Efficacy of folate-coupled quantum dots immunomagnetic beads method for detecting circulating tumor cells in epithelial ovarian cancer and the association of circulating tumor cells with clinicopathological features / 肿瘤研究与临床

Objective:To explore the effectiveness of folate-coupled quantum dots (FA-QD) immunomagnetic beads method for detecting circulating tumor cells (CTC) in epithelial ovarian cancer and the association of CTC with clinicopathological features of tumor patients.Methods:A total of 67 ovarian cancer patients in Shanxi Provincial People's Hospital from August 2019 to January 2020 were selected. Ovarian cancer SKOV-3 cells were divided into 5 cell number gradients (0, 100, 150, 200, 300), the detection rates of CTC were compared by using epithelial cell adhesion molecule (EpCAM) immunomagnetic beads (single standard method) and FA-QD immunomagnetic beads method (double standard method). The number of CTC in peripheral blood of ovarian cancer patients was detected by using FA-QD immunomagnetic beads method, and those with positive CTC under fluorescence microscope were treated as CTC positive patients. The association of CTC with clinicopathological factors and tumor markers of tumor patients was analyzed.Results:The average capture efficiency rate of CTC in SKOV-3 cells detected by FA-QD immunomagnetic beads method (83.4%) was higher than that by EpCAM immunomagnetic beads method (70.3%). Among 67 patients of ovarian cancer, the proportion of CTC positive patients was 30.0% (3/10) in stage Ⅰ-Ⅱ, 91.9% (34/37) in stage Ⅲ, 95.0% (19/20) in stage Ⅳ, and the difference was statistically significant ( P < 0.05). The proportion of CTC positive patients with lymph node metastasis was higher than that of patients without lymph node metastasis [97.1% (33/34) vs. 69.7% (23/33)], and the difference was statistically significant ( P < 0.05). The proportion of CTC positive patients with human epididymis protein 4 (HE4)>110 pmol/L was lower than that of patients with HE4 ≤ 110 pmol/L [58.8% (10/17) vs. 92.0% (46/50)], and the difference was statistically significant ( P = 0.005). There were no statistically significant differences in the proportion of CTC positive patients stratified by age, menopause, pathological differentiation, distant metastasis, carbohydrate antigen (CA) 125, CA199, carcino-embryonic antigen (CEA) (all P > 0.05). Conclusions:FA-QD immunomagnetic beads method can effectively detect CTC in peripheral blood of patients with epithelial ovarian cancer. The level of CTC in patients with epithelial ovarian cancer is related to lymph node metastasis, clinical TNM stage and HE4 level.

Detection of N-ethyl-N-nitrosourea-induced Pig-a gene mutation in rats by immunomagnetic separation technique combined with flow cytometry / 中国实验动物学报

Objective To optimize the detection test of Pig-a gene mutation in peripheral blood of rats by enriching and detecting mutant erythrocytes, using immunomagnetic separation technique in combination with flow cytometry. Methods SD rats were administered with 20,40 and 80 mg/(kg·bw)doses of N-ethyl-N-nitrosourea(ENU) continually for 3 days. The peripheral blood samples of rats were collected on the 7th,14th and 28th days, respectively, after treatment. Immunomagnetic separation columns were used to enrich RETCD59- and RBCCD59- cells, and then flow cytometry was used to count the number of pre-column and post-column peripheral erythrocytes. Results Compared with the control group,the frequencies of RETCD59- and RBCCD59- were significantly increased in each ENU group(P<0.05). With immunomagnetic separation technique, the test of Pig-a gene mutation of a sample could be completed within 3 minutes,and the number of detected RETCD59- or RBCCD59-cells was up to 2×104or 9×104, respectively. Conclusions In this study,immunomagnetic separation in combination with flow cytometry is used to establish and optimize the Pig-a gene mutation test in rat peripheral blood,showing a high-throughput detection and improved accuracy and efficiency of the experiment.

