SE540289C2

SE540289C2 - Method, device and kit for the preparation of prp

Info

Publication number: SE540289C2
Application number: SE1650038A
Authority: SE
Inventors: Ghanbari Ahmad
Original assignee: Ghanbari Ahmad
Priority date: 2016-01-14
Filing date: 2016-01-14
Publication date: 2018-05-22
Also published as: SE1650038A1

Abstract

There if provided a medical device (100) comprising a first blood bag (101) for collection of whole blood from a patient and a second blood bag (109) for separation of a platelet rich plasma (PRP) fraction, wherein:a whole blood inlet (102) is provided at a first end (1011) of the first blood bag (101) and a first blood bag outlet (103) is provided at a second end of the first blood bag (101);a packed cells outlet (105) and a plasma outlet (106) are connected to the first blood bag outlet (103);a first flexible plastic tube (107) connects the plasma outlet (106) to a plasma inlet (108) provided at a first end (1091) of the second blood bag (109);a second blood bag outlet (111) is provided at a second end of the second blood bag (109).

Description

METHOD, DEVICE AND KIT FOR THE PREPARATION OF PRP TECHNICAL FIELD The present disclosure relates to the preparation of platelet-rich plasma (PRP).

BACKGROUND Platelet-rich plasma (PRP) is blood plasma that has been enriched with platelets. As a concentrated source of autologous platelets, PRP contains (and releases through degranulation) several different growth factors and other cytokines that stimulate healing, e.g. of bone and soft tissue.

PRP was first developed in the 1970s and first used in Italy in 1987 in an open heart surgery procedure. PRP therapy began gaining popularity in the mid 1990s. Today, PRP injections have been safely used in many fields including wound healing, burn wound treatment, sports medicine, orthopaedics, cosmetics, faciomaxillary, gynaecology and urology.

The efficacy of certain growth factors in healing various injuries and the concentrations of these growth factors found within PRP are the theoretical basis for the use of PRP in tissue repair. The platelets collected in PRP may be activated by the addition of thrombin and calcium chloride, which induces the release of the mentioned factors from alpha granules. The growth factors and other cytokines present in PRP normally comprise: platelet-derived growth factor; transforming growth factor beta; fibroblast growth factor; insulin-like growth factor 1; insulin-like growth factor 2; vascular endothelial growth factor; epidermal growth factor; Interleukin 8; keratinocyte growth factor; and connective tissue growth factor.

Presently there are two methods of PRP preparation approved by the U.S. Food and Drug Administration. Both processes involve the collection of the patient's whole blood that is anticoagulated with citrate dextrose before undergoing two stages of centrifugation designed to separate the PRP aliquot from platelet-poor plasma (PPP) and red blood cells. In humans, the typical baseline blood platelet count is approximately 200,000 per ?L. Typically, the therapeutic PRP concentrates the platelets by roughly five-fold. There is, however, broad variability in the production of PRP by various concentrating equipment and techniques.

SUMMARY The present inventor has identified a number of problems of the prior art PRP preparation systems.

The first problem of the prior art systems is that blood is taken/collected from the patient with a syringe. The inventor has realized that the vacuum pressure in the syringe causes damage to the platelets’ sensitive membrane, which decreases the effectiveness of the resulting PRP. Therefore, it is beneficial to use a blood bag for the collection of blood instead of a syringe.

A second problem of the prior art systems are that they are “open” and thus exposed to the risk of infectious contamination. To improve sterility, it is preferred to have a closed system.

A third problem of the prior art is that a substantial portion of the platelets are damaged by contact with the hard walls of the tubes during centrifugation. This problem is avoided by the using bags instead of tubes in the centrifugation.

A fourth problem of the prior art is that high volumes of blood cannot be taken from anaemic patients or patients having large burn wounds.

The problems of the prior art has been addressed by the inventor and as a first aspect of the present disclosure, there is provided a medical device comprising a first blood bag for collection of whole blood from a patient and a second blood bag for separation of a platelet rich plasma (PRP) fraction, wherein: a whole blood inlet is provided at a first end of the first blood bag and a first blood bag outlet is provided at a second end of the first blood bag; a packed cells outlet and a plasma outlet are connected to the first blood bag outlet; a first flexible plastic tube connects the plasma outlet to a plasma inlet provided at a first end of the second blood bag; a second blood bag outlet is provided at a second end of the second blood bag.

