Hi,
I have been trying to induce platelets aggregation using different cancer cell lines but it did not work although this is reported many times in literature.
I tried the following cancer cells (MCF7, MDA 231, PC3, PC3M, BXPC3, CAPAN-2 and HPAF2), Also I tried different dissociation media to detach cells (Trypsin, EDTA, EGTA, accutase and scratching)
I used platelets prepared by different methods (Human washed platelets with ACD, PGI2 or PGE1), (PRP with either 3.2% or 3.8% citrate)
Passage of cancer cells from 25 to 60
Platelets from healthy volunteers who did not take any medications two weeks prior to bleeding.
It has been tried by more than one student and it is not working as well.
Wondering if anyone has done this before?
Thanks
Dear Omar,
Some suggestions, consult the authors they are here on RG:
Article [Changes of platelet aggregation in cancer patients]
Article Baicalein inhibits agonist- and tumor cell-induced platelet ...
Article Von Willebrand factor fibers promote cancer-associated plate...
Article The Potential of Enzastaurin to Enhance Platelet Aggregation...
Omar,
What volume of PRP and what volume of cancer cells you are taking. Perhaps you may have to vary both the volumes and try again. I would add more of platelets and less of cancer cells.
Omer
Hi Omer, thanks for your reply.
I am using 90 µL of 300 ×10e6/ml of washed human platelets, + 180 ul Tyrode then after 1 min stirring in aggregometer, I add 30 µL of 1 ×10e6/ml of cancer cells.
For PRP, the same except the platelets concentration is less (300 ×10e5/ml)
Hi Maria,
If you are looking at cancer cell-induced aggregation of human WASHED platelets, no, It never worked consistently with me although trying different platelets isolation protocols/anticoagulants, and cancer cells prep protocols (e.g. different cell detachment reagents). However, I found that it works consistently if 1-using PRP instead of washed platelets, 2- using platelets without washing steps (resuspending platelets pellet directly after removing plasma), 3- the addition of a very low amount of plasma to the aggregation reaction (0.5% plasma makes it work consistently).
Long story short: try adding 0.5-1% plasma to your aggregation reaction, if it did not work this means the cell line you are looking at is not able to induce platelets aggregation (maybe it can induce platelets activation, secretion or spreading but not aggregation)
Thanks
Hi Omar!
Thank you very much for the reply. What you tell me is that the assay works only when some of plasma proteins are present (by adding them at 0.5%, using PRP or do not fully washing).
But... How this can be possible?! TCIPA with washed platelets is EXTENSIVELY reported in the literature. When you say that it was not consistent in your hands, means that you got some positive result? In my case worked ONCE! but never again.
Thank you again for your answer!
Best wishes.
- Badges
- Science topic
- Similar topics
- Computer Communications (Networks)
- Simulators