In this study, a computer code is developed to numerically investigate a magnetic bead micromixer under different conditions. The micromixer consists of a microchannel and numerous micro magnetic particles which enter...In this study, a computer code is developed to numerically investigate a magnetic bead micromixer under different conditions. The micromixer consists of a microchannel and numerous micro magnetic particles which enter the micromixer by fluid flows and are actuated by an alternating magnetic field normal to the main flow. An important feature of micromixer which is not considered before by researchers is the particle entrance arrangement into the micromixer. This parameter could effectively affect the micromixer efficiency. There are two general micro magnetic particle entrance arrangements in magnetic bead micromixers: determined position entrance and random position entrance. In the case of determined position entrances, micro magnetic particles enter the micromixer at specific positions of entrance cross section. However, in a random position entrance,particles enter the microchannel with no order. In this study mixing efficiencies of identical magnetic bead micromixers which only differ in particle entrance arrangement are numerically investigated and compared.The results reported in this paper illustrate that the prepared computer code can be one of the most powerful and beneficial tools for the magnetic bead micromixer performance analysis. In addition, the results show that some features of the magnetic bead micromixer are strongly affected by the entrance arrangement of the particles.展开更多
The organic monomer- molecule with nanometer magnetic powder by means of reforming the surface of nanometer magnetic powder have been synthesized. Magnetic beads in diameter of 2μm or so are obtained by controlling c...The organic monomer- molecule with nanometer magnetic powder by means of reforming the surface of nanometer magnetic powder have been synthesized. Magnetic beads in diameter of 2μm or so are obtained by controlling conditions . Ovary cancer cells of ascites are separated and ovary cancer cells of blood are detected by using immuno-magnetic beads linked with ovary cancer cell mono-antibodies. Results show that the specificity is 85% , sensitivity is 87% , accuracy is 84% , cells acquiring purity is 90% , cells activity is 92% and detection sensitivity is 25 × 10-7 .展开更多
The transcription activator-like effector nuclease(TALEN) technique combined with the somatic cell nuclear transfer(SCNT) method has been successfully applied for creating genetically modified pigs. However, methods f...The transcription activator-like effector nuclease(TALEN) technique combined with the somatic cell nuclear transfer(SCNT) method has been successfully applied for creating genetically modified pigs. However, methods for isolating cells with biallelic indels requires further improvement because of the relatively low enrichment efficiency of mutated somatic cells. Moreover, little is known regarding the off-target effects of the TALEN system and the heredity of TALEN-modified pigs. In this study, an efficient method to increase the enrichment efficiency of TALEN-mediated biallelic knockout(KO) cells was established, and corresponding genetically modified pigs with the expected genotype were generated whose off-target effect, fertility and heredity characteristics were aslo evaluated. Two TALEN pairs were constructed to target the porcine α-1,3-galactosyltransferase(GGTA1) gene locus. TALEN m RNA was transfected into the ear fibroblasts followed by the enrichment of α-Gal null cells of minipigs using isolectin B4(IB4) lectin and m agnetic beads. A total of 115 cell c olonies were formed and validated to be GGTA1 KO cells by sequencing and 10 biallelic KO cell colonies were used as nuclear donors for SCNT. Thirty GGTA1 biallelic KO piglets were successfully delivered and grew normally. Seventeen potential off-target sites were investigated, and no off-target events were detected in the live piglets. To determine t he fertility and heredity characteristics of TALEN-modified pigs, 10 mature founders w ere mated with each other and the mutations were determined to be transmitted to the F1 piglets. We established a robust and safe technology for developing genetically modified pig lines with expected genotypes for agricultural breeding and biomedical application.展开更多
A protocol for enrichment and adsorption of karyocyte from whole blood by using magnetic nanometer beads as solid phase absorbents was presented. The PCR amplification could be accomplished by using the nanobeads with...A protocol for enrichment and adsorption of karyocyte from whole blood by using magnetic nanometer beads as solid phase absorbents was presented. The PCR amplification could be accomplished by using the nanobeads with karyocyte as template directly and the PCR products were applied on an oligonucleotide array to do gene typing. The HLA A PCR amplification system and a small HLA A oligonucleotide microarray were applied as the platform and an experiment protocol of separating karyocyte from whole blood using the magnetic nanometer beads (Fe 2O 3) were set up. The experimental conditions were also discussed. It showed that pH level of PBS eluent, Taq enzyme quantity and fragment length of products could influent the amplification results, and the magnetic nano beads could succeed in sample preparation in microarray to provide a promising way in automatic detection and lab on a chip.展开更多
