Are there techniques other than SEM to confirm bacterial outer membrane vesicles. Actually, our SEM is out of order for some time, so, I want to confirm them by some other techniques.
Hello, easy way is SIM. Other way is using some assay for respiratory chain enzymes or other integral IM protein but It is hard to distinguish the membrane vesicles or Plebs from membrane vesicles made from lysed cells. However, the best approach is to determine the percentage of the cells that lysed by assaying the supernatant for the presence of a constitutive intracellular enzyme - like glucose-6-Phosphate dehydrogenase. This way you can determine the percentage of lysed cells and their potential contribution to your total isolated membrane vesicles. Of course you need to determine the total activity of G6PD in a given volume of the culture (i.e., Cells + supernatant).
Therefore, % of lyses can be calculated by the way (Total G6PD activity per ml of culture / G6PD activity per ml of supernatant) 100.
Also several quantitative methods exists, see attached.
Hello, easy way is SIM. Other way is using some assay for respiratory chain enzymes or other integral IM protein but It is hard to distinguish the membrane vesicles or Plebs from membrane vesicles made from lysed cells. However, the best approach is to determine the percentage of the cells that lysed by assaying the supernatant for the presence of a constitutive intracellular enzyme - like glucose-6-Phosphate dehydrogenase. This way you can determine the percentage of lysed cells and their potential contribution to your total isolated membrane vesicles. Of course you need to determine the total activity of G6PD in a given volume of the culture (i.e., Cells + supernatant).
Therefore, % of lyses can be calculated by the way (Total G6PD activity per ml of culture / G6PD activity per ml of supernatant) 100.
Also several quantitative methods exists, see attached.