In an alternative (less preferred) configuration of the first aspect, there is not a packed cells outlet and a plasma outlet, but only the first blood bag outlet connected to the plasma inlet of the second blood bag by the first flexible tube.

Packed cells recovered from the packed cells outlet can be reinjected in the blood stream of a critically ill patient and thus allow for taking higher volumes of blood from the critically ill patient.

In one embodiment, the packed cells outlet comprises a breakable lock. Such a lock stops flow though the packed cells outlet and prevents contamination until it has been broken. When the medical device is used as intended, the breakable lock of the packed cells outlet is not broken when the plasma is transferred to the second blood bag. It is only broken when reinfusion of packed cells is needed.

A first branching may be connected to the first blood bag outlet. In such case, the first branching comprises the packed cells outlet and the plasma outlet.

The volume of the plasma (containing platelets) transferred to the second bag is typically about half of the volume of the whole blood provided in the first bag. Accordingly, the volume of the first blood bag may be higher than the volume of the second blood bag.

In one embodiment, the first flexible plastic tube comprises a breakable lock. Such a lock stops flow though the first flexible plastic tube until it has been broken.

Preferably, a platelet-rich plasma (RPR) outlet and a platelet-poor plasma (PPP) outlet are connected to the second blood bag outlet.

PPP recovered from the PPP outlet may be injected to the skin for moisturizing purposes. It may also be used in a moisturizing créme.

Preferably, calcium and eucerin is added to the PPP for such a créme application.

Alternatively, PPP recovered from the PPP outlet may be used in an créme or ointment for topical treatment of a burn wound. Preferably, calcium and eucerin is added to the PPP for such an application. Accordingly, the medical device enables the provision of packed cells, PRP and PPP, which can all be used for the treatment of a patient having a burn wound. The medical device is particularly beneficial when the burn wound is large.

In one embodiment, the platelet-rich plasma (RPR) outlet comprises a breakable lock. Such a lock stops flow though the PRP outlet and prevents contamination until it has been broken. When the medical device is used as intended, the breakable lock of the RPR outlet is not broken until shortly before the PRP fraction is recovered from the second blood bag. For example, the lock in question is intended to be unbroken when the PPP fraction is transferred through the PPP outlet.

Preferably, the device further comprises a third blood bag for collection of platelet-poor plasma (PPP). In such case, a second flexible plastic tube may connect the PPP outlet to a PPP inlet provided at a first end of the third blood bag. Further, the third blood bag may comprise an outlet, for example provided at a second end of the third blood bag, for recovery of PPP from the third bag. Such an outlet may comprise a breakable lock.

In one embodiment, the second flexible plastic tube comprises a breakable lock. Such a lock stops flow though the second flexible plastic tube until it has been broken.

The medical device may comprise a second branching that is connected to the second blood bag outlet. In such case, the second branching comprises the PRP outlet and the PPP outlet.

The walls of the first blood bag, the second blood bag and (if present) the third blood bag are preferably composed of a flexible plastic material according to blood bag standards.

The first blood bag may comprise a first bag extension extending from the walls of the first blood bag. Such an extension enables fixation of the first blood bag during centrifugation. For example, holes penetrable by pins on a centrifuge may be provided in the first bag extension to allow the first blood bag to be fixed, preferably in an extended (non-creased) configuration, during centrifugation.

Likewise, the second blood bag may comprise a second bag extension extending from the walls of the second blood bag. Such an extension enables fixation of the second blood bag during centrifugation. For example, holes penetrable by pins on a centrifuge may be provided in the second bag extension to allow the second blood bag to be fixed, preferably in an extended (non-creased) configuration, during centrifugation.

To reduce the impact on the platelets and to facilitate the separation of platelets from packed cells, the second end of the first blood bag may be funnel-shaped. Alternatively, corners of the second end of the first blood bag may be rounded. However, a funnel shape is more preferred for the first blood bag.

To reduce the impact on the platelets and to facilitate the separation of platelets from (platelet poor) plasma, corners of the second end of the second blood bag may be rounded, e.g. such that the bottom end of the second blood bag has the shape of a half sphere. Alternatively, the second end of the second blood bag may be funnel-shaped.

As a second aspect of the present disclosure, there is provided a use of a medical device according to the first aspect for separation of platelet-rich plasma (RPR) fraction from whole blood.

As a third aspect of the present disclosure, there is provided a method of preparing a platelet-rich plasma (PRP) fraction using the medical device according to the first aspect.