Fluorescent Microspheres Lateral Flow Assay Based on Immunomagnetic Separation for Detection of S.choleraesuis / 分析化学

Immunomagnetic separation (IMS) was coupled with fluorescent microspheres lateral flow assay (FM-LFA) for rapid detection of S.choleraesuis in this study.The target bacteria were firstly enriched from sample by immunomagnetic beads (IMBs),then eluted by heat treatment and detected by fluorescent microspheres lateral flow test strip.The IMBs was labeled with 30 μg/mg antibody,and the capture efficiency was greater than 90％ against 102-106 CFU/mL of S.choleraesuis with great specificity.The immunofluorescent microspheres were prepared by coupling 300 μg of 11 D8-D4 monoclonal antibody with 1 mg of fluorescent microspheres at pH 6.Monoclonal antibody 5F11-B11 (2.0 mg/mL) and donkey anti-mouse IgG (1.0 mg/mL) were sprayed on nitrocellulose membrane as test line and control line,respectively.The FM-LFA based on IMS was used to detect S.choleraesuis in PBS and milk.The limits of detection in PBS buffer and milk were 1.5×105 CFU/mL and 7.6×105 CFU/mL respectively,which were 10 and 200 times lower than that of traditional fluorescent microspheres lateral flow assay,respectively.The results showed that the method,which could enrich S.choleraesuis in milk effectively,could avoid matrix interference and improve the detection sensitivity,thus had a good application prospect.

Mycobacterium paratuberculosis detection in cow's milk in Argentina by immunomagnetic separation-PCR

Abstract The aim of this study was to standardize a diagnosis procedure to detect Mycobacterium avium subsp. paratuberculosis (Map) DNA in raw cow milk samples under field conditions. A procedure that combines both immunomagnetic separation and IS900 -PCR detection (IMS-IS1 PCR) was employed on milk samples from 265 lactating Holstein cows from Map infected and uninfected herds in Argentina. IMS-IS1 PCR results were analyzed and compared with those obtained from milk and fecal culture and serum ELISA. The extent of agreement between both tests was determined by the Kappa test. IMS-IS1 PCR showed a detection limit of 101 CFU of Map/mL of milk, when 50:50 mix of monoclonal and polyclonal antibodies were used to coat magnetic beads. All of the 118 samples from the Map uninfected herds were negative for the set of the tests. In Map infected herds, 80 out of 147 cows tested positive by milk IMS-IS1 PCR (55%), of which 2 (1.4%) were also positive by milk culture, 15 (10%) by fecal culture, and 20 (14%) by serum ELISA. Kappa statistics (95% CI) showed a slight agreement between the different tests (<0.20), and the proportions of agreement were ≤0.55. The IMS-IS1 PCR method detected Map in milk of the cows that were not positive in other techniques. This is the first report dealing with the application of IMS-IS1 PCR in the detection of Map in raw milk samples under field conditions in Argentina.

Método imunomagnético associado ao meio MesenCult® na obtenção de células mononucleares da medula óssea de coelhos negativas para o anticorpo monoclonal CD45 / Immunomagnetic method associated with MesenCult® medium to obtain bone marrow mononuclear cells from rabbits negative for CD45 monoclonal antibody

O objetivo detse artigo é de descrever um protocolo de isolamento das células mononucleares da medula óssea de coelhos, seguido de purificação celular por depleção negativa com o anticorpo monoclonal CD45 e posterior expansão em meio de cultura MesenCult®. Dez coelhos machos adultos, da raça Nova Zelândia, com idade média de 1,0±0,2 anos e peso médio 3,5±0,24kg, foram utilizados para padronização da metodologia. O isolamento das células mononuclares da medula óssea foi realizado pelo gradiente de densidade Ficoll-paque® e a purificação e obtenção das células- pela depleção negativa com o anticorpo monoclonal CD45 em base imunomagnética. A população celular obtida foi expandida posteriormente em meio de cultura MesenCult®. No isolamento pelo gradiente de icoll-Paque® foi obtido um rendimento médio de 7,31x106 células/mL. Após purificação e obtenção das possíveis células-tronco mesenquimais pela base imunomagnética, houve um decréscimo do rendimento para 2,28x106 células/mL, mas o processo de expansão foi incrementado pelo cultivo celular. Os resultados indicaram que as células obtidas da fração mononuclear da medula óssea, cultivadas in vitro foram capazes de gerar células aderentes 24 horas após o cultivo, com predominância de células fibroblastóides sugestivas de células-tronco mesenquimais. Concluiu-se que a obtenção de células-tronco mesenquimais pode ser alcançada após purificação das células mononucleares da medula óssea de coelhos pelo método imunomagético, o meio de cultura MesenCult® proporciona um ambiente adequado para a rápida expansão in vitro e o número de passagens exerce influência negativa sobre as características morfológicas das células.

