PhD student Polly came into the lab on Monday morning to find that her purified protein had started to aggregate after a weekend in the fridge. The profile she got from her Dynamic Light Scattering (DLS) measurement clearly showed two peaks, with the second in the micron range. Her supervisor suggested she tried Nanoparticle Tracking Analysis (NTA) to get more information on the different sized aggregate species in her sample.
Monday – 1pm
Polly: Hi Professor. I’ve got the NTA data you suggested. You were right – it was pretty quick and easy to get the data. I just checked with Fred that the image looked good and took five 60 second video captures. It was cool to see the particles move!
Here’s what the profile looks like by NTA:
I’ve a range of sizes from 35 nm to 500 nm and there are definitely a few peaks in between.
Prof: So what do you think this is telling you about your protein?
Polly: Well ……um…I think that I have aggregates that are probably bigger than a trimer in my sample and some larger order oligomers?
Prof: So with these two techniques do you have a fuller picture of what’s in your sample?
Polly: Yes! By DLS I see I have some monomer and from the width of that peak there is also presence of oligomers, it also shows me I have some larger aggregates. The NTA gives greatly increased resolution on the smaller to larger aggregates.
Prof: So if the aggregation is the outcome, what’s going on in the sample?
Polly: I guess some of the protein has lost its tertiary structure or denatured and this has caused the aggregates.
Prof: So what do we do next?
Check back later this week to find out….
Missed Part 1?: