Dear Trisha Bansal,

Yes. It can be done.

Plasma Protein Binding Assay

Understand the distribution potential of your compound using our plasma protein binding assay.

Plasma protein binding is one of Cyprotex's in vitro ADME screening services. Cyprotex deliver consistent, high quality data with cost-efficiency that comes from a highly automated approach.

Determination of fraction unbound in plasma using equilibrium dialysis

The extent of binding to plasma influences the way in which a drug distributes into tissues in the body.

Etensive plasma protein binding also limits the amount of free compound available to access sites of action in the cell, and metabolism and elimination may be slower.

Equilibrium dialysis is the most widely accepted method for assessing plasma protein binding as non specific binding effects are minimised compared with other methods such as ultrafiltration.

Cyprotex's Plasma Protein Binding assay is performed using an equilibrium dialysis method and delivers a value of fraction of compound unbound to proteins (fu).

There is a choice of three methods for assessing plasma protein binding using three different percentages of plasma to provide flexibility depending on budget and compound characteristics.

Equilibrium dialysis is the preferred method to determine the free drug fraction, because it is less susceptible to experimental artifacts.

Kariv I, Cao H and Oldenburg KR. (2001) J Pharm Sci 90(5); 580-587

For the protocol details, please use the following link:

http://www.cyprotex.com/admepk/protein_binding/plasma-protein-binding

Hoping this will be helpful,

Rafik

thanks, But I would need an interaction in a cell free system between the drug and the protein which I obtained via in vitro transcription coupled translation and subsequent purification.

Is your protein a membrane-bound, or a cytoplasmic free-floating protein, or a membrane-associated protein?  This is vital to creating a similar in-vitro micro-environment to simulate reality, for your protein and drug.  If your protein is cytoplasmic or nuclear, and your drug doesn't diffuse through membranes, you will never get an interaction in the body similar to an interaction that you may see in-vitro.

membrane bound proteins:  try using microsomes

cytoplasmic: use any plasma or protein appropriate media

membrane-associated protein:  this is tough, because you have to also introduce the membrane-bound protein in microsomes or vesicles, to then bind your new protein.

The main point of this is, that you can put any drug and any protein in a plate or microwell together. The interaction you see may or may not be an accurate picture of reality.

Inclusion of liver microsomes can give an added bonus of watching how your drug is metabolized at the same time as you are watching for the drug to interact with your protein.