Objective To evaluate the application of weak cation exchange (WCX) magnetic bead-based Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) in detecting differentially expressed...Objective To evaluate the application of weak cation exchange (WCX) magnetic bead-based Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) in detecting differentially expressed proteins in the urine of renal clear cell carcinoma (RCCC) and its value in the early diagnosis of RCCC.Methods Eleven newly diagnosed patients (10 males and 1 female, aged 46-78, mean 63 years) of renal clear cell carcinoma by biopsy and 10 healthy volunteers (all males, aged 25-32, mean 29.7 years) were enrolled in this study. Urine samples of the RCCC patients and healthy controls were collected in the morning.Weak cation exchange (WCX) bead-based MALDI-TOF MS technique was applied in detecting differential protein peaks in the urine of RCCC. ClinProTools2.2 software was utilized to determine the characteristic proteins in the urine of RCCC patients for the predictive model of RCCC.Results The technique identified 160 protein peaks in the urine that were different between RCCC patients and health controls; and among them, there was one peak (molecular weight of 2221.71 Da) with statistical significance (P=0.0304). With genetic algorithms and the support vector machine, we screened out 13 characteristic protein peaks for the predictive model.Conclusions The application of WCX magnetic bead-based MALDI-TOF MS in detecting differentiallyexpressed proteins in urine may have potential value for the early diagnosis of RCCC.展开更多
Heavy metal ion is one of the major environmental pollutants.In this study,a Cu(Ⅱ)ions imprinted magnetic chitosan beads are prepared to use chitosan as functional monomer,Cu(Ⅱ)ions as template,Fe_(3)O_(4) as magnet...Heavy metal ion is one of the major environmental pollutants.In this study,a Cu(Ⅱ)ions imprinted magnetic chitosan beads are prepared to use chitosan as functional monomer,Cu(Ⅱ)ions as template,Fe_(3)O_(4) as magnetic core and epichlorohydrin and glutaraldehyde as crosslinker,which can be used for removal Cu(Ⅱ)ions from wastewater.The kinetic study shows that the adsorption process follows the pseudosecond-order kinetic equations.The adsorption isotherm study shows that the Langmuir isotherm equation best fits for the monolayer adsorption processes.The selective adsorption properties are performed in Cu(Ⅱ)/Zn(Ⅱ),Cu(Ⅱ)/Ni(Ⅱ),and Cu(Ⅱ)/Co(Ⅱ)binary systems.The results shows that the ⅡMCD has a high selectivity for Cu(Ⅱ)ions in binary systems.The mechanism of ⅡMCD recognition Cu(Ⅱ)ions is also discussed.The results show that the ⅡMCD adsorption Cu(Ⅱ)ions is an enthalpy controlled process.The absolute value of DH(Cu(Ⅱ))and DS(Cu(Ⅱ))is greater than DH(Zn(Ⅱ),Ni(Ⅱ),Co(Ⅱ))and DS(Zn(Ⅱ),Ni(Ⅱ),Co(Ⅱ)),respectively,this indicates that the Cu(Ⅱ)ions have a good spatial matching with imprinted holes on ⅡMCD.The FTIR and XPS also demonstrates the strongly combination of function groups on imprinted holes in the suitable space position.Finally,the ⅡMCD can be regenerated and reused for 10 times without a significantly decreasing in adsorption capacity.This information can be used for further application in the selective removal of Cu(Ⅱ)ions from industrial wastewater.展开更多
Due to the complexity of bioactive ingredients in biological samples,the screening of target proteins is a complex process.Herein,a feasible strategy for directing protein immobilization on silica magnetic beads for l...Due to the complexity of bioactive ingredients in biological samples,the screening of target proteins is a complex process.Herein,a feasible strategy for directing protein immobilization on silica magnetic beads for ligand fishing based on SpyTag/SpyCatcher(ST/SC)-mediated anchoring is presented.Carboxyl functional groups on the surface of silica-coated magnetic beads(SMBs)were coupled with SC using the 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride/N-hydroxysulfosuccinimide method,named SC-SMBs.The green fluorescent protein(GFP),as the capturing protein model,was ST-labeled and anchored at a specific orientation onto the surface of SC-SMBs directly from relevant cell lysates via ST/SC self-ligation.The characteristics of the SC-SMBs were studied via electron microscopy,energy dispersive spectroscopy,and Fourier transform infrared spectroscopy.The spontaneity and site-specificity of this unique reaction were confirmed via electrophoresis and fluorescence analyses.Although the alkaline stability of ST-GFP-ligated SC-SMBs was not ideal,the formed isopeptide bond was unbreakable under acidic conditions(0.05 M glycine-HCl buffer,pH 1e6)for 2 h,under 20%ethanol solution within 7 days,and at most temperatures.We,therefore,present a simple and universal strategy for the preparation of diverse protein-functionalized SMBs for ligand fishing,prompting its usage on drug screening and target finding.展开更多