The objective of this study was to describe guidelines for the isolation of bone marrow mononuclear cells from rabbits, followed by cell purification by negative depletion with CD45 monoclonal antibody, and further expansion in MesenCult® medium. Ten adult male New Zealand White rabbits, age average of 1.0±0.2 years and weighting 3.5±0.24kg, were used to obtain a standardized method. The mononuclear cells of the bone marrow were isolated with Ficoll-paque® density gradient centrifugation, and the cell purification and acquisition was completed by negative depletion with CD45 monoclonal antibody in immunomagnetic base. The cell population obtained was expanded in MesenCult® medium. Through isolation with Ficoll-paque® density gradient was possible to obtain an average yield of 7.31x106 cells/mL. After purification and acquisiton of potential mesenchymal stem cells by the immunomagnetic base, there was a yield decrease to 2.28x106 cells/mL; however the expansion process was increased in cell culture. The results indicated that cells obtained from the mononuclear fraction of bone marrow and cultivated in vitro were capable to generate adherent cells 24 hours after culture, with predominance of fibroblastoid cells suggestive of mesenchymal stem cells. It can be concluded that mesenchymal stem cells can be achieved with purified rabbit bone marrow mononuclear cells through the immunomagnetic method, as the MesenCult® medium provides a suitable environment for a quick in vitro expansion, and the number of passages exerts negative influence on the morphological characteristics.

Research progress on enrichment and isolation of circulating tumor cells using immunomagnetic separation based microfluidic technology / 中华检验医学杂志

Circulating tumor cells ( CTCs ) exists in peripheral blood of patients with a variety of malignant tumors , the accurate detection of CTCs helps early detection of cancer , and can be used to guide individualized treatment of cancer patients , rapid assessment of tumor chemotherapy drugs and detection of tumor recurrence .However , the CTCs in peripheral blood of cancer patients were extremely low , their detection requires efficient and specific enrichment and separation methods .The new microfluidic technology can accurately control the microliquid preciously , combined with the immunomagnetic separation technology , has been applied in detecting CTCs , which can save the blood samples of CTCs , has the advantages of simple operation , fast sorting speed , high specificity and high purity .

Extraction of prostate cancer stem cells using self-designed magnetic beads / 中国组织工程研究

BACKGROUND:Effective sorting of prostate cancer stem cels is the basis of experimental studies in prostate cancer developing. The most common sorting method is magnetic-activated cel sorting. OBJECTIVE:To separate CD133+/CD44+ cels in prostate cancer tissues using self-designed magnetic beads folowed by culture, passage and immunological identification. METHODS:Self-designed magnetic microspheres were applied to establish immunomagnetic beads to sort CD133+/CD44+ cels in prostate cancer tissues. The sorted cels were cultured in serum-free medium. The sphere formation, cel morphology, and proliferation ability after cel passage were statisticaly compared between the sorted cels and the normal tumor cel lines. Immunofluorescence detection was performed to detect the expression of specific antibodies. RESULTS AND CONCLUSION:Self-designed immunomagnetic beads had smal diameter and a high-sorting effect. The sorted cels possessed a high capacity of microsphere formation. After cel culture and passage, the cels highly expressed CD133 and CD44 antigens. The sorted cels with no induction had varying shapes and grew vigorously. After induction with transforming growth factor-β, the cultured cels were noted to have a single shape and grow slowly. The cel proliferation ability of sorted cels in these two groups differed significantly from that of the normal cancer cel lines (bothP < 0.05). In conclusion, the CD133+/CD44+ cels sorted from prostate tumor cels possessed cel morphology and function characteristics of stem cels which can provide a basis for extraction and culture of prostate cancer stem cels. Cite this article:Gong R, Li SY, Huo ZX, Ding H, Sun EL. Extraction of prostate cancer stem cels using self-designed magnetic beads. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):36-41.

