In the exosome isolation protocol by Thery et al it is recommended to prepare 30 % sucrose cushion using Tris and D2O. I would like to know if the sucrose cushion can be made in PBS instead of Tris and D2O. Will it affect the purity of vesicles?
The hydrogen atoms in heavy water (D2O) contain a neutron, making those heavy water hydrogen atoms about twice as heavy as normal hydrogen. The additional neutron in each hydrogen atom does not change the volume of the water noticeably; however, the increased molecular weight does make the water slightly more dense. In its pure form, it has a density about 11% greater than water. The isolation of exosome is dependent on the unique density of exosomes and the buffer system. The density of D2O is 1.1056, if you can make a solution with the D2O density, it is possible to replace D2O from other solution. I am afraid that the density of PBS may different from D2O, if they have similar density, I think you can use PBS to replace D2O.
Yes. You can dissolve sucrose with PBS. I have done it a lot without any problem. In addition, D2O is relatively expensive.
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