Latex beads phagocytosis capacity and ecto-NAD glycohydrolase activity of rat brain microglia cells in vitro

Conditions are described which allow the preparation in vitro of pure (> 95%) microglial cell cultures isolated from newborn rat brain. Such ameboid cells cultivated in vitro can efficiently phagocytize opsonized latex beads and are capable of ingesting more (100–200 beads of 1.1 μm diameter per cell) and larger (6.4 μm) particles than other nerve cells, such as oligodendrocytes and astroglia. The microglial cells also show an important ecto-NAD⁺ glycohydrolase activity which is characteristic of phagocytic cells. We noted that the phagocytic capacity and ecto-NAD⁺ glycohydrolase of these cells were correlated and increased notably during the in vitro culture.Microglia cultivated in vitro appear to be a good model to study the activation of phagocytic properties in the central nervous system and corresponding modulation by natural or pharmacological immunomodulators.     

Use of microcarrier beads to facilitate epithelial outgrowth inside collagen gel

Summary Multicellular aggregates of some epithelial cell types rapidly produce ductlike protuberances when cultured inside a matrix of hydrated collagen gel. However, some epithelioid cells aggregate poorly. These cell types will produce outgrowths rapidly if they are attached to microcarrier beads before embedding them in the collagen matrix. Methods are described for preparing such cultures.     

塑料球固相二抗通用分离剂在RIA中的应用

本文报道以聚苯乙烯塑料球为载体．经化学处理制成固相二抗，作为ＲＩＡ技术中的通用分离剂，并先后用于半抗原及大分子蛋白质类的测定。实验结果表明：塑料球固相二抗制备法工艺简便，最佳包被条件为１００ｍｍｏｌ／ＬＰＨ４．８柠檬酸盐缓冲液，不需特殊条件，便于推广应用。本方法不需离心，且各项指标均达对照ＰＲ分离法的标准。     

The use of Cytodex microcarrier beads in patch-clamp studies on cultured epithelial cells

This paper describes an economical attachment substratum for use in patch-clamp experiments on cultured epithelial cells. It consists of Cytodex microcarrier beads on which growing cells present a unique profile perspective that increases the ease with which patch-clamp seals can be obtained. Cells from the mouse mandibular gland cell line, ST₈₈₅, were grown on the beads and studied using standard patch-clamp techniques. The results were indistinguishable from those obtained from cells grown in monolayers on Petri dishes. The method can be applied in many types of patch-clamp experiment and used with a wide variety of cell types. It is especially useful if one wishes to do experiments in the cell-attached or whole-cell configurations and needs to add drugs or hormones to the external surface of the cell.This project was supported by the National Health and Medical Research Council of Australia.     

Immunomagnetic exclusion of E-cadherin-positive hepatoblasts in fetal mouse liver cell cultures impairs morphogenesis and gene expression of sinusoidal endothelial cells

Previous studies have shown that various cell-cell interactions between hepatoblasts and nonparenchymal cells, including sinusoidal endothelial cells and stellate cells, are indispensable for the development of fetal murine hepatic architecture. The present study was undertaken to determine the effects of hepatoblasts on the sinusoidal structural formation using a culture system of fetal mouse livers. Primitive sinusoidal structures extensively developed in fetal livers, and were composed of LYVE-1- and PECAM-1-positive endothelial cells, desmin-positive stellate cells and F4/80-positive macrophages. When fetal liver cells at 12.5 days of gestation were cultured in vitro, hepatoblasts spread on glass slides and gave rise to hepatocytes on day 5. Desmin-positive stellate cells also spread on the glass slides. PECAM-1-positive endothelial cells became slender and developed into anastomosing capillary networks. When fetal liver cells were cultured without hepatoblasts, which were excluded by an immunomagnetic method using anti-E-cadherin antibodies, endothelial cells had impaired growth and capillary formation. These results demonstrated that capillary formation of endothelial cells was induced by the presence of hepatoblasts. VEGF and the conditioned medium containing humoral factors produced by hepatoblasts/hepatocytes did not induce capillary formation of endothelial cells in cultures of nonparenchymal cells, although they significantly increased the number of endothelial cells on the glass slides. The presence of hepatoblasts also significantly stimulated expression of CD32b mRNA, which is a sinusoidal endothelial marker. Hepatoblasts may work as a positive stimulator of sinusoid morphogenesis and maturation in liver development, in which a signal other than VEGF may play a decisive role, together with VEGF.     

