High-purity P-PRP preparation system and preparation method thereof

Abstract

The invention discloses a high-purity P-PRP preparation system and a preparation method thereof. The high-purity P-PRP preparation system comprises a sterile syringe for sampling, a centrifugal machine for providing a centrifugal effect, a centrifugal sleeve and a separator for purifying a sample of the centrifuged sterile syringe, the centrifugal sleeves are of two types and comprise an inverted centrifugal sleeve used for first-time centrifugation and an upright centrifugal sleeve used for second-time centrifugation. The preparation method comprises the steps of sample preparation, primary centrifugation, purification and re-sampling, secondary centrifugation, final sampling and the like, and finally the leukocyte-depleted platelet-rich plasma with high purity is obtained. According to the system and the method, a conventional medical general syringe can be directly used for the closed centrifugal sleeve, the problem that the syringe makes contact with the outside in the transferring and centrifuging process is effectively solved, the pollution risk in the PRP preparation process is reduced, meanwhile, P-PRP is conveniently extracted, the influence of liquid hanging on the wall on purity is avoided, the purity is improved, and residual pollution is rejected.

Description

Technical Field

[0001] This invention belongs to the field of medical devices, and specifically relates to a PRP preparation and extraction device system and method, specifically a high-purity P-PRP preparation system and its preparation method. Background Technology

[0002] Platelet-rich plasma (PRP) is an extract from autologous whole blood containing high concentrations of platelets and growth factors. Recent studies have shown a close relationship between the concentrations of interleukin 1β (IL-1β) and tumor necrosis factor α (TNF-α) in PRP and its leukocyte concentration, suggesting that platelet-rich plasma (L-PRP) has a strong anti-inflammatory effect. Simultaneously, high leukocyte concentrations may inhibit tissue repair. P-PRP has gained increasing attention in recent years, and the methods for its preparation are constantly evolving. Existing P-PRP preparation methods can be broadly classified into two categories: blood filtration and centrifugation. The latter has advantages such as simple operation, no need for special equipment and consumables, and low cost, making it more suitable for clinical use.

[0003] L-PRP: Platelet-rich plasma containing a high concentration of white blood cells, abbreviated as L-PRP (Leukocyte Platelet-Rich Plasma, containing a small number of red blood cells), is mainly used for the clinical treatment of refractory wounds, diabetic foot, gout with non-healing wounds, bone repair, nonunion of bones, osteomyelitis, etc.

[0004] P-PRP: Platelet-rich plasma (P-PRP) is abbreviated as P-PRP (Pure Platelet-Rich Plasma, which does not contain red blood cells) and contains little or no white blood cells. It is mainly used for sports injuries and degenerative diseases, including meniscus injuries, ligament and tendon injuries, tennis elbow, knee arthritis, cartilage degeneration, lumbar disc herniation and other diseases.

[0005] The PRP discussed in this invention refers to P-PRP.

[0006] Currently, sterile syringes are mainly used for PRP sampling. After the first centrifugation, the blood sample separates into layers, from top to bottom:

[0007] 1. Plasma layer: This is usually the uppermost layer and is typically a pale yellow, transparent liquid containing water, plasma globulins, serum proteins, inorganic salts, and other substances.

[0008] 2. Leukocyte and platelet layer: This is the middle layer, usually a thin white layer, which mainly contains leukocytes such as neutrophils, basophils, and monocytes, as well as platelets;

[0009] 3. Red blood cell layer: This is the lowest layer, mainly composed of red blood cells. It is dark red and opaque solid.

[0010] The plasma layer accounts for approximately 40-44% of the total volume, the intermediate layer accounts for approximately 1-5% of the total volume, which is pure PRP, but because it is relatively small, it is easy to mix with the upper and lower layers. Red blood cells (account for approximately 55% of the total volume) are also present. The extraction of PRP is the focus of this invention. In actual use, PRP contains a portion of the plasma layer.

[0011] However, at present, most domestic and foreign countries use open manual secondary centrifugation method to prepare PRP. The existing P-PRP preparation method has the following defects: (1) The open preparation system is easily contaminated by the outside world; (2) During the sample extraction process, it is easy to cause residues of removed substances (some red blood cell residues or plasma layer residues), and the final sample concentration is not high enough and not pure enough.