The method comprises the steps of: a) subjecting the first blood bag containing whole blood to a first centrifugation to separate packed cells (primarily RBC) from plasma (containing platelets); b) transferring plasma (containing platelets) from the first blood bag through the first flexible tube (107) to the second blood bag; c) subjecting the second blood bag containing plasma (containing platelets) to a second centrifugation to settle platelets; and d) removing a platelet poor plasma (PPP) fraction from the second blood bag to obtain the PRP fraction in the second blood bag.

The platelet count of the PRP fraction is typically 3-8 times higher than that of the whole blood.

Step b) may further comprise recovering of a packed cells fraction from the first blood bag after the plasma has been transferred to the second bag. At least part of such a packed cells fraction can be reinjected into the bloodstream of a patient, such as a critically ill or an anaemic patient, which preferably is the patient from which the whole blood was taken.

The removal of the PPP fraction of step d) may be transferring the PPP fraction to the third blood bag. Uses of the PPP fraction are discussed above and below.

The whole blood of step a) is preferably mixed with an anticoagulant, such as CPDA1.

The first centrifugation can for example be carried out for a time period of 5-40 minutes at 500-3000 rpm. Preferably, the time period is 10-30 minutes at 750-1800 rpm.

The second centrifugation can for example be carried out for a time period of 1-10 minutes at 1500-4000 rpm. Preferably, the time period is 2-8 minutes 2000-3500 rpm.

During the first and the second centrifugation, the first and second blood bag, respectively, are preferably fixed in an extended configuration. In the extended configuration, the walls of the blood bag in question are not creased to any substantial degree. Pins provided on the centrifuge used for the first and second centrifugations matching holes in extensions from the walls of the blood bags may provide such a fixation.

The method may further comprise taking whole blood from a subject using a needle connected to the whole blood inlet to obtain the first blood bag containing whole blood of step a) and administration of at least part of the PRP fraction obtained in step d) to the subject.

Such a method comprising the taking of whole blood and administration of PRP may be therapeutic or non-therapeutic. An example of a non-therapeutic method is a cosmetic method.

The administration method may for example be spraying, which is particularly beneficial in the treatment of large burn wounds (e.g. > 0.2 m<2>) where it is necessary to cover a large area with a limited amount of available PRP. Spraying is further discussed below with reference to Fig 3.

The present disclosure thus provides the concept of treating a patient having a burn wound by spraying PRP onto the burn wound. The PRP for this concept can be any type of PRP, but it is preferably PRP prepared according to the present disclosure.

As a fourth aspect of the present disclosure, there is provided a kit-of-parts comprising the medical device according to the first aspect and a centrifuge. The centrifuge may be provided with pins capable of fixing a blood bag in an extended configuration during centrifugation.

BRIEF DESCRIPTION OF THE DRAWINGS The invention is now described, by way of example, with reference to the accompanying drawings, in which: Fig 1 shows an embodiment of the system/medical device of the present disclosure.

Fig 2 shows a folded configuration of the medical device of Fig 1.

Fig 3 show administration of PRP to a burn wound by means of spraying.

DETAILED DESCRIPTION Non-limiting examples of embodiments of the system and the method of the present disclosure are described below.

The exemplary system is a three-bag system 100. The three-bag system may also be referred to as a medical device.

A first bag 101 of the three-bag system 100 is a blood bag for collection of whole blood from a patient. The walls of the first bag 101 are composed of a flexible plastic material according to blood bag standards. The first bag 101 further comprises a first bag extension 101a extending from the walls of the first bag 101. In the first bag extension 101a, holes 101b are provided to allow the first bag 101 to be fixed in an extended configuration during centrifugation. A whole blood inlet 102 is arranged at a first end 1011 of the first bag 101 and a blood bag outlet 103 is arranged at a second end of the first blood bag 101. To reduce the impact on the platelets and to improve the separation of platelets from packed cells, the second end of the first bag 101 is funnel-shaped.

