Pelleting in a centrifuge will occur only if the substance has higher density than the medium. Assuming this is true, pelleting may take longer than you have tried, or require higher g force to occur in a reasonable amount of time. You could try using an ultracentrifuge. Alternatively you could remove excess protein and ions by dialysis or ultrafiltration. How big are the particles and what is the molecular weight of the protein?

Hello A.B Shapiro,

I am using BSA stabilised gold nanoclusters (~2 nm size) and trying to avoid the dialysis as it induces some changes in the spectral characteristics after dialysis which I want to avoid. I want to obtain nanoclusters in the form of precipitate  for some  further work and at the same time want to obtain pure gold nanoclusters free of ions. So I am concentrating on centrifugation technique. Please suggest .

What conditions of centrifugation have you tried? If the particles won't pellet at 100,000 x g in the ultracentrifuge, I would suspect the reason is that they are suspended in a dense medium. If so, diluting the medium with a buffer to reduce its density may allow the particles to pellet. However, the BSA may also pellet. Have you considered density gradient ultracentrifugation?

Hello Sir,

I have tried centrifuging at 25000 rpm but not density gradient centrifugation. As I want to precipitate nanoclusters, I have tried both in water as well as buffer. After running centrifugation for 30 min. at 25000 k, 4 degree celcius, very little amount of particles (i.e BSA stabilised gold nanoclusters) pellets down and major amount remains in the supernatant. Please suggest.

Have you tried precipitating the clusters with ammonium sulfate? It's often used to precipitate proteins in a reversible, non-denaturing way. The required concentration varies, but is usually several Molar (e.g. 3 M).

Hi. Anushree.. Even I am facing the same problem. I am also trying to seperate GNCs from unreacted gold and BSA.

But in this paper you can find some way to seperate clusters but in a complicated way. Try see whether u can get this.

http://pubs.rsc.org/en/content/articlehtml/2009/cc/b916919a