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There has been a buzz about PRP globally due to its wide applications. However, the definition of PRP for each clinical indication is different in terms of volume, Platelet count and leukocyte addition etc.
The PRP is in fact a platelet concentrate (PC) with high number of platelets of the baseline count in liquid plasma.
There are largely 2 variants of PRP used for clinical indications
- Leukocyte poor PRP – No or less leukocytes
- Leukocyte Rich PRP – Contain Leukocytes
The major factor of PRP injection is the number of platelets used for specific indication rather than the plasma volume.
In certain indications, it is necessary to inject high number of platelets i.e muscle injuries , Gynecology etc, in small volume i.e 1-2ml. Therefore it is mandatory to concentrate Platelets to 5-8 times (X) of base line to get high number in low plasma volume.
This document contains the basic facts about PRP used for various clinical applications and acknowledges false claims or misapprehension about product.
DEPA classification of PRP preparations
| DEPA classification | Subgroup | Description |
|---|---|---|
| Dose of injected platelets | Very high High Medium Low |
>5 Billion injected platelets 3-5 Billion injected platelets 1-3 Billion injected platelets <1 Billion injected platelets |
| Efficiency of production | High device efficiency Medium device efficiency Low device efficiency Poor device efficiency |
Recovery rate in platelets >90% Recovery rate in platelets 70%-90% Recovery rate in platelets 30%-70% Recovery rate in platelets <30% |
| Purity of the PRP | Very pure PRP Pure PRP Heterogeneous PRP Whole-blood PRP |
Platelets in the PRP >90% Platelets in the PRP 70%-90% Platelets in the PRP 30%-70% Platelets in the PRP <30% |
| Activation process | Autologous thrombin Calcium chloride |
Modified from Magalon et al. BMJ Open Sport Exerc Med 2016;2:e000060 [11]. DEPA, dose of injected platelets, efficiency of production, purity of the PRP, activation of the PRP; PRP, platelet-rich plasma.
How to chose the Concentration devices
Better to avoid ACD Gel tubes (Specially for concentrated injections in ortho and gynic)
- Because gel tubes are meant for separation of plasma for Diagnostics testing only
- Separation gel is made up of Fumed silica and other micro plastic materials those are never approved for human injection ( Like ACD is approved for injection )
- Especially at high concentrated platelets injection at smaller sites may cause more inflammation due to silica materials and cause more damage than repair.
- Regular use of these devices for multiple small injection (Dermatologists) is not causing issue because small injections at multiples sites reducing the effect.
- Further ACD gel tubes cannot give 5-6X concentrated platelets with single step centrifugation
Devices to get Leukocyte rich plasma and leukocyte poor Plasma
- For Wounds, burns care and some Gynic conditions, better to collect leukocyte rich Platelets plasma as it is effective in healing wounds and induce local repair. So chose a device which can provide Leukocyte rich plasma. For orthopedic applications, better to inject Leukocyte poor plasma but at highly concentrated platelets (6-7X). So dual centrifugation.is required to concentrate the platelets.
How to calculate platelet count in PRP
Normal Blood Platelet Conc. 150-450 million / ml (i. e 150-450 103/ul). We need to have 8-10 billion as therapeutic dose for OA specially...
Ex: If Blood Report shows 250x103/ul that means 1 ml of Blood contains 250 million platelets. 20ml blood contains 250x20ml = 5000 million = 5 billion platelets 40 ml blood contains 250 x40ml= 10000 million = 10 billion platelets
- Knee OA treated with 10 billion platelets / Knee has given sustainable results for 3 year follow up. (Himanshu et al 2021)
- In Gynecology 5 billion platelets in endometrium yields good results. (Yogita et al, JBRA Assisted Reproduction 2022)
When we need to inject 10 Billion platelets, we need to concentrate all platelets from 40-50ml of blood into 5-6 ml of plasma.
Platelet Activation
Activation means breaking the platelets to release GF and Cytokines to Plasma liquid. However, external Activation of platelets is not required as native collagen of body is effective activator and releases growth factors to show the activity at injured site.
Further external activation shortens the activity of growth factors and degrades within 1 hr of injection. Intact platelets release growth factors slowly and provides long lasting sustainable repair.
Claims and facts about PRP preparation devices
Claiming of GFC (Growth factor concentrate) & Claiming Addition of growth factors to device:
- No growth factors need to add to device as our own PRP contains all required growth factors required for healing.
- Falsely claims the growth factors addition is not specified with the name of growth factors.
- Addition of Growth factors is not Possible without permission of CDSCO as Growth factors are Biopharmaceutical products
Cryo freezing and thawing increases platelet growth factors
- Cryo freezing and thawing breaks down platelets and release GF and Cytokines into plasma solution and it is considered as more growth factors. After injection, platelets get activated by native collagen and local release of growth factors are better.
- All released contents outside are degraded immediately in the body within 70 Min.
Claiming ACD is better anticoagulant:
- Heparin Maintains better platelet quality for longer periods. ACD is cheaper alternative and stable at room temperatures. So ACD is easy to use but if available heparin maintains good quality platelets
Devices to get Leukocyte rich plasma and leukocyte poor Plasma
- Platelets in blood circulated at 37°C. Even in environment also they are suspended in Plasma and does not activate / breakdown in 20 -30 min. of process time and 2-3 hrs of storage.
- There are more than 100 PRP devices approved by FDA in USA and none of them need to process at lower temperatures.
Claiming that single centrifugation sufficient for 6-7X concentration
- Definitely Need dual centrifugation step for 6-7X concentrated platelets. 1st centrifugation yields 2X concentration and 2nd centrifugation concentrates 2X platelet plasma to 6-7X to the base line blood.
- All most all high concentrate devices follow dual centrifugation protocol in order to achieve high concentration
PRP preparation method by centrifugation
Dual centrifugation is an optimal method for concentration of platelets from Blood. The 1st centrifugation step separates Platelets and plasma (2X of platelets) from RBC and WBC. Then Plasma containing platelets has to be centrifuged again at higher speed to concentrate platelets into small volume of plasma for further injection.
Platelet concentration Devices Variants
There are 2 varieties of devices available in the market.
concentrate devices - Yields 2X concentration with single centrifugation
- Small gel tubes with ACD anticoagulant
- Laboratory plain tubes (Need to add anticoagulant)
High concentrate devices - 6-8X concentration with dual centrifugation
- Dual centrifugation based devices for yielding high concentrated low volume plasma
Anticoagulation Myth and Cooling temperature
Anticoagulant – ACD and Heparin are Equally good and no effect on Platelet quality
No effect of temperature has been observed on platelet quality in 20-30 min of processing since they are suspended plasma. Platelets get activated / de-granulated when they are exposed to Citrate chelators, Collagen and Very low temperatures.
Sterility
Should be Sterile for infusion
Processing Blood Volume
Clinical Indication dependent
Required Platelet Number
Clinical Indication dependent
Concentration
Device should be able to concentrate up to 6-8 times of the Base Level with Dual centrifugation.
Biocompatible and fit for Infusion
All used materials shall be Biocompatible including ACD/ Heparin, Plastics, Silicone etc (ACD gel is not proven to be safe for infusion)
Why to concentrate platelets for injection
Any degeneration / damage occurs due to high inflammation or short supply of growth factors (for repair) in the particular microenvironment area.
High concentration of platelets at Microenvironment normalizes inflammation (by cytokines) and induces growth by supplementing growth factors.
Certain cases, more than 3-5ml cannot be injected to the injury site and thus supplementation of high concentration of platelets (10 billion platelets in 4ml) in low volumes is required.
PRP types
Leukocyte Rich PRP: Containing Mononuclear cells (regenerative Monocytes also) along with Platelets. This product works better in Wound healing, High grade OA and Certain gynic conditions.
Leukocyte poor PRP: Contains only platelets in plasma (No MNC or WBC etc)
Certain cases, more than 3-5ml cannot be injected to the injury site and thus supplementation of high concentration of platelets (10 billion platelets in 4ml) in low volumes is required.
About RBC contamination in platelets
Higher RBC concentration may cause inflammation at the site of Injury. But lower concentration of RBC with WBC will be better product for injury repair.