What is Platelet-Rich Plasma (PRP)? PRP is platelet concentrate made of autologous blood in an adequate volume of plasma. After three decades of Platelet-Rich Plasma use in various medical fields, there is yet to be a universally-accepted definition for what PRP is. Today there are currently number of PRP Practitioners in the world that wrongfully believe that as long as an «Enrichment» sequence is performed, PRP, at any level of concentration, compared to the baseline, is attained…

1

2

What is Effective PRP Treatment?The known range for fragile platelet count in human blood is anywhere between 150,000 and 450,000 per microliter of Whole Blood (with an average count of 200,000/µl). Based on recorded effective bone and soft tissue healing enhancement treatments, Medical Professionals conclude that a minimum of 1,000,000 platelet-count per microliter of PRP must be obtained in order to achieve Effective PRP. In other words, with 1,000,000 platelet count, one can achieve three to ten times the baseline. Any concentration level below three times will not be considered effective PRP; and further studies indicate that any level of platelet concentration above ten times the baseline does not offer any enhanced healing.

That said, there are three factors that influence the effectiveness of any PRP treatment:

A.High Concentration of PlateletsB.Healthiness of fragile PlateletsC.The right volume of PRP (treatment-specific) Why Healthy platelets?1,000,000 fragile platelet-count per 1 microliter of PRP attains concentration levels of three to five times the baseline. In other words, to have an effective PRP treatment, the healthiness of the platelets must be well protected, because platelets are super fragile and easily become degranulated by the slightest trauma, releasing their Growth Factors before the right time, making the treatment ineffective. Therefore, one must make sure that the Growth Factors are not released until adequate platelet count is achieved.

3

Ho w Fr ag ile Platelets Become Degranulated at the Wrong Time?Fragile platelet survival rate against any type of stress is ten to fifteen times less than the stress level that red blood cells can sustain.There are many preventable factors that destroy fragile platelets during the PRP preparation cycle, including but

4

not limited to, needle gauge used for blood collection, vacuum pressure in blood collection devices, transfer containers types and material, centrifugation speed, force, and duration, centrifuge machine type, and number of centrifugation cycles…

• Needle GaugeBlood-flow inside the needle is not laminar, it is turbulent. Therefore, using small gauge, long, and multiple needles for blood collection are one of the causes of fragile platelet's breakup or degranulation.

In vacutainer blood collection devices, there are two needles. One for blood collection, and the other for collected blood's transfer into the transfer container. In such devices, the needle gauge for blood collection

is usually smaller than the needle used for transfer into transfer container. Thus, doubling the risk of fragile platelets' degranulation.

• Vacuum PressureThe vacuum (PULL) pressure inside the blood collection devices such as syringe or vacutainer, where collected blood is forcefully drawn inside, can be another cause of fragile platelets' damage, caused by the crashing of the blood cells into each other and against the hard interior surfaces of the collection devices.In addition, there are two other types of Positive (PUSH) Pressures that fragile platelets must defy for their survival: A) the pressure caused by the force intended to push the blood inside the blood collection container out into the transfer container, which attaches and squeezes all the cells together that can cause more fragile platelets' degranulation, and B) the pressure that is present,

5

when the collected blood is forced through the needle into the transfer container, which in turn forces the crash of additional number of outlived fragile platelets into other blood cells and the hard interior surfaces of the transfer container.

• Blood Container Hard Surfaces

As per American Association of Blood Banks' recommendation, in order to protect all the blood cells, the blood collection containers must have a soft surface.• When using vacutainer

tubes for blood collection, the risk of damaging the blood cells, including the fragile platelets, is increased, because

the vacuum pressure inside the tube, jets the collected blood cells against the tube's hard surface.

• During the centrifugation spin, all blood cells inside the container are pushed against the back wall of the container and the first layer of the cells crash against the container surface.

• In addition, since each blood cell type has a different density, then while moving inside the container during the spin cycle, it will have a turbulent effect on the other lighter cells, which in turn crashes those cells against the container›s wall.

• In some kits, the angular shape of the PRP preparation container (e.g. hourglass type), will cause damage due to the centrifugal force and also the turbulent effect that is created due to the device's reduced (bottle-

6

neck) diameter• The presence of any

other object inside the PRP kits will also create further turbulence, causing more fragile platelets' degranulation.

• Centrifuge Machine Type

Many of the current centrifuge machine types that are used for PRP preparation are «fixed» rotor-head type.

