteins. This was carried out using an in-house monoclonal antibody (ATC2) specific for placental alkaline phosphatase (PLAP) as a primary antibody for conjugation to CNBr beads. The phosphatase activity (ALP) of PLAP was measured by colorimetric assay (MEDC). The data from this study has so far indicated that: 1. HPLC analysis of molecules following isolation with ATC2-conjugated beads showed high degree of purity. This could be achieved using protein mixtures prepared from lysates of tumour cell lines or tumour fragments. 2. HPLC-isolated PLAP maintained phosphatase activity. 3. Out of the four dissociation reagents used, diethyl amine (DEA) was found to be the best reagent for dissociation of antigen, ie PLAP, but not mAb from CNBr beads. 4. The profile of ALP activity was different for samples prepared from testis and kidney fragments, both in terms of the HPLC peak profile as well as the sensitivity. These data confirmed that the immuno-bead separation technique in conjunction with HPLC were powerful tools for identifying an individual protein from a pool of proteins. These approaches are being used for the identification of PLAP molecules, as a tumour marker in patients suspected of testicular malignancies with equivocal ultrasound.     

Hydrophobic charge‐induction resin with 5‐aminobenzimidazol as the functional ligand: preparation,protein adsorption and immunoglobulin G purification

A new hydrophobic charge‐induction chromatography resin was prepared with 5‐aminobenzimidazol as functional ligand and polyacrylic ester beads as matrix. Adsorption isotherms and adsorption in columns were investigated using human immunoglobulin G and bovine serum albumin as model proteins, and the influence of pH and NaCl concentration was discussed. Results showed that the ligand density was 195 μmol/mL gel, and protein selectivity can be improved by controlling pH and salt addition. An optimized purification process (sample loading at pH 8.0 with 0.2 M NaCl and elution at pH 5.0) was performed to purify human immunoglobulin G from bovine serum albumin containing feedstock, which resulted in human immunoglobulin G purity of 99.7% and recovery of 94.6%. A similar process was applied for the purification of monoclonal antibody from cell culture supernatant, which showed antibody purity of 94.9% and recovery of 92.5%. The results indicated that the new resin developed had comparable performance as Protein A chromatography and would be suitable for antibody purification from complex feedstock.