A first branching 104 is connected to the blood bag outlet 103. The first branching 104 comprises a packed cells outlet 105 and a plasma outlet 106 (for plasma containing platelets). The packed cells outlet 105 comprises a breakable lock 105a that i.a. prevents contamination. A first flexible plastic tube 107 connects the plasma outlet 106 to a plasma inlet 108 arranged at a first end 1091 of a second bag 109. The walls of the second bag 109 are composed of a flexible plastic material according to blood bag standards. The second bag 109 further comprises a second bag extension 109a extending from the walls of the second bag 109. In the second bag extension 109a, holes 109b are provided to allow the second bag 109 to be fixed in an extended configuration during centrifugation. An outlet 111 is arranged at a second end of the second bag 109. To reduce the impact on the platelets and to improve the separation of platelets from (platelet poor) plasma, the corners 109c of the second end of the second bag 109 are rounded, e.g. such that the bottom end of the second bag 109 has the shape of a half sphere. The first flexible plastic tube 107 comprises a breakable lock 107a. Further, a first clamp 110 is arranged on the first flexible plastic tube 107 such that flow through the first flexible plastic tube 107 can be stopped also after the breakable lock 107a has been broken. The first clamp 110 may be arranged upstream the breakable lock 107a.

A second branching 112 is connected to the outlet 111 of the second bag 109. The second branching 112 comprises a platelet-rich plasma (RPR) outlet 113 and a platelet-poor plasma (PPP) outlet 114. The PRP outlet 113 comprises a breakable lock 113a that i.a. prevents contamination. A second flexible plastic tube 115 connects the PPP outlet 114 to a PPP inlet 116 arranged at a first end 1181 of a third bag 118 for collection of PPP. The walls of the third bag 118 are composed of a flexible plastic material according to blood bag standards. The second flexible plastic tube 115 comprises a breakable lock 115a. Further, a second clamp 117 is arranged on the second flexible plastic tube 115 such that flow through the second flexible plastic tube 115 can be stopped also after the breakable lock 115a has been broken. The second clamp 117, may be arranged upstream the breakable lock 115a.

In the method, whole blood is first collected from a patient in the first bag 101 (i.e. the blood bag). A clamp 120 is then used to prevent blood from leaking through the whole blood inlet 102 of the first bag 101. The collected volume of whole blood may for example be about too ml. In the first bag 101, the whole blood is mixed with an anticoagulant, preferably CPDA1. The collection and mixing with the anticoagulant are well known to the skilled person. The first bag 101 containing the whole blood mixed with the anticoagulant is then subjected to a first centrifugation, e.g. for 18 minutes at 1100 rpm, for separation of red blood cells (RBC) from plasma (containing platelets).

During centrifugation, pins of the centrifuge penetrate the holes 101b of the first bag extension 101a to fix the first bag 101 in an extended configuration (compaction of the bag is thus avoided). The orientation of the first bag 101 during the first centrifugation is such that a packed cells fraction is formed at the first end 1011 of the first bag 101.

After the first centrifugation, the breakable lock 107a of the first flexible tube 107 is broke open and the first bag is gently squeezed such that the supernatant, i.e. the platelet-containing plasma, is gently pushed through the plasma outlet 106 and the first flexible tube 107 to the second bag 109. Then, the flow through the first flexible tube 107 is stopped by the first clamp 110. The packed cells fraction remains in the first bag and can be obtained through the packed cells outlet 105 after that the breakable lock 105a has been broke open. The packed cells fraction may be reinjected into the blood stream of the patient, in particular when the patients is anaemic or has large burn wounds.

The concentration of platelets in the plasma transferred to the second bag 109 is typically about 1.5 higher than in the whole blood. That means that if the platelet count in the whole blood was 200000 per ?l, it is typically 300000 per ?l in the plasma. If the original volume of whole blood was too ml, the volume of plasma transported to the second bag 109 is typically about 50 ml (which means that about 25% of the platelets could not be separated from the packed cells if the platelet concentration 1.5 times higher in the separated plasma than in the whole blood). To further increase the platelet count, the second bag 109 containing the plasma is subjected to a second centrifugation, e.g. for 4 minutes at 2700 rpm. During the second centrifugation, the orientation of the second bag is such that platelets settle at the first end 1091 of the second bag 109 (the first end 1091 is the end by the inlet 108 of the second bag 109). During centrifugation, pins of the centrifuge penetrate the holes 109b of the second bag extension 109a to fix the second bag 109 in an extended configuration. After the second centrifugation, the second bag is rested, e.g. for 30-90 minutes to allow expansion of the settled platelets.

The breakable lock 115a of the second flexible tube 115 is then broke open and the second bag 109 is squeezed such that the supernatant, i.e. the plateletpoor plasma (PPP), is pushed through the PPP outlet 106 and the second flexible tube 107 to the third bag 118. The PPP collected in the third bag 118 may for example be used as a skin moisturizer.