Overthelastseveraldecades,harmfulalgalblooms(HABs)havebecomeaseriousenvironmental problem in many parts of the world. A rapid and accurate detection process for HAB algae has yet to be developed. Heterosigma akashiwo ...Overthelastseveraldecades,harmfulalgalblooms(HABs)havebecomeaseriousenvironmental problem in many parts of the world. A rapid and accurate detection process for HAB algae has yet to be developed. Heterosigma akashiwo is one of the most important HABs species in China. The objective of this study was to develop an immunologic technique that can rapidly and sensitively count H. akashiwo cells. Five HABs species (Alexandrium catenella, Thalassiosira sp., Cryptomonas sp., Alexandrium tamarense and Symbiodinium sp.,) were used in this study to evaluate the analysis process we developed. A polyclonal antibody with high titers against H. akashiwo was obtained by injecting H. akashiwo cells into rabbits. Immuno-magnetic beads (IMB) were produced via conjugated polyclonal antibodies with magnetic beads and applied to isolate and count H. akashiwo cells from the culture. Results show that 66.7%-91.6% of the cells were captured from unialgal culture by IMBs, and only 5.3%-12.5% of the four other HAB microalgae species were captured, indicating that the constructed IMBs combined specifically with the H. akashiwo cells. At the same time, flow cytometry (FCM) sorting was exploited to screen H. akashiwo cells after labeling with FITC conjugated polyclonal antibodies. Using the FCM technique, 91.7% of the targeted cells were sorted out from mixed microalgae samples in just a few minutes. These results indicate that both antibody-involved IMB and antibody-based FCM techniques are highly effective at detecting and quantifying HAB species. These techniques, especially immuno-magnetic separation, have low associated cost, and are fast and simple processes compared with other techniques currently in use.展开更多
AIM: To find new biomarkers for uveal melanoma(UM) by analyzing the serum peptidome profile. METHODS: Proteomic spectra in patients with UM before and after operation were analyzed and compared with those of healthy c...AIM: To find new biomarkers for uveal melanoma(UM) by analyzing the serum peptidome profile. METHODS: Proteomic spectra in patients with UM before and after operation were analyzed and compared with those of healthy controls. Magnetic affinity beads were used to capture serum peptides and matrix-assisted laser desorption/ionization time-of-flight(MALDI-TOF) mass spectrometer were used to compile serum peptide profiles. RESULTS: A panel of 49 peptides were differentially expressed between UM patients and controls, of which 33 peptides were of higher intensities in patient group and 16 peptides were of higher intensities in control group. Based on combined use of these potential markers, peptides with mean molecular masses of 1467 and 9289.0 Da provide high sensitivity(83.3%), specificity(100%) and accuracy rate(93.0%) together to differentiate melanoma patients from healthy controls. At the time point of 6mo postoperatively, the levels of many peptides differentially expressed before surgery showed no more statistical difference between the patients and the control group. Fibrinogen α-chain precursors were identified as potential UM markers.CONCLUSION: We have shown that a convenient and fast proteomic technique, affinity bead separation and MALDITOF analysis combined with bioinformatic software, facilitates the identification of novel biomarkers for UM.展开更多
Peptides in shrimp hemolymph play an important role in the innate immune response.Analysis of hemolymph will help to detect and identify potential novel biomarkers of microbial infection.We used magnetic bead-based pu...Peptides in shrimp hemolymph play an important role in the innate immune response.Analysis of hemolymph will help to detect and identify potential novel biomarkers of microbial infection.We used magnetic bead-based purification(ClinProt system) and matrix-assisted laser desorption/ionization time of flight mass spectrometry(MALDI-TOF MS) to characterize shrimp hemolymph peptides.Shrimp serum and plasma were used as the source of samples for comparative analysis,and it was found that serum was more suitable for shrimp hemolymph peptidomic analysis.To screen potential specific biomarkers in serum of immune-challenged shrimps,we applied magnetic bead-based MALDI-TOF MS to serum samples from 10 immune-challenged and 10 healthy shrimps.The spectra were analyzed using FlexAnalysis 3.0 and ClinProTools 2.1 software.Thirteen peptide peaks significantly different between the two groups were selected as candidate biomarkers of lipopolysaccharide(LPS)-infection.The diagnostic model established by genetic algorithm using five of these peaks was able to discriminate LPS-challenged shrimps from healthy control shrimps with a sensitivity of 90% and a specificity of 100%.Our approach in MALDITOF MS-based peptidomics is a powerful tool for screening bioactive peptides or biomarkers derived from hemolymph,and will help to enable a better understanding of the innate immune response of shrimps.展开更多