That is, during the separation cycle, containers are arranged inside the machine carriages at a fixed angle well below 180°. In other words, since the centrifugal force is parallel to the horizon at a 180°, the blood cells are propelled to the container's back wall. The consequence of this crash is the rupture and degranulation of fragile platelets. Therefore, using a fixed rotor-head centrifuge machine cannot fully protect the fragile platelets from degranulation.

7

. Number of Centrifugation CyclesUnfortunately, many Physician and PRP Practitioners around the world are led to believe by their devices' instruction that a single centrifugation cycle can produce the proper concentration level of platelets that are required in an effective PRP treatment.

The truth of the matter is, that although «enrichment» is possible in all PRP preparation devices, fact remains that at the end of a preparation cycle, where a single centrifugation is applied, adequate concentration level of platelets from the baseline is not obtained. Effective PRP treatment will happen when minimum three times to ten times the concentration level from the baseline is attained.

8

• To demonstrate this point, let's say that 20ml of blood is collected from a donor. Obviously, then at the completion of the single centrifugation cycle, the original 20ml of Whole Blood is now separated into approximately 10ml of Red & White blood cells and 10ml of Plasma-containing platelets (assuming donor's Hct is 50).

• Although it is impossible for every single fragile platelet to be separated from the Whole Blood after the completion of the first centrifugation cycle (because they are suppressed by the larger RBCs), then hypothetically speaking, if every single fragile platelet has survived the journey and are now accumulated inside the plasma, then the maximum concentration level of the fragile platelets in the 10ml of plasma is only twice the baseline! Remind you,

effective PRP treatment is when minimum three to ten times concentration of fragile platelets from the baseline is achieved!

• Risk of InfectionIf not cautious when performing a PRP treatment, considering the treatment type and specific risk factors involved with each type of treatment, the risk of infection is varied, and if not controlled can be very high, seriously endangering the wellbeing of the patient.Note here that the risk of infection increases, each time the collected blood is transferred into another container (for example: from blood collection syringe or vacutainer into tubes, for centrifugation cycle).EPRP® PRP Preparation Device is a CLOSED system, which reduces the risk of cross-contamination and any possible infection risk, to none.

9

Right Volume for the Right Treatment (Simple Mathematics)

Have you ever asked yourself what is the right volume of PRP for any specific treatment; and how much blood must be collected from your patient in order to obtain that required volume of PRP?

Well, it is simple mathematics. Here is an example:• Treatment Type: Hair

Resuscitation• Treatment Area on the Scalp:

10cm X 15cm• Number of PRP injection

points: 150• Minimum PRP volume per

injection: 0.1ml• Total Volume of PRP

required: 15ml (150 X 0.1ml)• If for Effective treatment

minimum 1,000,000 platelets/µl (microliter) of PRP is required, then 15ml PRP X 1,000,000,000 platelets/ml (milliliter) = 15,000,000,000 platelets (1ml = 1,000µl)

• If on average, there are 200,000,000 platelets in every milliliter of Whole Blood, then 15,000,000,000 ÷ 200,000,000 = 75ml Whole Blood

• Conclusion: To obtain 15ml of PRP for effective hair resuscitation procedure, then minimum 75ml blood-collection is required!

It is reiterated here that for successful PRP treatments, one must achieve ALL three attributes: A) High Concentration of platelets, B) Healthy, intact platelets, and C) Adequate volume of PRP.If any of the mentioned attributes are missing, then most likely your PRP treatment will NOT be effective.

10

• Why EPRP® P r e p a r a t i o n Device?

• Platelets are a blood cell; and PRP is a Blood Product! That is why EPRP® PRP Preparation Device is designed based on the same standards and functionalities that are considered in the production of blood collection bags; whereby every single blood cell can be effectively separated, while keeping most of the

cells intact and healthy.• EPRP® PRP Preparation

Device concept takes into account the complex requirements of Blood Banks, while at the same time making it very user-friendly for the physicians to achieve excellent separation of blood cells in their own private clinics.

• EPRP® PRP Preparation Device is designed to attain all three attributes of Effective PRP Treatment: Healthy platelets, High platelet Concentrates, and Accurate Volume of PRP.

• How Does EPRP® Deliver Healthy Fragile platelets?

