Normally I use Minisart, 0.22-μm pore size filter for phage isolation from E. coli. But sometimes it is so difficult to filtrate the phage. It needs a lot of pressure and it hurts my hand. Is there any problem if I use 0.45-μm pore size filter? Or suggest any alternative way?
Try to centrifuge the phage/bacteria mixture before filtration. As the bacterial cells pellet, a very few of them remains in the supernatant in which phages are present. Then filtrate the supernatant through 0.22 μm syringe filter and it should be an easy job. According to Pierucci (1978), Dimensions of Escherichia coli at various growth rates: model for envelope growth, E. coli is large enough not to pass through 0.45 μm pore size filter. It's up to you, but I think that pelleting the bacterial cells is sufficient even while using 0.22 μm filter.
Hi Marian Varga, Thank for suggestion. In my current protocol I also use 13,300 rpm centrifuge before filtration. But still then filtration is very difficult. What happen if bacteria pass through the 0.45 μm filter? Then pellet not looks like glassy.
Hi, which the diameter of your filters? You should choose the proper size filters according to your supernatant volume. Also, you can let it through 0.45μm filters first, then0 .22μm ones. Otherwise, if you have a large volume, you may consider to use a membrane filter with vacuum pump.
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