| Isolation and Purification of Murine Microglial Cells for Flow Cytometry
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Author:
Date:
2016-01-05
[Abstract] The detailed protocol is used to isolate different cell types from murine brain as glial cells, including microglia, using an enzymatic digestion that minimizes cellular mortality. A Percoll gradient (30% to 80%) separation allows a maximal recovery of isolated murine microglial cells prior to flow cytometry analysis.
[摘要] 详细的协议是用于分离不同的细胞类型从鼠脑作为神经胶质细胞，包括小胶质细胞，使用最小化细胞死亡率的酶消化。 Percoll梯度（30％至80％）分离允许在流式细胞术分析之前分离的小鼠胶质细胞的最大恢复。
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| Human Blood Component Vaccinia Virus Neutralization Assay
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Author:
Date:
2015-12-05
[Abstract] Many therapeutic viruses, such as oncolytic viruses, vaccines, or gene therapy vectors, may be administered by the intravenous route to maximize their delivery to target tissues. Blood components, such as antibody, complement and blood cells (such as neutrophils, monocytes, T cells, B cells or platelets) may result in viral neutralization and therefore reduce the therapeutic efficacy. This protocol will describe an in vitro assay by which to test the interaction of viruses with blood components. The effect of various factors can be isolated through fractionation. While whole blood can offer the most physiologically relevant snapshot, plasma can investigate the effects of antibody in concert with complement, and heat inactivated plasma will interrogate the effect of antibody ...
[摘要] 许多治疗性病毒，例如溶瘤病毒，疫苗或基因治疗载体，可以通过静脉内途径施用以最大化它们向靶组织的递送。 血液组分，例如抗体，补体和血细胞（例如嗜中性粒细胞，单核细胞，T细胞，B细胞或血小板）可导致病毒中和并因此降低治疗功效。 该方案将描述用于测试病毒与血液组分的相互作用的体外测定。 各种因素的影响可以通过分馏分离。 尽管全血可以提供最具生理学相关性的快照，但血浆可以研究抗体与补体一起的作用，而热灭活的血浆将会询问单独的抗体的作用。
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| Longitudinal Bioluminescent Quantification of Three Dimensional Cell Growth
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Author:
Date:
2013-12-05
[Abstract] The use of three-dimensional (3D) cell culture systems is widely accepted as representing a more physiologically relevant means to propagate mammary epithelial and breast cancer cells. However, 3D cultures systems are plagued by several experimental and technical limitations as compared to their traditional 2D counterparts. For instance, quantifying the growth of mammary epithelial or breast cancer organoids longitudinally is particularly troublesome using standard [3H]thymidine or MTT assay systems, or using computer-assisted area calculations. Likewise, the nature of the multicellular aggregates and organoids formed by breast cancer cells under 3D conditions precludes efficient recovery of the cells from 3D matrices, an event that is time consuming and leads to spurious ...
[摘要] 使用三维（3D）细胞培养系统被广泛接受为代表更多生理相关手段以增殖乳腺上皮和乳腺癌细胞。然而，与其传统的2D对应物相比，3D培养系统受到若干实验和技术限制的困扰。例如，使用标准的[3 H]胸苷或MTT测定系统或使用计算机辅助面积计算，纵向量化乳腺上皮或乳腺癌组织的生长是特别麻烦的。同样，在3D条件下由乳腺癌细胞形成的多细胞聚集体和类器体的性质阻止了细胞从3D基质的有效回收，这是耗时的并且导致假的结果的事件。这里描述的测定利用萤火虫荧光素酶的稳定表达作为定量在3D矩阵内传播的细胞的纵向生长的手段。该技术的主要优点包括其高通量性质和在规定的时间段内纵向跟踪单个孔的能力，从而降低与测定性能相关的成本。最后，该技术可以容易地与药物治疗和/或遗传操作组合以测定它们对3D类器官生长的影响。
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