Lönnbro Per, Nordenfelt Pontus, Tapper Hans
Division of Infection Medicine, Department of Clinical Sciences, Lund University, SE-221 84 Lund, Sweden.
BMC Cell Biol. 2008 Jun 27;9:35. doi: 10.1186/1471-2121-9-35.
There is a growing awareness of the importance of intracellular events in determining the outcome of infectious disease. To improve the understanding of such events, like phagosome maturation, we set out to develop a versatile technique for phagosome isolation that is rapid and widely applicable to different pathogens.
We developed two different protocols to isolate phagosomes containing dead or live bacteria modified with small magnetic particles, in conjunction with a synchronized phagocytosis protocol and nitrogen cavitation. For dead bacteria, we performed analysis of the phagosome samples by microscopy and immunoblot, and demonstrated the appearance of maturation markers on isolated phagosomes.
We have presented detailed protocols for phagosome isolation, which can be adapted for use with different cell types and prey. The versatility and simplicity of the approach allow better control of phagosome isolation, the parameters of which are critical in studies of host-bacteria interaction and phagosome maturation.
人们越来越意识到细胞内事件在决定传染病结局中的重要性。为了更好地理解诸如吞噬体成熟等事件，我们着手开发一种通用的吞噬体分离技术，该技术快速且广泛适用于不同病原体。
我们开发了两种不同的方案，通过结合同步吞噬作用方案和氮空化作用，利用小磁性颗粒分离含有死菌或活菌的吞噬体。对于死菌，我们通过显微镜和免疫印迹对吞噬体样本进行分析，并证明了分离出的吞噬体上出现了成熟标记物。
我们展示了详细的吞噬体分离方案，该方案可适用于不同的细胞类型和猎物。该方法的通用性和简单性使得对吞噬体分离的控制更好，而其参数在宿主 - 细菌相互作用和吞噬体成熟的研究中至关重要。
