Preparing platelet rich plasma (PRP) from whole blood requires centrifugation to isolate and concentrate platelets. Centrifugation harnesses the principle of centrifugal force to speed up the natural sedimentation process. Practitioners who administer PRP with a significant concentration of viable platelets into damaged tissues see increased tissue regeneration and better outcomes. This article discusses the principles of effective centrifugation as it applies to PRP preparation

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PRP: How to Concentrate Platelets

Blood is composed of red blood cells, white blood cells, plasma and platelets. If a vial of blood were left out on a table, the blood would eventually stratify into three distinct layers with the heaviest components sinking to the bottom and the lightest rising to the top. Eventually three distinct layers would become visible; red blood cells (RBCs) at the base, white blood cells and platelets in a thin layer above the RBCs (also known as the ‘buffy coat’), and finally platelet poor plasma on top.

Centrifugation accelerates this innate sedimentation process and allows practitioners to isolate and re-inject platelets in a timely manner. Under centrifugation, the particles in suspension experience radial force which moves them away from the axis of rotation. Particles of a higher density travel at a faster rate and become separated out from less dense and smaller particles.

Best PRP Centrifuge Rates and Protocols

researcher analyzing prp centrifuge

Calculating the exact speed, force, and length of time required to stratify blood requires a deep understanding of physics. Many factors influence how the centrifugation process unfolds, including the width, length and weight of the PRP kit, as well as the speed and force exerted by the centrifuge. Additionally, centrifugal force increases with the distance from the axis of rotation. Thus, given the same speed, a larger centrifuge will exert a larger force compared to a smaller centrifuge.

Relative centrifugal force is an expression of g-force; the earth’s gravitational pull. When calculating g-force, remember the g-force acting on the particles is exponential to the speed of rotation. This exponential is expressed in revolutions per minute (rpm). The exponential is especially important when considering speed of centrifugation; for example, doubling the speed quadruples the force.

Commercially available centrifuges are designed to maximize sedimentation and isolate platelets when used with their respective PRP kit. Variation in weight and size of commercially available PRP kits necessitates a distinct centrifuge for each kit. The manufacturer establishes protocols to optimize the sedimentation process based on the exact kit size.

Centrifugation and Temperature

The most important factors to consider in centrifugation are time, speed, and temperature, but none of the commercially available PRP centrifuges appear to control for temperature. Is this industry oversight leading to disparate results in orthobiologics?

researchers determine best PRP centrifuge design

Temperature is an essential variable to consider whenever working with biological material. High speeds create friction and can heat the sample during centrifugation. Controlling for temperature becomes even more important if centrifugation takes place outside of a vacuum. Centrifugation outside of a vacuum creates additional drag and temperature increase. Research biologists traditionally cool centrifuges to offset the increase in temperature from friction.

If the sample is heated during centrifugation, platelets may activate early. Additionally, heat could damage platelets and lower viability. Cooling the centrifuge slows biological processes and could help to retain platelet viability. This well known biological phenomenon has been acknowledged by researchers as it pertains to enhancing platelet activation and the release of alpha granules¹, but rarely as a detriment to platelet viability during centrifugation. The literature reveals one single-author review recommending centrifuges be cooled between 12-16 degrees C (53.6-60.8 F) to preserve platelet viability.²

More research is necessary to better understand the role of temperature in PRP preparation. Replicating results when working with biological materials depends on precisely controlling variables, and conflicting results in the field of orthobiologics could be linked to varying sample temperatures. The best PRP centrifuge and centrifugation protocols depend on the exact specifications of the PRP kit. Ideal centrifugation rate and duration is calculated based on the mass and dimensions of the PRP kit. Researchers and clinicians would benefit from experimentation designed to assess temperature increase during centrifugation as related to platelet viability.

References

  1. Egidi, M. G., D’Alessandro, A., Mandarello, G. & Zolla, L. Troubleshooting in platelet storage temperature and new perspectives through proteomics. Blood Transfus 8(Suppl 3), s73–81 (2010).[ncbi]
  2. Dhurat R, Sukesh M. Principles and Methods of Preparation of Platelet-Rich Plasma: A Review and Author’s Perspective. Journal of Cutaneous and Aesthetic Surgery. 2014;7(4):189-197. doi:10.4103/0974-2077.150734.[ncbi]

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