A simple and non-invasive method for detecting endometrial cancer in women with abnormal uterine bleeding is required. For this purpose, we prepared immuno-magnetic beads conjugated with anti-human EpCAM rat monoclona...A simple and non-invasive method for detecting endometrial cancer in women with abnormal uterine bleeding is required. For this purpose, we prepared immuno-magnetic beads conjugated with anti-human EpCAM rat monoclonal antibody (mAb) for isolating exfoliated endometrial cells including endometrial cancer cells in vaginal discharge. The affinities of the anti-human EpCAM rat mAbs were analyzed by flow cytometry and immunocytochemistry and then magnetic beads were conjugated with the mAbs. The rate of retrieval of endometrial cells using the immuno-magnetic beads was calculated. Endometrial cells were isolated using the immuno-magnetic beads from the vaginal discharges of 22 patients with endometrial cancer and 16 non-malignant controls. The isolated cells were stained using endometrial cancer specific-mAbs and analyzed by flow cytometry and imaging cytometry. The immuno-magnetic beads conjugated with high-affinity mAb (clone 1456) appeared to have very low auto-fluorescence. Sufficient enrichment of Ep-CAMpositive cells using immuno-magnetic beads was observed in both simulation and clinical samples. The overall sensitivities of flow cytometry and imaging cytometry to detect endometrial cancer cells were 72.7% and 45.5%, respectively. Meanwhile, the overall specificities of flow cytometry and imaging cytometry for healthy controls were 75.0% and 81.3%, respectively. Our immuno-magnetic beads have very low auto-fluorescence, so they could be useful for fluorescent analysis, such as fluorescent immunochemical staining. In the future, these novel immuno-magnetic beads could be used for cytological study.展开更多
To investigate the technique of sorting high-purity precartilaginous stem cells from rat’s perichondrium, neonatal rat’s perichondrium cells suspensions were incubated with monoclone anti-body of anti-fibroblast gro...To investigate the technique of sorting high-purity precartilaginous stem cells from rat’s perichondrium, neonatal rat’s perichondrium cells suspensions were incubated with monoclone anti-body of anti-fibroblast growth factor receptor-3 (anti-FGFR-3), and the labeled cells were separated from the suspension in the magnetic field by immuno-beads coated with the second antibody. Purity of the sorted neural stem cells was found to be 93.0 %-99.0 %, with living cells amounting to 80 %-85 %. The magnetic cell sorting system could effectively separate precartilaginous stem cells from perichondrium cell suspensions.展开更多
On the basis of study on physical and chemical properties of magnetic bead (MB) in fly ash (FA), the paper gives out the separation methods of MB and results of three separating process. The result of comparative test...On the basis of study on physical and chemical properties of magnetic bead (MB) in fly ash (FA), the paper gives out the separation methods of MB and results of three separating process. The result of comparative test in size, density, stability, magnetic material content, specific magnetic susceptibility (SMS), medium recovery oxidation resistance and wear resistance between MB and magnetic fines currently used in dense medium separation leads to that using MB recovered from fly ash is used as medium solids in coal cleaning in stead of magnetic fines not only have no influence upon taryests of separation, but can bring good economic and social benefits.展开更多
目的评估自动化磁珠法提取血清脂溶性维生素应用液相色谱串联质谱法(liquid chromatography-tandem mass spectrometry,LC-MS/MS)检测的性能。方法收集200例临床剩余血清样本,采用自动化磁珠法提取血清中脂溶性维生素A,D_(2),D_(3),E和K...目的评估自动化磁珠法提取血清脂溶性维生素应用液相色谱串联质谱法(liquid chromatography-tandem mass spectrometry,LC-MS/MS)检测的性能。方法收集200例临床剩余血清样本,采用自动化磁珠法提取血清中脂溶性维生素A,D_(2),D_(3),E和K;同时联合LC-MS/MS检测脂溶性维生素A,D_(2),D_(3),E和K的线性、定量限、精密度、正确度、携带污染率等性能指标以及基质效应。并比较此方法与传统萃取法检测结果的一致性。结果自动化磁珠法提取脂溶性维生素A,D_(2),D_(3),E和K线性相关系数均>0.99;五种物质的定量限分别为5,0.25,0.25,125和0.025ng/ml;批内精密度和批间精密度分别为0.66%~4.83%,0.15%~3.70%;平均加标回收率为87.05%~111.11%;基质效应为95.43%~99.07%;高-低值样本循环进样结果均值与低-低值样本循环进样结果均值之差,均小于低-低值样本循环进样结果均值的3s;统计学结果显示自动化磁珠法和传统萃取法提取的脂溶性维生素结果相关性良好(r>0.99),两种方法的检测结果无显著偏倚。结论自动化磁珠法提取脂溶性维生素的检测性能良好,有望提高样品通量和分析效率。展开更多
构建一种基于杂交链式反应(hybridization chain reaction,HCR)扩增的适配体磁珠荧光传感器。巧妙设计序列HP和发卡序列H1、H2,其中HP是由适配体序列与触发序列结合而成的,并且序列互补形成稳定的二级结构。然后采用戊二醇反应和亲和素...构建一种基于杂交链式反应(hybridization chain reaction,HCR)扩增的适配体磁珠荧光传感器。巧妙设计序列HP和发卡序列H1、H2,其中HP是由适配体序列与触发序列结合而成的,并且序列互补形成稳定的二级结构。然后采用戊二醇反应和亲和素-生物素反应进行适配体功能化磁珠的制备。将阪崎肠杆菌与适配体磁珠一起孵育,HP中的适配体序列识别靶标,引起HP构象变化,露出触发序列,通过HCR触发H1和H2的链状组装,产生长双链DNA。荧光指示剂SYBR Green I以插层和小槽结合的方式与HCR产物的长双链结合。最后加入氧化石墨烯(graphene oxide,GO)后,游离的H1、H2和SYBR Green I将通过π-π堆积紧密吸附在GO表面,荧光信号被猝灭。HCR产物不能被吸附在GO表面,因此与HCR产物结合的SYBR Green I发出依赖于靶浓度的强荧光信号,从而实现阪崎肠杆菌的定量检测。本方法在纯培养条件下的检出限为2CFU/mL,对奶粉的检出限为8CFU/g,对奶粉样品的检测结果与传统微生物培养法具有良好的一致性。该方法具有无需DNA提取,快速、稳定性高、高特异性和高灵敏度等优点,因此为阪崎肠杆菌的现场快速检测提供了一种很有潜力的方法。展开更多