Comparison Analysis of HLA-B27 Detected by PCR-SBT and IMS-ELISA in AS Patients / 现代检验医学杂志

Objective To compare PCR-SBT to IMS-ELISA in the HLA-B27 detection in the ankylosing spondylitis (AS)pa-tients.Methods Simultaneously,PCR-SBT and IMS-ELISA were used to detect the HLA-B27 expression in peripheral blood samples which were suspected patients with AS from 120 cases.Chisquare test of paired design and the area under curve of receiver operating characteristics of SPSS17.0 software were used to evaluate the value of PCR-SBT and IMS-ELISA in HLA-B27 detection of AS patients.Results Among 120 cases of suspected patients with AS,the positive rates of HLA-B27 detected by PCR-SBT and IMS-ELISA were 45.83%(55/120)and 37.50% (45/120),respectively.There was statistical difference between the two methods in the HLA-B27 detection (χ2 =59.455,P =0.000).The sensibility and spe-cificity of PCR-SBT were 96.36% and 96.92%,respectively.While the sensibility and the specificity of IMS-ELISA were 69.09% and 89.23%,respectively.Area under the curve of two methods were 0.966 and 0.792,respectively.Conclusion In comparison with IMS-ELISA,the sensibility and the specificity of PCR-SBT in HLA-B27 detection were higher in AS diag-nosis,that is to say,PCR-SBT is better in HLA-B27 detection and AS diagnosis.

A novel method for detection of circulating tumor cells in peripheral blood of patients with lung cancer using immunomagnetic beads device / 中华检验医学杂志

Objective To establish a novel method for detecting circulating tumor cells (CTCs) in phripheral blood of lung cancer patients with high sensitivity and specificity.Methods Experimental study.42 cases of initial treatment patient who underwent resection and diagnosed to be non-small cell lung cancer by biopsy were studied,including 7 patients at stage Ⅰ,9 patients at stage Ⅱ,16 patients at stage Ⅲ and 10 patients at stage Ⅳ.As a control group,20 cases of healthy volunteers were selected.A series of experiments was conducted to determine the efficiency of tumor cells isolation,in which varied concentration (50,100,200,500,1000 cells) of A549 cells spiked into 2 ml peripheral blood drawn from healthy donors.The blood was removed of unwanted erythrocytes by lysis buffer,and made the rest of nucleated cells incubate with anti-EpCAM magnetic beads,then separated and enriched by a specific detector.All epithelia cells were retained on a slide because of a magnetic force and identified by H&E staining protocol.On the basis of cell recovery rate we calculated the sensitivity of tumor cells isolation.20 blood samples taken from healthy individuals were also detected to validate the specificity of this method.Samples of 42 patients with lung cancer were assayed for CTCs detection by above method.The correction of CTCs quantity with the patients' clinical features,for example,ages,gender,clinical stage,tumor size was analyzed in lung cancer patients by chi-square statistics.The correction of recovery cells with the spiked cells were assayed by linear correlation.Results The recovery rate was ranging from 68％ to 82％ by spiking varying numbers of A549 lung cancer cells into 2ml blood samples of healthy volunteers.Regression analysis of number of recovered vs.spiked A549 cells yielded a regression equation of Y =0.6419X + 8.8875.The number of CTCs detected has signification correlate with the cells spiked (R2 =0.9916,P ＜ 0.05),Eighteen of the 42 patients (43％) were found have CTCs in peripheral blood.The detection rate of lung cancer cells was 0 at stage Ⅰ,the detection rate of lung cancer cells was 11.1％ at stage Ⅱ,the detection rate of lung cancer cells was 62.5％ at stage Ⅲ and the detection rate of lung cancer cells was 70％ at stage Ⅳ.The positive rate of CTCs has no signification correlate with ages and gender of patients and tumor size (P ＞ 0.05),has signification with the clinical stage (P ＜ 0.05).None of the peripheral blood samples of the 20 healthy subjects analyzed was found to have CTCs.Conclusions This novel immunomagnetic separation technology is a sensitive and specific method,which provides a new tool allowing for feasible and specific detection of CTCs in lung cancer patients.The level of CTCs increases with the clinical stage and tumor size increased,which has important value to discover the early stage micrometastasis and redefine the clinical stage.But further multicenter and large sample clinical research are needed to confirm its clinical value.