By following National and International Scientific Organizations, Associations and Societies' Guidelines (e.g. ISBT, AABB, WHO…) EPRP® PRP Preparation Device utilizes several methods to keep the fragile platelets healthy and undamaged:

• Needle Gauge:For EPRP® PRP Preparation Device the smallest recommended Needle Gauge/Size is 19. That is, larger the needle gauge, less fragile platelets are degranulated during blood collection phase.

• Soft mini-blood bagsEPRP® PRP Preparation Device's soft material maximizes the survival rate of fragile platelets at all times, during blood collection and separation cycles, because fragile platelets no longer crash into any hard surfaces.In fact, AABB (American Association of Blood Banks) strongly recommends that all blood containers must be made of soft material in order to maximize the survival rate of all blood cells.

• No Vacuum Pressure PresentEPRP® PRP Preparation Device›s functionality is similar to blood collection procedures and guidelines being applied in every blood bank around the world; where patients› blood is collected using natural venous pressure and actual force of gravity. There is no negative or vacuum pressure to damage the fragile platelets.

12

• How Does EPRP® Deliver High Platelet Concentrates?

As every physician and practitioner may have had to experience how to control massive bleeding in a patient, sometimes during their careers, they would tell you that they had to inject pure, highly concentrated platelets into the patient.For many decades, national blood banks are producing high concentration of pure platelets under industrial conditions; but today, EPRP® PRP Preparation Device's user-friendly design helps physicians to obtain the same results, in their own private clinics. The only difference is the level of concentration. The recommended platelet concentration levels for an effective PRP treatment is between three times to ten times the baseline. Anything below three times the baseline cannot be considered PRP; or anything above ten times the base line offers enhanced healing.EPRP® PRP Preparation Device is created to enable the physicians and practitioners to achieve high concentration of healthy platelets at the right volumes of PRP, in their own private practices.

• Double CentrifugationTo achieve 1,000,000/µl fragile platelet count, we need to delicately separate RBCs and WBCs from fragile platelets, contained within the plasma. This is attained by the first centrifugation cycle. Obviously, a small percentage of fragile platelets cannot make the journey because they are held back by the bigger RBCs.

13

Don't forget that at the end of the first centrifugation cycle, the concentration level of platelets is less than twice the baseline. That is exactly why AABB recommends double-centrifugation procedure, because this is the only way that fragile platelets will separate from plasma, and minimum three times the base line concentration of platelets can be achieved. Therefore, the right PRP concentration is obtained only after the second spin

cycle, where fragile platelets are separated from Platelet-Poor Plasma (PPP); resulting in the highest collectable platelet count.

• Medical Fields that can highly benefit from EPRP®

• Aesthetic- High Concentration of

14

Healthy platelets, for effective PRP treatments, every time.- The right volume of PRP is attained, due to blood collection capacities, especially for treatments that right volume of high-concentrated healthy platelets are required (i.e. hair recitation, skin rejuvenation).

• Orthopedics- Infection is the leading concern of all Orthopedists around the world, and EPRP® has eliminated this fear, because it is a 100% closed system. No risk of infection. No risk of contamination.- High Concentration of Healthy platelets, for effective PRP treatments, every time.

• Surgery & Stem Cell Therapy

- Stem Cell Therapy can be most effective, when mixed with pure platelets. However, stem cell therapists rather not mix Platelets with Stem Cell! The reason being the low yields of Platelets in the

ordinary PRP kits.- EPRP® PRP Preparation Device offers pure platelets (more than 20x the baseline).

• Wound Healing /Burn Patient

- EPRP® is the only PRP Preparation Device that with «closed-system» properties. After PRP is produced then the packed cells can be re-transfused into the same patient's body.- High Concentration of Healthy platelets, for effective PRP treatments, every time.- The right volume of PRP is attained, due to blood collection capacities, where right volume of high-concentrated healthy platelets are required for effectiveness of the treatment.- Infection is the leading concern of all medical facilities around the world, especially for burn patients. EPRP® is a 100% closed system. No risk of infection. No risk of contamination. 

15

the packed cells and Poor-Platelet Plasma can be re-transfused into patient.

• Bio Filler / Plasma Filler & Cream

In the Triple-Bag Preparation Device, the transferred PPP (Platelet-Poor Plasma) to the third bag, can be used to produce Bio Filler / Plasma Filler for the same patient.The PPP can also be used to produce moisturizing cream for the same patient.