基金Supported by Shanghai Science and Technology Development Program, No. 03DZ14024the National High Technology 863 Programs, No. 2002BAC11A11National Development Program (973) for Key Basic Research of China, No. 2001CB510205
文摘In this study, a computer code is developed to numerically investigate a magnetic bead micromixer under different conditions. The micromixer consists of a microchannel and numerous micro magnetic particles which enter the micromixer by fluid flows and are actuated by an alternating magnetic field normal to the main flow. An important feature of micromixer which is not considered before by researchers is the particle entrance arrangement into the micromixer. This parameter could effectively affect the micromixer efficiency. There are two general micro magnetic particle entrance arrangements in magnetic bead micromixers: determined position entrance and random position entrance. In the case of determined position entrances, micro magnetic particles enter the micromixer at specific positions of entrance cross section. However, in a random position entrance,particles enter the microchannel with no order. In this study mixing efficiencies of identical magnetic bead micromixers which only differ in particle entrance arrangement are numerically investigated and compared.The results reported in this paper illustrate that the prepared computer code can be one of the most powerful and beneficial tools for the magnetic bead micromixer performance analysis. In addition, the results show that some features of the magnetic bead micromixer are strongly affected by the entrance arrangement of the particles.
文摘The organic monomer- molecule with nanometer magnetic powder by means of reforming the surface of nanometer magnetic powder have been synthesized. Magnetic beads in diameter of 2μm or so are obtained by controlling conditions . Ovary cancer cells of ascites are separated and ovary cancer cells of blood are detected by using immuno-magnetic beads linked with ovary cancer cell mono-antibodies. Results show that the specificity is 85% , sensitivity is 87% , accuracy is 84% , cells acquiring purity is 90% , cells activity is 92% and detection sensitivity is 25 × 10-7 .
基金The authors are grateful to the financial support for this research from the National Natural Science Foundation of China，the Research Fund for the Doctoral Program of Higher Education of China
基金supported by the National Basic Research Program of China(973 Program)(2015CB554103 and 2011CBA01004)
文摘The transcription activator-like effector nuclease(TALEN) technique combined with the somatic cell nuclear transfer(SCNT) method has been successfully applied for creating genetically modified pigs. However, methods for isolating cells with biallelic indels requires further improvement because of the relatively low enrichment efficiency of mutated somatic cells. Moreover, little is known regarding the off-target effects of the TALEN system and the heredity of TALEN-modified pigs. In this study, an efficient method to increase the enrichment efficiency of TALEN-mediated biallelic knockout(KO) cells was established, and corresponding genetically modified pigs with the expected genotype were generated whose off-target effect, fertility and heredity characteristics were aslo evaluated. Two TALEN pairs were constructed to target the porcine α-1,3-galactosyltransferase(GGTA1) gene locus. TALEN m RNA was transfected into the ear fibroblasts followed by the enrichment of α-Gal null cells of minipigs using isolectin B4(IB4) lectin and m agnetic beads. A total of 115 cell c olonies were formed and validated to be GGTA1 KO cells by sequencing and 10 biallelic KO cell colonies were used as nuclear donors for SCNT. Thirty GGTA1 biallelic KO piglets were successfully delivered and grew normally. Seventeen potential off-target sites were investigated, and no off-target events were detected in the live piglets. To determine t he fertility and heredity characteristics of TALEN-modified pigs, 10 mature founders w ere mated with each other and the mutations were determined to be transmitted to the F1 piglets. We established a robust and safe technology for developing genetically modified pig lines with expected genotypes for agricultural breeding and biomedical application.
文摘A protocol for enrichment and adsorption of karyocyte from whole blood by using magnetic nanometer beads as solid phase absorbents was presented. The PCR amplification could be accomplished by using the nanobeads with karyocyte as template directly and the PCR products were applied on an oligonucleotide array to do gene typing. The HLA A PCR amplification system and a small HLA A oligonucleotide microarray were applied as the platform and an experiment protocol of separating karyocyte from whole blood using the magnetic nanometer beads (Fe 2O 3) were set up. The experimental conditions were also discussed. It showed that pH level of PBS eluent, Taq enzyme quantity and fragment length of products could influent the amplification results, and the magnetic nano beads could succeed in sample preparation in microarray to provide a promising way in automatic detection and lab on a chip.