Viabilidad de un aislado de Escherichia coli O157: H7 en queso tipo guayanés usando separación inmunomagnética como herramienta de recuperación / The viability of an Escherichia coli O157: H7 strain in guayanés Ccheese using immunomagnetic separation as a tool for recovery

Se determinó la viabilidad de Escherichia coli O157:H7 en queso guayanés de manufactura artesanal, evaluando distintos esquemas de aislamiento basados en separación inmunomagnética (SIM). Unidades de queso (25 g) fueron inoculadas con 25 y 250 cel/g del patógeno y almacenadas a 4°C. Las piezas se analizaron los días 0, 2, 6, 8 y 10 post-inoculación a través de distintos esquemas de separación inmunomagnética (SIM) que incluían dos caldos de enriquecimiento: agua de peptona buferada sin inhibidores (APB-SI) y agua de peptona buferada con vancomicina, cefixime y telurito (APB-VCT) y dos agares de aislamiento del inmunoseparado: agar MacConkey sorbitol (MCS) y agar MacConkey sorbitol con telurito y cefixime (MCS-TC). Los resultados demostraron la viabilidad del patógeno hasta por 10 días post-inoculación y en el transcurso de este tiempo, para algunos de los esquemas aplicados sobre la base de SIM, se logró un incremento en los porcentajes de recuperación, lo que indica que el número de células inoculadas se elevó con el tiempo. En cuanto a la utilidad de la SIM para la recuperación del patógeno, se observó variaciones en los porcentajes de aislamiento en función del caldo de enriquecimiento y el nivel de células inoculadas. Los mayores porcentajes de recuperación se obtuvieron en las piezas inoculadas con 250 cel/g, con rangos del 35 al 85 por ciento (día 0 y 10 respectivamente) en el mejor de los esquemas SIM (APB-SI/SIM/MCS), mientras que para niveles de 25 cel/g, en el mejor de los casos (APB-SI/SIM/MCS), durante los primeros 6 días no superó el 15 por ciento. El caldo de enriquecimiento de mejor desempeño fue APB-SI (p <0,05) y no se observó diferencias en los porcentajes de recuperación (p>0,05) en función de los agares utilizados (MCS y MCS-TC) para la siembra del inmunoseparado

The viability of an Escherichia coli O157:H7 strain in cottage-industry Guayanes cheese was determined by evaluating several isolation protocols based on immunomagnetic separation (IMS). Cheese units (25 g) were inoculated with 25 and 250 cel/g of this pathogen and stored at 4°C. The pieces were analized at 0, 2, 6, 8 and 10 days post-inoculation through several IMS protocols including two enrichment broths: buffered peptone water without inhibitors (BPW-WI) and buffered peptone water with vancomicyn, cefixime and telurite (BPW-VCT) and two immunoseparation isolation agars: MacConkey-sorbitol agar (MSA) and MacConkey-sorbitol agar with cefixime and telurite (MSA-CT). Results demonstrated the viability of the pathogen for up to 10 days post-inoculation, and during this time, for some of the schemes applied on the IMS base, an increase in recovery percentages was achieved, indicating that the number of inoculated cells increased with time. In terms of the utility of IMS for recovering the pathogen, variations in the isolation percentages were observed in terms of the enrichment broth and the level of inoculated cells. The biggest recovery percentages were obtained in pieces inoculated with 250 cel/g, with ranges between 35 and 85 percent (days 0 and 10 respectively) in the best IMS scheme (BPW-WI/IMS/MSA), while, at levels of 25 cel/g, in the best case (BPW-WI/IMS/MSA), 15 percent was not surpassed during the first six days. The best performing enrichment broth was BPW-WI (p<0.05) and differences in the recovery percentages (p>0.05) were not observed in relation to the agars (MSA and MSA-CT) used for sowing the immunoseparator

Cryptosporidium Oocyst Detection in Water Samples: Floatation Technique Enhanced with Immunofluorescence Is as Effective as Immunomagnetic Separation Method

Cryptosporidium can cause gastrointestinal diseases worldwide, consequently posing public health problems and economic burden. Effective techniques for detecting contaminated oocysts in water are important to prevent and control the contamination. Immunomagnetic separation (IMS) method has been widely employed recently due to its efficiency, but, it is costly. Sucrose floatation technique is generally used for separating organisms by using their different specific gravity. It is effective and cheap but time consuming as well as requiring highly skilled personnel. Water turbidity and parasite load in water sample are additional factors affecting to the recovery rate of those 2 methods. We compared the efficiency of IMS and sucrose floatation methods to recover the spiked Cryptosporidium oocysts in various turbidity water samples. Cryptosporidium oocysts concentration at 1, 10(1), 10(2), and 10(3) per 10 microliter were spiked into 3 sets of 10 ml-water turbidity (5, 50, and 500 NTU). The recovery rate of the 2 methods was not different. Oocyst load at the concentration < 10(2) per 10 ml yielded unreliable results. Water turbidity at 500 NTU decreased the recovery rate of both techniques. The combination of sucrose floatation and immunofluorescense assay techniques (SF-FA) showed higher recovery rate than IMS and immunofluorescense assay (IMS-FA). We used this SF-FA to detect Cryptosporidium and Giardia from the river water samples and found 9 and 19 out of 30 (30% and 63.3%) positive, respectively. Our results favored sucrose floatation technique enhanced with immunofluorescense assay for detecting contaminated protozoa in water samples in general laboratories and in the real practical setting.

