Testing the ability of drugs to modulate platelet function is generally done using washed platelets (WP). I understand that most of the publications use WP to avoid any interference of the plasma compounds with the drug during the platelet function test. However, to find new platelet drugs wouldn't be better to use first platelet-rich plasma (which I consider to be more physiological)? In my opinion WP should be used to confirm if the effect of the drug is dependent of any plasma compound. Does it make sense?
I completely agree with your opinion, using PRP is the best way in general, however in some cases use of washed platelets is required. For example, if the incubation with drug has to be done at 37 C, then platelets in PRP get activated at this temp ex vivo as opposed to at RT, in this case washed platelets may be better choice.
Dear Tarun,
thank you very much for your feedback.
Currently I am using the following protocol for PRP:
1) 1/10 3.8% Sodium Citrate
2) 1100 rpm/ 200 G 15 min - PRP
3) 2500 rpm / 1100 G 15 min - PPP
I am adjusting the platelet concentration using the PPP.
Do you have any suggestion for this protocol?
Best,
The procedure used for preparation of PPP will of course depend on the platelet count you are aiming at in your plasma. It may be a good idea to stick to the CLSI guideline H21-A5. According to this guideline PPP should be prepared by centrifugation at 2000g for no less than 15 min, ensuring a platelet count below 10x10^9/L. However, a check of the platelet count is a good idea.
It depends of what you are looking at. Use of washed platelets allows more precise determination of mechanism of drug action. This is a better approach for early stages of investigation. For in vitro mechanistic studies washed platelets work well. However, use of PRP would make sense if you want to make sure that your drug (or drug candidate) is active in more physiological media, and its binding is not altered by plasma proteins.
Hi Augusto,
The the spinning speed is ok, use of 3.8% Trisodium citrate is good while duration depends on size of the tube, for upto 10 ml whole blood, 15 min is Ok. Duration may be increased upto 20 min for higher volume.
Hello, I read your discussion about PRP and WP. A month ago, I used PRP for study aggregation and everything went well. Now, I tried using WP and it did not work after induced with collagen. In the previous, I thought I make a mistake during CaCl2 preparation, but after I tried to make a new one, aggregation still did not work. If you all know or have some experiments like this please let us discuss about this. Thank you. Augusto Martins Lima Tarun Tyagi Yuliya Dobrydneva Johannes Jakobsen Sidelmann
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