Then, the flow through the second flexible tube 115 is stopped by the second clamp 117 and a plasma fraction containing the expanded platelets settled at the first end 1091 of the second bag 109, i.e. the platelet-rich plasma (PRP), is recovered through the PRP outlet 113 by gently squeezing the second bag 109 after the breakable lock 113a of the PRP outlet has been broke open. The platelet count of the PRP fraction is typically 3-8 times higher than that of the whole blood.

Figure 2 shows a folded configuration of the medical device of Figure 2 that can be used during the centrifugations. Pins 201 penetrate holes 101b, 109b, 118b made in the extensions 101a, 109a, 118a from the walls of the bags 101, 109, 118. Thereby, the bags 101, 109, 118 are fixed in extended configurations during the centrifugations. During the first centrifugation, the folded configuration preferably has the orientation shown in Figure 2 such that packed cells are formed (settle) at the first end 1011 of the first bag 101.

During the second centrifugation, the folded configuration of figure 2 is preferably turned upside-down such that platelets settle at the first end 1091 of the second bag 109.

Figure 3 shows a spraying device 300 by means of which PRP is sprayed onto a large burn wound 301. The spaying device 300 preferably composed of such material(s) that it can be sterilized by autoclaving. An example of such a material is metal, preferably stainless steel.

The invention has mainly been described above with reference to a few embodiments. However, as is readily appreciated by a person skilled in the art, other embodiments than the ones disclosed above are equally possible within the scope of the invention, as defined by the appended patent claims.

Claims

1. A medical device (100) comprising a first blood bag (101) for collection of whole blood from a patient and a second blood bag (109) for separation of a platelet rich plasma (PRP) fraction, wherein: a whole blood inlet (102) is provided at a first end (1011) of the first blood bag (101) and a first blood bag outlet (103) is provided at a second end of the first blood bag (101); a packed cells outlet (105) and a plasma outlet (106) are connected to the first blood bag outlet (103); a first flexible plastic tube (107) connects the plasma outlet (106) to a plasma inlet (108) provided at a first end (1091) of the second blood bag (109); a second blood bag outlet (111) is provided at a second end of the second blood bag (109) wherein a first branching (104) is connected to the first blood bag outlet (103), which first branching (104) comprises the packed cells outlet (105) and the plasma outlet (106).

2. The medical device of claim 1, wherein the packed cells outlet (105) comprises a breakable lock (105a).

3. The medical device of any one of claim 1-2, wherein the volume of the first blood bag (101) is higher than the volume of the second blood bag (109).

4. The medical device of any one of claims 1-3, wherein the first flexible plastic tube (107) comprises a breakable lock (107a).

5. The medical device of any one of claims 1-4, wherein a platelet-rich plasma (RPR) outlet (113) and a platelet-poor plasma (PPP) outlet (114) are connected to the second blood bag outlet (111).

6. The medical device of claim 5, wherein the platelet-rich plasma (RPR) outlet (113) comprises a breakable lock (113a).

7. The medical device of any one of claims 5-6, further comprising a third blood bag (118) for collection of platelet-poor plasma (PPP) and wherein a second flexible plastic tube (115) connects the PPP outlet (114) to a PPP inlet (116) provided at a first end (1181) of the third blood bag (118).

8. The medical device of claim 7, wherein the second flexible plastic tube (115) comprises a breakable lock (115a)

9. The medical device of anyone of claims 5-8, wherein a second branching (112) is connected to the second blood bag outlet (111), which second branching (111) comprises the PRP outlet (113) and the PPP outlet (114).

10. Use ex vivo of a medical device according to any one of the preceding claims for separation of platelet-rich plasma (PRP) fraction from whole blood.

11. Method of preparing a platelet-rich plasma (PRP) fraction using the medical device (100) according to any one of claims 1-9, comprising the steps of: a) subjecting the first blood bag (101) containing whole blood to a first centrifugation to separate packed cells from plasma; b) transferring plasma from the first blood bag (101) through the first flexible tube (107) to the second blood bag (109); c) subjecting the second blood bag (109) containing plasma to a second centrifugation to settle platelets; and d) removing a platelet poor plasma (PPP) fraction from the second blood bag to obtain the PRP fraction in the second blood bag (109).