• Cost-BenefitsEPRP® PRP Preparation Device is the most cost-beneficial, value-for-money, PRP preparation device available in the market. It is offered in three convenient blood collection volume sizes:• 100cc (Maximum 85cc

Blood Collection Capacity), perfect for surgery and Lipotransfer procedures.

• 80cc (Maximum 70cc Blood Collection Capacity), standard

• OtherO u t s t a n d i n g Features of EPRP®

PRP Preparation Device• Re-transfuse-abilityIn some critical medical situations, where taking more blood would possibly jeopardize patients' life (e.g. burn victims), and loss of more blood is not an option; while high concentration of platelets are required to aid in speeding the healing process, only EPRP® PRP Preparation Device makes it possible to re-transfuse patient›s own blood (packed cells and plasma) back into the patient's own body.This priceless feature is also advantageous for huge Lipotransfer operations, where initially patient loses excessive volume of blood, during the fat harvesting procedure; where at the completion of PRP preparation procedure,

16

volume for aesthetic procedures like hair resuscitation and skin rejuvenation.

• 40cc (Maximum 35cc Blood Collection Capacity), ideal for PRP treatment procedures in orthopedics and dentistry.

With one blood collection, patient can be treated to several PRP treatments in one session.

• PlasmaTherapy Is NOT PRP Treatment!

The result of effective PRP treatment is the stimulation of fibroblast growth factors for production of collagens (known as Collagen Induction Therapy (CIT)); the effect for which will be apparent to the patient only after few months of multiple treatments. Unfortunately, there is a common misinterpretation that exists among Aesthetic Physicians about PRP treatment in skin rejuvenation therapy

17

verses simple plasma injection.Many physicians confuse the quick effect of «skin hydration» that is due to the injection of plasma (which contains proteins, electrolytes and many other components, that trigger the skin's natural moisture to charge to the surface of the skin via osmotic pressure, giving the skin a temporary, healthy-looking, reduced-

wrinkles, glowing effect); as the indication of the effectiveness of their PRP Treatment.Again, the true intention for applying PRP treatment for Skin Rejuvenation is to induce cell growth in the skin, which results in retarding and reversing the aging process, also known as Collagen Induction Therapy (CIT); which takes few months of multiple treatment sessions.

18

Booklet References

Platelet-rich plasma (PRP): what is PRP and what is not PRP?Marx RE Implant Dent. 2001;10(4):225-8.PMID:11813662

Platelet-Rich Plasma: A Literature ReviewArora, Navneet S. DDS, MPH; Ramanayake, Thaminda MS; Ren, Yan-Fang DDS, MD, PhD; Romanos, Georgios E. DDS, DrMedDent, PhDImplant Dentistry: August 2009 - Volume 18 - Issue 4 - p 303-310doi: 10.1097/ID.0b013e31819e8ec6Clinical Science and Technology

Optimized Preparation Method of Platelet-ConcentratedPlasma and Noncoagulating Platelet-DerivedFactor Concentrates: Maximization of PlateletConcentration and Removal of FibrinogenAraki J1, Jona M, Eto H, Aoi N, Kato H, Suga H, Doi K, Yatomi Y, Yoshimura K.Tissue Eng Part C Methods. 2012 Mar;18(3):176-85. doi: 10.1089/ten.TEC.2011.0308. Epub 2011 Nov 22.

Evaluation of the effects of the use of platelet-rich plasma (PRP) on alveolar bonerepair following extraction of impacted third molars: Prospective studyGuilherme de Marco Antonello*, Ricardo Torres do Couto, Caroline Comis Giongo, Marcos Britto Corrêa,Otacílio Luiz Chagas Júnior, Carmen Helena Jacques LemesJournal of Cranio-Maxillo-Facial Surgery 41 (2013) e70ee75

Quality Parameters of Platelet Concentrate at Kenyatta National Hospital’s Blood Transfusion UnitNjoroge N Richard1, 2,, Maturi P Mwamba2, Githanga J2, Jamilla Rajab21Garissa District Hospital, Ministry of Health

19

Booklet References

International Journal of Hematological Disorders, 2014 1 (1), pp 35-40. DOI: 10.12691/ijhd-1-1-7

Comparative Study of Four Platelet-Rich PlasmaMethods for Preparing Platelet ConcentratesPourmokhtar M 1*, Salek Moghaddam E 2, Abbasi F 2, Zarei N 21. Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.2. Tehran Blood Transfusion Center, Tehran, Iran.IJBC 2014;6(3): 103-107