[0012] In the development process of this invention, based on actual needs and with reference to some existing literature, some of the patent documents referenced are as follows:

[0013] Reference 1: Chinese Patent Application No. CN2016110745999 discloses a method for preparing platelet-rich plasma (PRP) and the uses of the platelet-rich plasma. The method for extracting platelet-rich plasma from blood includes the following steps: (a) placing the collected whole blood in a container containing an anticoagulant and mixing the blood and anticoagulant thoroughly; (b) placing the blood mixed with the anticoagulant in a centrifuge tube and centrifuging for the first time, so that the blood is basically separated into three layers; (c) extracting the top layer and most of the middle layer and transferring them to a new centrifuge tube, mixing them evenly, and centrifuging for the second time; (d) discarding the plasma on the top layer of the centrifuge tube and using the remaining plasma to resuspend the precipitated platelets, thereby obtaining platelet-rich plasma. First, the method does not specify the centrifugation and separation equipment used; the scheme is only a rough outline, and ordinary technicians in the field do not have suitable equipment to perform the separation operation. Second, when preparing the final sample, the method states that "the plasma on the top layer of the centrifuge tube is discarded, and the remaining plasma is used to resuspend the precipitated platelets to obtain platelet-rich plasma (PRP)." In actual operation, some residual plasma will remain on the centrifuge tube wall during the process of discarding the plasma on the top layer of the centrifuge tube.

[0014] Reference 2: Chinese Patent Application No. CN202211284730X discloses a PRP preparation device and preparation method, including a first centrifuge tube, a second centrifuge tube, and a pipette connecting the cavities of the first centrifuge tube and the second centrifuge tube; each of the first centrifuge tube and the second centrifuge tube has a sealing cap at one end that can be pierced by a needle; one end of the pipette is inserted into the cavity of the first centrifuge tube, and the other end is inserted into the cavity of the second centrifuge tube; the insertion depth of the pipette end inserted into the first centrifuge tube is adjustable; and the pipette end inserted into the second centrifuge tube is fixedly connected to the second centrifuge tube.

[0015] Reference 3: Chinese Patent Application No. 2024116583808 discloses a PRP preparer and a PRP preparation method. The preparer includes a tube body, an upper cover, a lower cover, and a sealing plug. The tube body includes an upper section tube, a middle section tube, and a lower section tube. The upper cover cooperates with the upper section tube to form an upper cavity. The upper cover can be moved up and down to switch between a first position and a second position. The upper cover is provided with an injection hole, an aspiration tube, and a sealing element. The lower cover cooperates with the lower section tube to form a lower cavity. The lower cover can be moved up and down to adjust the volume of the lower cavity. The sealing plug is used to control the opening and closing of the injection hole and the aspiration tube. The portion of the aspiration tube located in the upper cavity has a set length. The set length allows the aspiration tube to be located within the platelet-rich plasma layer in the second position and to maintain a distance from the platelet-rich plasma layer, and allows the port of the aspiration tube to be located outside the platelet-rich plasma in the first position.

[0016] References 2 and 3 both use funnel-like containers for centrifuging blood samples, which requires additional blood drawing and is relatively complicated. Additional medical contact with blood requires higher-level drug registration, which is costly and has poor versatility.

[0017] Reference 4: Chinese Patent Application No. CN202010262345X discloses a PRP centrifuge sleeve, a PRP centrifuge, and a PRP preparation method. This method involves passing a pulsed alternating current through a spiral coil during centrifugation, causing the coil to generate a magnetic field with changing direction that acts on the charged growth factors in the blood collection tube. This method directly applies electrophysiological and chemical properties to the growth factors; whether this affects their biological activity, whether its therapeutic effect is compromised, and whether there are other adverse effects require further investigation.

[0018] Reference 5: Chinese Patent Application No. 2021226175525 discloses a centrifuge for PRP preparation. This is the prior art of the present patent applicant and is the centrifuge used in the technical solution of this application.

[0019] Reference 6: Chinese Patent Application No. 2021209222756 discloses a closed centrifuge sleeve device, which is the prior art of the present patent applicant and is a positive centrifuge sleeve used in the technical solution of this application.