文摘Objective To evaluate the application of weak cation exchange (WCX) magnetic bead-based Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) in detecting differentially expressed proteins in the urine of renal clear cell carcinoma (RCCC) and its value in the early diagnosis of RCCC.Methods Eleven newly diagnosed patients (10 males and 1 female, aged 46-78, mean 63 years) of renal clear cell carcinoma by biopsy and 10 healthy volunteers (all males, aged 25-32, mean 29.7 years) were enrolled in this study. Urine samples of the RCCC patients and healthy controls were collected in the morning.Weak cation exchange (WCX) bead-based MALDI-TOF MS technique was applied in detecting differential protein peaks in the urine of RCCC. ClinProTools2.2 software was utilized to determine the characteristic proteins in the urine of RCCC patients for the predictive model of RCCC.Results The technique identified 160 protein peaks in the urine that were different between RCCC patients and health controls; and among them, there was one peak (molecular weight of 2221.71 Da) with statistical significance (P=0.0304). With genetic algorithms and the support vector machine, we screened out 13 characteristic protein peaks for the predictive model.Conclusions The application of WCX magnetic bead-based MALDI-TOF MS in detecting differentiallyexpressed proteins in urine may have potential value for the early diagnosis of RCCC.
文摘Heavy metal ion is one of the major environmental pollutants.In this study,a Cu(Ⅱ)ions imprinted magnetic chitosan beads are prepared to use chitosan as functional monomer,Cu(Ⅱ)ions as template,Fe_(3)O_(4) as magnetic core and epichlorohydrin and glutaraldehyde as crosslinker,which can be used for removal Cu(Ⅱ)ions from wastewater.The kinetic study shows that the adsorption process follows the pseudosecond-order kinetic equations.The adsorption isotherm study shows that the Langmuir isotherm equation best fits for the monolayer adsorption processes.The selective adsorption properties are performed in Cu(Ⅱ)/Zn(Ⅱ),Cu(Ⅱ)/Ni(Ⅱ),and Cu(Ⅱ)/Co(Ⅱ)binary systems.The results shows that the ⅡMCD has a high selectivity for Cu(Ⅱ)ions in binary systems.The mechanism of ⅡMCD recognition Cu(Ⅱ)ions is also discussed.The results show that the ⅡMCD adsorption Cu(Ⅱ)ions is an enthalpy controlled process.The absolute value of DH(Cu(Ⅱ))and DS(Cu(Ⅱ))is greater than DH(Zn(Ⅱ),Ni(Ⅱ),Co(Ⅱ))and DS(Zn(Ⅱ),Ni(Ⅱ),Co(Ⅱ)),respectively,this indicates that the Cu(Ⅱ)ions have a good spatial matching with imprinted holes on ⅡMCD.The FTIR and XPS also demonstrates the strongly combination of function groups on imprinted holes in the suitable space position.Finally,the ⅡMCD can be regenerated and reused for 10 times without a significantly decreasing in adsorption capacity.This information can be used for further application in the selective removal of Cu(Ⅱ)ions from industrial wastewater.
基金supported by the Zhejiang Foundation Public Welfare Research Project(Authorization No.:LGF19B060006)。
文摘Due to the complexity of bioactive ingredients in biological samples,the screening of target proteins is a complex process.Herein,a feasible strategy for directing protein immobilization on silica magnetic beads for ligand fishing based on SpyTag/SpyCatcher(ST/SC)-mediated anchoring is presented.Carboxyl functional groups on the surface of silica-coated magnetic beads(SMBs)were coupled with SC using the 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride/N-hydroxysulfosuccinimide method,named SC-SMBs.The green fluorescent protein(GFP),as the capturing protein model,was ST-labeled and anchored at a specific orientation onto the surface of SC-SMBs directly from relevant cell lysates via ST/SC self-ligation.The characteristics of the SC-SMBs were studied via electron microscopy,energy dispersive spectroscopy,and Fourier transform infrared spectroscopy.The spontaneity and site-specificity of this unique reaction were confirmed via electrophoresis and fluorescence analyses.Although the alkaline stability of ST-GFP-ligated SC-SMBs was not ideal,the formed isopeptide bond was unbreakable under acidic conditions(0.05 M glycine-HCl buffer,pH 1e6)for 2 h,under 20%ethanol solution within 7 days,and at most temperatures.We,therefore,present a simple and universal strategy for the preparation of diverse protein-functionalized SMBs for ligand fishing,prompting its usage on drug screening and target finding.