Detection of salmonella sp in chicken cuts using immunomagnetic separation

The immunomagnetic separation (IMS) is a technique that has been used to increase sensitivity and specificity and to decrease the time required for detection of Salmonella in foods through different methodologies. In this work we report on the development of a method for detection of Salmonella in chicken cuts using in house antibody-sensitized microspheres associated to conventional plating in selective agar (IMS-plating). First, protein A-coated microspheres were sensitized with polyclonal antibodies against lipopolysacharide and flagella from salmonellae and used to standardize a procedure for capturing Salmonella Enteritidis from pure cultures and detection in selective agar. Subsequently, samples of chicken meat experimentally contaminated with S. Enteritidis were analyzed immediately after contamination and after 24h of refrigeration using three enrichment protocols. The detection limit of the IMS-plating procedure after standardization with pure culture was about 2x10 CFU/mL. The protocol using non-selective enrichment for 6-8h, selective enrichment for 16-18h and a post-enrichment for 4h gave the best results of S. Enteritidis detection by IMS-plating in experimentally contaminated meat. IMS-plating using this protocol was compared to the standard culture method for salmonellae detection in naturally contaminated chicken cuts and yielded 100 percent sensitivity and 94 percent specificity. The method developed using in house prepared magnetic microespheres for IMS and plating in selective agar was able to diminish by at least one day the time required for detection of Salmonella in chicken products by the conventional culture method.

A separação imunomagnética (IMS) é uma técnica que tem sido associada a diferentes métodos de detecção de Salmonella em alimentos para aumentar a sensibilidade e a especificidade e diminuir o tempo de análise. Neste trabalho é comunicada a obtenção de microesferas magnéticas sensibilizadas com anticorpos anti-Salmonella e seu uso em associação com a metodologia de cultivo convencional para desenvolver um método de detecção de salmonelas em cortes de frango (IMS-plaqueamento). Inicialmente, microesferas cobertas com proteína A foram sensibilizadas com anticorpos policlonais contra lipopolissacarídeo e flagelo de salmonelas e usadas na padronização de um procedimento que captura Salmonella Enteritidis em cultivo puro e faz posterior detecção em ágar seletivo. A seguir, amostras de carne de frango experimentalmente contaminadas com S. Enteritidis foram analisadas imediatamente após a contaminação e após 24h de refrigeração utilizando três protocolos de enriquecimento. O limite de detecção foi cerca de 2x10 UFC/mL. O protocolo que incluiu enriquecimento não-seletivo por 6-8h, enriquecimento seletivo por 16-18h e pós-enriquecimento por 4h foi o que proporcionou melhor resultado na detecção de S. Enteritidis em carne de frango experimentalmente contaminada. Este protocolo foi comparado à metodologia convencional em estudo de detecção de salmonelas em cortes de frango naturalmente contaminados e obteve 100 por cento de sensibilidade e 94 por cento de especificidade. O método desenvolvido foi capaz de diminuir em pelo menos um dia o tempo de detecção de salmonelas em cortes de frango pela metodologia convencional.

Magnetic activated cell sorting combined with a panel of monoclonal antibodies in detecting free cancer cells in analogue malignant ascites / 中华消化杂志