[0020] Reference 7: Chinese Patent Application No. 2022202070259 discloses an inverted PRP centrifuge sleeve, which is the prior art of the present patent applicant and is an inverted centrifuge sleeve used in the technical solution of this application.

[0021] Reference 8: Chinese Patent Application No. CN2021112675686 discloses a centrifugation system and method for producing leukocyte-depleted platelet-rich plasma. The system includes a sterile syringe for sampling, a centrifuge for centrifugation, an inverted centrifuge sleeve for use with the centrifuge, and a separator for purifying the centrifuged sterile syringe sample. The production method includes steps such as sample preparation, first centrifugation, purification and re-sampling, second centrifugation, and final sampling, ultimately obtaining high-purity leukocyte-depleted platelet-rich plasma.

[0022] This is the technical solution first applied for by the patent applicant. The solution is a secondary centrifugation operation performed by simply inverting the centrifuge tube. However, in the actual use of this solution for P-PRP extraction, there is a problem of adhesion to the wall, which leads to a low sample concentration.

[0023] The above-mentioned patented technologies do not disclose the technical components of the present invention or provide any inspiration for the technical components of the present invention. Summary of the Invention

[0024] The purpose of this invention is to address the above-mentioned problems by providing a high-purity P-PRP preparation system and its preparation method. This system allows conventional medical sterile syringes to be directly used in closed centrifuge tubing, effectively solving the problem of contact with the external environment during transfer and centrifugation, and reducing the risk of contamination during P-PRP preparation. Furthermore, multiple tubings can be used for separate centrifugation, improving clinical efficiency and reducing usage costs.

[0025] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0026] A high-purity P-PRP preparation system includes a sterile syringe for sampling, a centrifuge for centrifugation, a centrifuge sleeve used in conjunction with the centrifuge, and a separator for purifying the sample from the centrifuged sterile syringe. The centrifuge sleeve is detachably filled with the sterile syringe to be centrifuged, and the injection port of the sterile syringe after centrifugation contains the sample component to be used. The separator is a three-way tube with an end seal at one end, and the other two ends are connected to the centrifuged sterile syringe and the sterile syringe for extraction, respectively. There are two types of centrifuge sleeves: an inverted centrifuge sleeve for the first centrifugation and an upright centrifuge sleeve for the second centrifugation.

[0027] Furthermore, after the first centrifugation, the top of the sterile syringe placed upside down inside the centrifuge sleeve is connected to a three-way valve. The other opening of the three-way valve is connected to the injection port of another sterile syringe, which is used to extract the upper plasma layer and the middle white blood cell and platelet layer (PRP).

[0028] Furthermore, after the second centrifugation, the bottom of the sterile syringe placed inside the centrifuge sleeve is connected to a three-way valve. The other opening of the three-way valve is connected to the injection port of another sterile syringe, which is used to extract white blood cells and platelet-rich plasma (PRP) from the lower end.

[0029] Furthermore, the fixing device provided on the top inner side of the sealing cap of the inverted centrifuge sleeve is ring-shaped and is matched and coupled to the end seal of the syringe containing the test sample.

[0030] Furthermore, the bottom of the receiving cavity of the upright centrifuge sleeve is provided with an inner support platform, and the center of the inner support platform is provided with a support hole, the size of which matches the injection port of the sterile syringe.

[0031] This invention also discloses its preparation method: a method for preparing high-purity P-PRP, comprising the following steps:

[0032] (1) Sample preparation: Use a sterile syringe to draw whole blood samples to be centrifuged;

[0033] (2) First centrifugation: After removing the plunger from the end of the sterile syringe, place the sterile syringe upside down into the inverted centrifuge tube, and place the inverted centrifuge tube on the centrifuge for the first centrifugation operation. The centrifugation speed is 150g-210g and the duration is 10-15min. After centrifugation, the liquid in the sterile syringe will separate into 3 layers.