免疫磁珠两步法体外分离纯化小鼠脾脏CD4+CD25+调节性T细胞的应用研究

目的 研究免疫磁珠两步法体外分离纯化小鼠脾脏CD4+CD25+调节性T细胞(regulatory T cells,Treg)的效果.方法 将C57BL/6(H-2b)小鼠脾脏剪碎、过滤、去除红细胞后离心洗涤,获得小鼠脾细胞悬液.用Ficoll法分离获得单个核细胞后,通过免疫磁珠阴性加阳性选择两步法分离获得CD4+CD2...     

免疫磁珠法分离纯化人肾癌CD133~+细胞实验研究

目的:本研究通过分析CD133+细胞在肾细胞癌患者肿瘤组织的表达情况,寻找肾癌中CD133+细胞分离纯化方法,探寻CD133+细胞与肾癌发生中的作用。方法:①应用胎盼蓝拒染试验检测细胞活性。②应用免疫磁珠法分离纯化肿瘤组织中CD133+细胞。③应用流式细胞仪检测肾透明细胞癌标本中CD133+细胞表达情况。结果:30例肾透明细胞癌中CD133+比为(13.08±1.39)%,与病理分级、临床分期无关。免疫磁珠法分离纯化CD133+纯度为(75±2.4)%,细胞活性不受影响。结论:免疫磁珠技术纯化肾癌CD133+细胞成熟有效,CD133+细胞可能是肾癌干细胞。     

免疫磁珠捕获法分离抗酸杆菌的实验研究

目的建立免疫磁珠捕获法,用于分离抗酸杆菌,以提高痰菌阴性肺结核的诊断率。方法选取浸润型肺结核患者19例作为实验组,对照组6例,为肺部普通细菌感染者。受检者留取痰液,分别做直接涂片抗酸染色检查及免疫磁珠分选后抗酸染色检查。结果 19例浸润型肺结核患者中,7例痰涂片抗酸染色检查阳性,免疫磁珠分选后抗酸染色仍阳性;在12例痰涂片阴性肺结核患者中,经免疫磁珠分选后6例抗酸染色阳性。6例普通细菌性肺炎患者经免疫磁珠分选后抗酸染色均为阴性。结论用免疫磁珠捕获法分离抗酸杆菌能显著提高痰菌阴性肺结核的诊断效率。     

Biofunctionalized magnetic nanoparticles for specifically detecting biomarkers of Alzheimer's disease in vitro

Magnetic nanoparticles biofunctionalized with antibodies against β-amyloid-40 (Aβ-40) and Aβ-42, which are promising biomarkers related to Alzheimer's disease (AD), were synthesized. We characterized the size distribution, saturated magnetizations, and stability of the magnetic nanoparticles conjugated with anti-Aβ antibody. In combination with immunomagnetic reduction technology, it is demonstrated such biofunctionalized magnetic nanoparticles are able to label Aβs specifically. The ultralow-detection limits of assaying Aβs in vitro using the magnetic nanoparticles via immunomagnetic reduction are determined to a concentration of ～10 ppt (10 pg/mL). Further, immunomagnetic reduction signals of Aβ-40 and Aβ-42 in human plasma from normal samples and AD patients were analyzed, and the results showed a significant difference between these two groups. These results show the feasibility of using magnetic nanoparticles with Aβs as reagents for assaying low-concentration Aβs through immunomagnetic reduction, and also provide a promising new method for early diagnosis of Alzheimer's disease from human blood plasma.