Effect of Platelet-Rich Plasma on Ischemia-ReperfusionInjury in a Skin Flap Mouse ModelDong Kyun Rah1, Hyung Jun Min, Yang Woo Kim, and Young Woo CheonInt. J. Med. Sci. 2017, Vol. 142017; 14(9): 829-839. doi: 10.7150/ijms.19573

Determining the Effect of Preparation and Storage: AnEffort to Streamline Platelet Components as a Sourceof Growth Factors for Clinical ApplicationAtul Sonker Anju DubeyTransfus Med Hemother 2015;42:174–180DOI: 10.1159/000371504

High-Shear Stress Sensitizes Platelets to SubsequentLow-Shear ConditionsJAWAAD SHERIFF,1 DANNY BLUESTEIN,1 GAURAV GIRDHAR,1 and JOLYON JESTYAnnals of Biomedical Engineering (_ 2010)DOI: 10.1007/s10439-010-9936-2

Platelet-rich plasma for bone healing to use or not to use?Sebastian Lippross, Mauro Alini AODIALOGUE 1 | 07

20

Booklet References

A standardized research protocol for platelet-rich plasma (PRP) preparation in rats

Michel Reis Messora, Maria José Hitomi Nagata, Flávia Aparecida ChavesFurlaneto, Rita Cássia Menegati ornelles, Suely Regina Mogami Bomfim ,Tatiana Miranda Deliberador, Valdir Gouveia Garcia, Alvaro Francisco BoscoRSBO. 2011 Jul-Sep;8(3):299-304

Producing accurate platelet counts for platelet rich plasma: validation of a hematology analyzer and preparation techniques for counting.Woodell-May JE, Ridderman DN, Swift MJ, Higgins J.J Craniofac Surg. 2005 Sep;16(5):749-56; discussion 757-9.PMID:16192851

Platelet Rich Plasma (PRP) GuidelinesInternational Cellular Medicine Society – 2011

PRP in OA knee – update, current confusions and future optionsMandeep S. Dhillon, Sandeep Patel*, and Rakesh JohnDepartment of Orthopaedics, Post Graduate Institute of Medical Education and Research, Chandigarh 160012, IndiaEDP Sciences, 2017DOI: 10.1051/sicotj/2017004

Platelet-rich plasma preparation for regenerative medicine: optimization and quantification of cytokines and growth factorsPaola Romina Amable1, Rosana Bizon Vieira Carias, Marcus Vinicius Telles Teixeira, Ítalo da Cruz Pacheco,Ronaldo José Farias Corrêa do Amaral, José Mauro Granjeiro and Radovan BorojevicStem Cell Research & Therapy 2013, 4:67

AABB Technical Manual 18th Edition ISBN-13: 978-1563958885

21

Booklet References

Platelet-Rich Plasma Peptides: Key for RegenerationDolores Javier S´anchez-Gonz´alez, EnriqueM´endez-Bolaina and Nayeli Isabel Trejo-Bahena

International Journal of PeptidesVolume 2012, Article ID 532519, 10 pagesdoi:10.1155/2012/532519Platelet-Rich Fibrin Promotes an Accelerated Healing of Achilles Tendon When Compared to Platelet-Rich Plasma in RatFranciele Dietrich1, Gustavo L. Duré, Caroline P. Klein, Vinícius F. Bampi, AlexandreV. Padoin, Vinícius D. Silva, Jefferson Braga-Silva

Platelet-Rich Plasma (PRP) for HairSanya Kalra Dr. Komal Mathurwww.ijetsr.com ISSN 2394 – 3386 Volume 4, Issue 6 June 2017

PRP modulates expression of bone matrix proteins in vivo without long-term effects on bone formationMichael Thorwarth, Falk Wehrhan, Stefan Schultze-Mosgau,Jörg Wiltfang, Karl Andreas SchlegelBone. 2006 Jan;38(1):30-40. Epub 2005 Oct 28.PMID:16257592DOI:10.1016/j.bone.2005.06.020

Platelet Rich Plasma and Its Use in Androgenetic Alopecia and Alopecia Areata: A Systematic ReviewMD. Scarlett Alvarado CarvajalSociedad Española de Medicina y Cirugía Cosmética March 2017 Barcelona Spine

WHO guidelines on drawing blood best practices in phlebotomyISBN 978 92 4 159922 1

22