Various viral infections still pose a great threat to human and animal health. A wide range of viral species are surrounded by a lipid shell, the envelope, containing viral and cellular proteins.
The group of enveloped viruses contains species of medical and biotechnical importance such as members of the families Retro- (HIV), Filo- (Ebola), Flavi- (West Nile, Zika, Dengue) and Orthomyxoviridae (Influenza) to name but a few. The contribution of protein elements of the envelope to viral reproduction and pathology is widely studied. We are more interested in the contribution of the lipid elements especially in the behavior of viral (lipid) envelopes after biophysical and biochemical stimuli. Furthermore we try and use this information in the modification of enveloped virus surfaces (i.e. Retro/Lentiviruses) for applications in gene therapy and vaccine development. One important tool we try to develop is multi-parametric single particle detection systems. The latter point is most important for acquiring reliable data on the homogeneity of the samples, answering questions such as: how strongly single viral particles differ from each other regarding morphological, biophysical or biochemical properties? Are different populations recognizable (i.e. contamination by other vesicles/particles or subpopulation of virus particles)? – TRPS provides an in-road into the topic by measuring size and charge of individual particles – key parameters for describing viral populations. Additionally, Since EVs often contaminate viral preparations, for our purposes a negative selection for EV would be most important (i.e. to remove EV from viral samples). However, a quick and reliable, cost-effective purification of virus as provided by qEV columns is highly welcome for our research.
Very. Somehow comparable to quality control in manufacturing processes: cells produce a large number of particles (not exclusively viral) and in order to do research on them we need to know how closely (or not) they resemble one another.
Mostly speed and the simple (and gentle) procedure reducing time and chemical stress. Both means we can keep the particles functional (and/or infectious) as much as possible.
I’d be tempted to say not a lot. As it’s emerging, the interplay between different particle types seems to be pretty close. Mostly the nature of both, the virus and the infected cell will decide. However, drawing clear lines can be very difficult.
For the first part, in some cases, I’m guessing yes. However, to my knowledge this has not been tried yet. Choosing the right pair would help. For the second part, not with molecular painting (currently, it doesn’t discriminate between different membrane types. However you may use it to discriminate membrane bound from protein capsid only particles). In any case, the quality of the marker(s) would be the critical issue.
Yes, so far we’re using quite complex biophysical techniques to assess bulk binding properties. It becomes most useful (and interesting) when we can link this information to virus functions.
The first part of the question is difficult to answer. Since data systematically comparing charge data between vesicles are lacking. So the probably not very helpful answer would be as high resolution as possible. For the second part: while membranes may not differ too much in charge, the dominant protein species on the vesicles may very well (as a result of their isoelectric points). I’m thinking going there might me most useful.
Quality control is for sure on the agenda. A quick way to measure infectious to total particles. A good substitute (set of) parameter(s) for infectivity (which requires per definition cumbersome cell culture work) would also be very welcome.
