Sullivan Heather A, Wickersham Ian R
Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139;
McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139.
Cold Spring Harb Protoc. 2015 Apr 1;2015(4):386-91. doi: 10.1101/pdb.prot075887.
Rabies viral and lentiviral vectors are very useful tools for neuroscientists, but high titer and purity are critical for in vivo applications. Here we present a protocol for concentration and purification of viral stocks by ultracentrifugation on a sucrose step gradient to remove impurities of both higher and lower densities than the virus itself, with sucrose removed by a subsequent pelleting step. The final stocks are concentrated in volume by a factor of up to 1000, with higher expected purity than is obtained following previously published protocols for preparing G-deleted rabies viral vectors.
狂犬病病毒载体和慢病毒载体对神经科学家来说是非常有用的工具,但高滴度和高纯度对于体内应用至关重要。在此,我们介绍一种通过在蔗糖阶梯梯度上进行超速离心来浓缩和纯化病毒原液的方案,以去除比病毒本身密度更高和更低的杂质,随后通过沉淀步骤去除蔗糖。最终的原液体积可浓缩至原来的1000倍,预期纯度高于之前发表的制备G缺失狂犬病病毒载体的方案。