Background Magnetic activated cell sorting(MACS)is a new immunomagnetic separation technique.It's principle is based on the specialized recognition between the antibody and antigen.When directed or indirected coupled with the 50 nm magnetic beads,the antibody can linked with the cells which express the corresponding antigen,and then leads to the magnetic cell separation in a high intensity and high-gradient magnetic field.The method has higher separative purity and recovery and been made use of enriching tumor cells and tumor stem cells.It could be also utilized to enrich the rare tumor cells in fluid specimen.Objective To evaluate the enriching efficiency of detecting free cancer cells in analogue malignant ascites using MACS and a panel of tumor-specific markers.Methods Five species of tumor cell lines correlated to the diseases resulting in malignant ascites were selected and the expressions of EpCAM,CAl25,CEA,TAG-72 and their mixture in these tumor cell lines were detected using immunofluorescence reactions and flow cytometry(FCM).The tumor cells were spiked into mononuclear cells by different ratios to analogue the main ingredients of malignant ascites.The efficiency of MACS combinded with mixture antibodies in separating tumor cells was compared with single antibody.Results The FCM revealed that the expression of the mixture antibodies in 5 tumor cancer cell lines was significantly higher than that in single antibody.The mean recovery rates of spiked tumor cells were enhanced by using a panel of monoclonal antibodies combined with MACS than single antibody,especially in two gastric cancer cells and colon cancer cells(69.18%±20.84%VS 45.23%±11.54%,78.75%±15.42%vs 59.73%±16.64%and 85.63%±12.30%VS 76.88%±8.65%,respectively),the ovarian cancer cells was the next(32.49%±3.58%vs31.79%±4.82%),and the liver cancer cells was the lowest(11.78%±0.43%VS 7.16%±0.46%).Conclusions The detection of free cancer cells from malignant ascites by MACS combined with a panel of monoelonal antibodies is more effectively than single antibody.The method has the potencial value of clinical application to the malignant ascites caused by gastroenteric tumor.

Immunomagnetic separation to isolate and purify precartilaginous stem cells from neonate rats / 中华创伤杂志

95%). Conclusion Immunomagnetic separation can effectively isolate and purify PSCs.

Investigation of Giardia lamblia and Crytosporidium parvum in Drinking Water in Tianjin and Shenyang / 环境与健康杂志

0.05).Conclusion The standard test methods are more effective in detecting G.lamblia than C.parvum.No G.lamblia and C.parvum contamination has been found in drinking water in the Tianjin and Shenyang.

DEVELOPMENT OF AN HIGH-EFFICIENT IMMUNOMAGNETIC ISOLATOR FOR THE PURIFICATION OF CD34~+ HEMATOPOIETIC CELLS / 解放军医学杂志

According to the principle of immunomagnetic separation, a novel high-efficient immunomagnetic isolator termed as CMSI 100 for the purification of human CD34 + hematopoietic cells was designed and successfully developed. To do this , neodymium iron boron having magnetic properties superior to any other permanent magnet materials was selected as the key parts of CMSI 100 isolator. The core of the magnet assembly was constructed by sandwiching mild steel bars between 4 pieces of magnet bars of neodymium iron boron. Each piece of magnet bars must be limited to the specifications of 3.5cm?2.0cm?0.3cm so that 2 700 gauss magnetic attractive force could be exerted at the magnet surface. Otherwise magnetic field above the surface of the magnet assembly is either very stronger or weaker, both of which are not beneficial to the enrichment of CD34 + hematopoietic cells. With CMSI 100 immunomagnetic isolator, more than 90% purity and over 95% viability of CD34 + hematopoietic cells could be obtained. These data indicate that CMSI 100 immunomagnetic isolator is as good as the internationally used Isolex TM 50 made by Baxter Company in USA.

Clinical significance of detecting circulating tumor cells in patients with lung cancer in early stage / 解放军医学杂志

Objective To discuss the feasibility and clinical significance of detecting circulating tumor cells(CTCs) in peripheral blood of patients with lung cancer in early stage. Methods Three groups of patients (Group A: 15 cases with early stage lung cancer; Group B: 20 cases with benign pulmonary disease; Group C: 20 healthy volunteers) were enrolled for detection of CTCs using immunomagnetic separation(IMS)+ immunocytochemistry(ICC) method and RT-PCR method. The patients in Group A were followed up for 6-9 months. Results By using IMS+ICC it was revealed that in 5 cases of Group A the result was positive, while it was negative in all cases of Group B and Group C. By using RT-PCR method, the result was positive in 7 cases of Group A, 8 cases of Group B and 6 cases of Group C. In the follow-up period, 2 patients of Group A, who were found positive for CTC before and after operation, showed recurrence. Conclusion IMS + ICC method was a sensitive method to detect CTCs in patients with early stage NSCLC. The detection of CTCs in patients with early stage lung cancer might be a relationship with the clinical prognosis of the patient.