[0034] (3) Purification and sampling: Take another sterile syringe and connect one end of the three-way connector, seal the other end of the three-way connector, and connect the third end of the three-way connector to the injection port of the sterile syringe after the first centrifugation; at this time, the injection port of the sterile syringe after centrifugation is facing upward, and draw out its upper layer liquid by taking another sterile syringe. At this time, the upper layer liquid contains the plasma layer as well as the white blood cell and platelet layers, which are clearly separated from the red blood cell layer. Its volume is about 40%-45% of the whole blood volume.

[0035] (4) Second centrifugation: Place the sterile syringe containing the upper liquid extracted in (3) into the upright centrifuge tube, and place the upright centrifuge tube on the centrifuge for a second centrifugation operation. The centrifugation speed is 240g-350g and the duration is 12-18min.

[0036] (5) Final sampling: The upper liquid is sampled again using the method in (3). At this time, the injection port of the sterile syringe after centrifugation is facing down. The lower liquid is extracted by taking another sterile syringe to obtain the final pure L-PRP. Its volume is about 20% of the liquid volume in the sterile syringe after the second centrifugation.

[0037] Furthermore, during purification and resampling in step (3), when pulling the plunger of the new sterile syringe, the liquid is drawn until the red blood cell layer is clearly separated and the bottom plunger plane of the sterile syringe from the first centrifugation is close to the surface. The volume drawn is about 40%-45% of the whole blood volume.

[0038] Furthermore, during the final sampling in step (5), when pulling the plunger of the new sterile syringe, the liquid is drawn until the plasma layer is clearly separated and the bottom plunger plane of the sterile syringe after the second centrifugation is close to the plane. The volume drawn is about 20% of the liquid volume in the sterile syringe after the second centrifugation.

[0039] Furthermore, in step (2), placing the sterile syringe upside down into the inverted centrifuge tube means that the plunger of the sterile syringe is facing down and the injection port is facing up.

[0040] Furthermore, in step (4), placing the sterile syringe upright into the upright centrifuge tube means that the plunger of the sterile syringe is facing upward and the injection port is facing downward.

[0041] The beneficial effects of this invention are:

[0042] This invention provides a high-purity P-PRP preparation system and method, which allows conventional medical sterile syringes to be directly used in closed centrifuge cannulas, effectively solving the problem of contact with the external environment during transfer and centrifugation, and reducing the risk of contamination during P-PRP preparation. Furthermore, multiple cannulas can be used for separate centrifugation, improving clinical efficiency and reducing usage costs.

[0043] 1. To address the issue of open preparation systems being susceptible to external contamination, this invention allows the direct use of standard medical sterile syringes in closed centrifuge tubing, effectively resolving the problem of contact with the external environment during transfer and centrifugation, and reducing the risk of contamination during P-PRP preparation. Furthermore, multiple tubings can be used for separate centrifugation.

[0044] 2. Regarding the problem of residual substances (partial red blood cell residue or plasma layer residue) that are easily caused during sample extraction, this invention breaks through the conventional constraint of using only one upright or upside-down sample placement. It adopts a centrifugation method of first placing the sample upside down and then upright, and uses two sets of centrifuge sleeves to effectively avoid the problem of sample sticking to the wall. In addition, the detachable base makes it easy to install and fix, and also makes it easy to clean.

[0045] 3. Using the present invention, the sample is first centrifuged upside down, and then centrifuged upright again. The chromatogram of the sample inside the tube is shown below. Figure 9 As shown, in the final third sterile syringe C, the leukocyte-depleted platelet-rich plasma (P-PRP) layer is located below the syringe outlet, making it convenient to directly remove the required P-PRP sample.

[0046] No need to be like Figure 8 The plasma layer is pushed out first, and then the P-PRP layer is pushed out, thus avoiding the plasma layer adhering to the wall and causing impurities in the P-PRP sample.

[0047] At the same time, it is not necessary to be like Figure 7 After the first centrifugation, as in the case described above, the red blood cell layer needs to be removed before centrifugation. At this time, the red blood cell layer will stick to the wall, resulting in red blood cell residue before the second centrifugation.

[0048] 4. For the inverted sleeve, the fixing device located on the top inner side of the sealing cap cooperates with the end seal of the syringe containing the test sample. This fixing device is annular and is coupled to the end seal of the syringe containing the test sample. In this way, the sealing cap and the bottom fixing seat can stably fix the syringe from top to bottom, avoiding shaking and failure during centrifugation.

