I mixed protein with silver nanoparticle in different protein concentration. We showed increase of protein concentration caused decrease of size measured by DLS.
How large was the change in size? How are you subtracting the protein contribution to the dynamic structure factor? QELSS measures a light scattering weighted average; a rapidly decaying component in S(q,t) due to the protein could create this illusion. How large is the protein concentration? Quasielastic scattering measures S(q,t), not size; how are converting from one to the other? What are the diffusion coefficients of teh protein and the silver nanoparticle? I am now asking questions rather than giving answers, because there are a whole bunch of ways to get this effect.
Despite not being expert in DLS and considering the commments from Dr. Phillies, I would like to suggest if the protein is not dissolving your nanoparticles. I explain myself, many proteins do have sulfur moieties that usually interact very well with metals, that is how the proteins use to anchor to Au for example. Could it be the case that sulfur as in cysteine of methionine moieties is in fact attaching with the nanoparticles thus reducing its size? Or if I understood well, the whole measured size is decreasing, could the increased concentration of protein be coupled with some degree of denaturalization, deagglomeration initiated by the nanoparticles... Just ideas in the spirit of some observations we had in semiconductor nanoparticles attached with thiols
thank you dear friends
Dr. phillies
we saw more than 40nm change. we really compare the DLS of stock nanoparticle and the DLS of nanoparticle interacted with proteins in same situation. we did not subtracting anythings!!! i am a biotechnologist and i do not the meaning oh QELSS. dose it necessary? the protein concentration is based on the percentage of acetylcholinestrase(my protein) than nanoparticle. in first percentage, we had nanoparticle with 42nm early measurement and 20% enzyme than nanoparticle.the size of nanoparticle after treat had been about 142nm. in other test, the size of nanoparticle in early step was 42nm and 50% enzyme than nanoparticle. after treatment we observed the DLS measurement of nanoparticles about 109nm.
i do not know what the Quasielastic scattering measures S(q,t) is?
how can we measure the diffusion coefficients of the protein and the silver nanoparticle?
Dr. Caballero-Briones
as i said my protein is a recombinant acetylcholinestrase and it has more than 5 cysteine amino acid. if the size decrement is the consequence of deagglomeration or... how can we prove this phenomenon?
Protein activity perhaps? X-ray diffraction to see if nanoparticles have dissolved? Scanning electron microscpoy to see changes in morphoolgy that would suggest deaglomeration, denaturalization?
my activity was reduced significantly. i am comparing the XRD from first and treated nanoparticles. is it useful? as my nanoparticle size is between 20nm to 120 nm i can not see the nanoparticle with SEM in good resolution.
The additional information you provided seems to indicate size increase after protein addition, not decrease. Or maybe I am not reading it correctly.
In any case, I agree with the previous responses. I think high resolution SEM or maybe TEM can really help. It seems you need to visualize what is going on. You can also measure the charge (zeta potential) on your zetasizer and see how it changes(?) between nanoparticle only, protein only, and the mixed systems of nanoparticle and protein.
Have you tried to measure hydrodynamic size over a period of time? Say an hour?
Dr. Adeleye
yes you right the nano particle size have increased after incubated with protein. but when we increased the protein concentration, the nanoparticle size has decreased. yes we did. we measured the size in the two times, after 30 and 200 minutes. at second time, the size of nanoparticle has increased.
At times, large aggregates may settle out of suspension leaving the smaller aggregates, which are detected by DLS and may be interpreted as disaggregation (decrease in size). It might be helpful to also keep an eye on average particle countrate (I assume you use a zetasizer nano). But again, no single technique can provide conclusive explanations. You may also have to investigate what is going on using surface charge (how it changes with protein addition), electron microscopy, and an xray technique. Best wishes.
thank you Dr. Adeley
the "kcps" factor measured for pristine nanoparticle and incubated nanoparticle showed for the first DLS this factor was about 4000, while this factor for second DLS was about 2000. in this situation is it possible the large aggregates may settle out of suspension leaving the smaller aggregates?
I think it is possible. But to be sure you can set up a size measurement that will run continuously for a long time (e.g. 200 min). That way you can continuously monitor size and kcps change with time. If you need help with setting up this kind of measurements let me know.
i am really appreciating you. what should i do? i want to carry out a continuously running.
Hi. You can start with this SOP attached. You will need to manually make this SOP in your zetasizer using the "New SOP" button. Let me know if there are other parameters you are unsure of when making your SOP but most will be the same you use for size measurement. This one will run size continously for an hour but you can adjust it as needed.
Key control experiment. Do QELSS from the protein solution without the particles being present. Protein scattering will give you a fast earlier decay in your spectrum that may confuse your data interpretation, even if the intensity is not that large.
Second issue: What is your background ionic strength? If you are at very low salt concentration, all sorts of interesting things can happen.