基金Supported by Specialized Research Fund for the Doctoral Program of Higher Education of China (No. SRFDP 20100132110007)Shandong Provincial Natural Science Foundation, China (No. ZR2011DZ002)
文摘Overthelastseveraldecades,harmfulalgalblooms(HABs)havebecomeaseriousenvironmental problem in many parts of the world. A rapid and accurate detection process for HAB algae has yet to be developed. Heterosigma akashiwo is one of the most important HABs species in China. The objective of this study was to develop an immunologic technique that can rapidly and sensitively count H. akashiwo cells. Five HABs species (Alexandrium catenella, Thalassiosira sp., Cryptomonas sp., Alexandrium tamarense and Symbiodinium sp.,) were used in this study to evaluate the analysis process we developed. A polyclonal antibody with high titers against H. akashiwo was obtained by injecting H. akashiwo cells into rabbits. Immuno-magnetic beads (IMB) were produced via conjugated polyclonal antibodies with magnetic beads and applied to isolate and count H. akashiwo cells from the culture. Results show that 66.7%-91.6% of the cells were captured from unialgal culture by IMBs, and only 5.3%-12.5% of the four other HAB microalgae species were captured, indicating that the constructed IMBs combined specifically with the H. akashiwo cells. At the same time, flow cytometry (FCM) sorting was exploited to screen H. akashiwo cells after labeling with FITC conjugated polyclonal antibodies. Using the FCM technique, 91.7% of the targeted cells were sorted out from mixed microalgae samples in just a few minutes. These results indicate that both antibody-involved IMB and antibody-based FCM techniques are highly effective at detecting and quantifying HAB species. These techniques, especially immuno-magnetic separation, have low associated cost, and are fast and simple processes compared with other techniques currently in use.
基金Supported by the National Natural Science Foundation of China(No.81570891No.81272981)+2 种基金the Beijing Natural Science Foundation(No.7151003)Advanced Health Care Professionals Development Project of Beijing Municipal Health Bureau(No.2014-2-003)Beijing Municipal Administration of Hospitals Clinical Medicine Development of Special Funding Support(No.ZYLX201307)
文摘AIM: To find new biomarkers for uveal melanoma(UM) by analyzing the serum peptidome profile. METHODS: Proteomic spectra in patients with UM before and after operation were analyzed and compared with those of healthy controls. Magnetic affinity beads were used to capture serum peptides and matrix-assisted laser desorption/ionization time-of-flight(MALDI-TOF) mass spectrometer were used to compile serum peptide profiles. RESULTS: A panel of 49 peptides were differentially expressed between UM patients and controls, of which 33 peptides were of higher intensities in patient group and 16 peptides were of higher intensities in control group. Based on combined use of these potential markers, peptides with mean molecular masses of 1467 and 9289.0 Da provide high sensitivity(83.3%), specificity(100%) and accuracy rate(93.0%) together to differentiate melanoma patients from healthy controls. At the time point of 6mo postoperatively, the levels of many peptides differentially expressed before surgery showed no more statistical difference between the patients and the control group. Fibrinogen α-chain precursors were identified as potential UM markers.CONCLUSION: We have shown that a convenient and fast proteomic technique, affinity bead separation and MALDITOF analysis combined with bioinformatic software, facilitates the identification of novel biomarkers for UM.
基金Supported by the National Natural Science Foundation of China(No.30600458)
文摘Peptides in shrimp hemolymph play an important role in the innate immune response.Analysis of hemolymph will help to detect and identify potential novel biomarkers of microbial infection.We used magnetic bead-based purification(ClinProt system) and matrix-assisted laser desorption/ionization time of flight mass spectrometry(MALDI-TOF MS) to characterize shrimp hemolymph peptides.Shrimp serum and plasma were used as the source of samples for comparative analysis,and it was found that serum was more suitable for shrimp hemolymph peptidomic analysis.To screen potential specific biomarkers in serum of immune-challenged shrimps,we applied magnetic bead-based MALDI-TOF MS to serum samples from 10 immune-challenged and 10 healthy shrimps.The spectra were analyzed using FlexAnalysis 3.0 and ClinProTools 2.1 software.Thirteen peptide peaks significantly different between the two groups were selected as candidate biomarkers of lipopolysaccharide(LPS)-infection.The diagnostic model established by genetic algorithm using five of these peaks was able to discriminate LPS-challenged shrimps from healthy control shrimps with a sensitivity of 90% and a specificity of 100%.Our approach in MALDITOF MS-based peptidomics is a powerful tool for screening bioactive peptides or biomarkers derived from hemolymph,and will help to enable a better understanding of the innate immune response of shrimps.
文摘A simple and non-invasive method for detecting endometrial cancer in women with abnormal uterine bleeding is required. For this purpose, we prepared immuno-magnetic beads conjugated with anti-human EpCAM rat monoclonal antibody (mAb) for isolating exfoliated endometrial cells including endometrial cancer cells in vaginal discharge. The affinities of the anti-human EpCAM rat mAbs were analyzed by flow cytometry and immunocytochemistry and then magnetic beads were conjugated with the mAbs. The rate of retrieval of endometrial cells using the immuno-magnetic beads was calculated. Endometrial cells were isolated using the immuno-magnetic beads from the vaginal discharges of 22 patients with endometrial cancer and 16 non-malignant controls. The isolated cells were stained using endometrial cancer specific-mAbs and analyzed by flow cytometry and imaging cytometry. The immuno-magnetic beads conjugated with high-affinity mAb (clone 1456) appeared to have very low auto-fluorescence. Sufficient enrichment of Ep-CAMpositive cells using immuno-magnetic beads was observed in both simulation and clinical samples. The overall sensitivities of flow cytometry and imaging cytometry to detect endometrial cancer cells were 72.7% and 45.5%, respectively. Meanwhile, the overall specificities of flow cytometry and imaging cytometry for healthy controls were 75.0% and 81.3%, respectively. Our immuno-magnetic beads have very low auto-fluorescence, so they could be useful for fluorescent analysis, such as fluorescent immunochemical staining. In the future, these novel immuno-magnetic beads could be used for cytological study.
基金This project was supported by a grant from the National Natural Sciences Foundation of China (No. 30170944).
文摘To investigate the technique of sorting high-purity precartilaginous stem cells from rat’s perichondrium, neonatal rat’s perichondrium cells suspensions were incubated with monoclone anti-body of anti-fibroblast growth factor receptor-3 (anti-FGFR-3), and the labeled cells were separated from the suspension in the magnetic field by immuno-beads coated with the second antibody. Purity of the sorted neural stem cells was found to be 93.0 %-99.0 %, with living cells amounting to 80 %-85 %. The magnetic cell sorting system could effectively separate precartilaginous stem cells from perichondrium cell suspensions.
文摘On the basis of study on physical and chemical properties of magnetic bead (MB) in fly ash (FA), the paper gives out the separation methods of MB and results of three separating process. The result of comparative test in size, density, stability, magnetic material content, specific magnetic susceptibility (SMS), medium recovery oxidation resistance and wear resistance between MB and magnetic fines currently used in dense medium separation leads to that using MB recovered from fly ash is used as medium solids in coal cleaning in stead of magnetic fines not only have no influence upon taryests of separation, but can bring good economic and social benefits.
文摘目的评估自动化磁珠法提取血清脂溶性维生素应用液相色谱串联质谱法(liquid chromatography-tandem mass spectrometry,LC-MS/MS)检测的性能。方法收集200例临床剩余血清样本,采用自动化磁珠法提取血清中脂溶性维生素A,D_(2),D_(3),E和K;同时联合LC-MS/MS检测脂溶性维生素A,D_(2),D_(3),E和K的线性、定量限、精密度、正确度、携带污染率等性能指标以及基质效应。并比较此方法与传统萃取法检测结果的一致性。结果自动化磁珠法提取脂溶性维生素A,D_(2),D_(3),E和K线性相关系数均>0.99;五种物质的定量限分别为5,0.25,0.25,125和0.025ng/ml;批内精密度和批间精密度分别为0.66%~4.83%,0.15%~3.70%;平均加标回收率为87.05%~111.11%;基质效应为95.43%~99.07%;高-低值样本循环进样结果均值与低-低值样本循环进样结果均值之差,均小于低-低值样本循环进样结果均值的3s;统计学结果显示自动化磁珠法和传统萃取法提取的脂溶性维生素结果相关性良好(r>0.99),两种方法的检测结果无显著偏倚。结论自动化磁珠法提取脂溶性维生素的检测性能良好,有望提高样品通量和分析效率。
文摘构建一种基于杂交链式反应(hybridization chain reaction,HCR)扩增的适配体磁珠荧光传感器。巧妙设计序列HP和发卡序列H1、H2,其中HP是由适配体序列与触发序列结合而成的,并且序列互补形成稳定的二级结构。然后采用戊二醇反应和亲和素-生物素反应进行适配体功能化磁珠的制备。将阪崎肠杆菌与适配体磁珠一起孵育,HP中的适配体序列识别靶标,引起HP构象变化,露出触发序列,通过HCR触发H1和H2的链状组装,产生长双链DNA。荧光指示剂SYBR Green I以插层和小槽结合的方式与HCR产物的长双链结合。最后加入氧化石墨烯(graphene oxide,GO)后,游离的H1、H2和SYBR Green I将通过π-π堆积紧密吸附在GO表面,荧光信号被猝灭。HCR产物不能被吸附在GO表面,因此与HCR产物结合的SYBR Green I发出依赖于靶浓度的强荧光信号,从而实现阪崎肠杆菌的定量检测。本方法在纯培养条件下的检出限为2CFU/mL,对奶粉的检出限为8CFU/g,对奶粉样品的检测结果与传统微生物培养法具有良好的一致性。该方法具有无需DNA提取,快速、稳定性高、高特异性和高灵敏度等优点,因此为阪崎肠杆菌的现场快速检测提供了一种很有潜力的方法。