[0049] 5. This centrifuge tube can be placed in a conventional centrifuge tube rack, solving the problem of contamination caused by contact with the external environment during the transfer and centrifugation process of P-PRP preparation. It is also compatible with standard medical sterile syringes, offering strong compatibility. Attached Figure Description

[0050] Figure 1 This is a schematic diagram of the plasma obtained after the first centrifugation using an existing sterile syringe.

[0051] Figure 2 This is a perspective view and a top view of an existing centrifuge device.

[0052] Figure 3 This is a schematic diagram of the centrifuge sleeve filling process during centrifuge operation according to the present invention.

[0053] Figure 4 This is a schematic diagram of the upright cannula of the present invention and its filling with a sterile syringe.

[0054] Figure 5 This is a schematic diagram of the inverted cannula and its filling with a sterile syringe according to the present invention.

[0055] Figure 6 This is a schematic diagram of the assembly of the sterile syringe and the three-way valve of the present invention.

[0056] Figure 7 This invention provides a method for preparing P-PRP by positive-side-up two-stage centrifugation.

[0057] Figure 8 This invention provides a method for preparing P-PRP through inverted double centrifugation.

[0058] Figure 9 This invention relates to a method for preparing P-PRP by first inverting the container and then centrifuging it twice.

[0059] Figures 10-11 This is a sample composition analysis table of Wang XX's whole blood and PRP prepared for laboratory testing.

[0060] Figures 12-13 This is the test report for the whole blood and PRP samples prepared by Li XX during hospital testing.

[0061] Figures 14-15 This is the test report for the whole blood and PRP samples prepared for hospital testing of Lei XX.

[0062] Figures 16-17 This is the test report for whole blood and PRP samples prepared by Jiang XX during hospital testing.

[0063] The text labels in the diagram represent: 80, sterile syringe; 801, plunger rod; 802, injection port; 60, tee tube; 601, threaded interface; 602, screw cap; 603, rotary switch;

[0064] 90. Centrifuge; 91. Cover plate; 901. Cover plate cavity;

[0065] 92. PRP centrifuge stand; 921. Fixing bracket; 922. Fixing cylinder;

[0066] 93. Centrifuge sleeve; 94. Cap;

[0067] 931. Fixed seat; 932. Inner support platform; 933. Support hole. Detailed Implementation

[0068] To enable those skilled in the art to better understand the technical solution of the present invention, the present invention will be described in detail below with reference to the accompanying drawings. The description in this part is only exemplary and explanatory, and should not be used to limit the scope of protection of the present invention in any way.

[0069] Figure 1 This diagram illustrates the plasma obtained after the first centrifugation using a sterile syringe. The sterile syringe 80 in the diagram has had its plunger removed and the bottom needle replaced with an end seal. The whole blood sample to be centrifuged is sealed between the plunger and the end seal. The stratification after the first centrifugation is as follows. Figure 1 The example shown is for ease of understanding of obtaining P-PRP using a two-stage centrifugation method and the phenomenon of blood centrifugation stratification.

[0070] Figure 2 This is a perspective view and a top view of an existing centrifuge device. Figure 3 This is a schematic diagram of the centrifuge sleeve filling during centrifuge operation according to the present invention. It is a schematic diagram of a conventional PRP centrifuge, showing the fixed mounting bracket portion of the centrifuge (only parts are omitted). The diagram includes a PRP centrifuge bracket 92, a fixing frame 921, and a fixing cylinder 922 for fixing the sample. The PRP centrifuge bracket 92 is disposed within the cover cavity 901 of the centrifuge 90. Existing technology involves placing the syringe in... Figure 2 When centrifuging on equipment, it is not easy to fix the sample and it is easy to contaminate the sample.

[0071] Example 1: This is one embodiment of the present invention. There are other possible similar structures. The structural description herein does not constitute a limitation on the claims of this application. The specific details are as follows:

[0072] Combination Figure 1-6 ,as well as Figure 9 This invention relates to a method for preparing P-PRP through a two-stage centrifugation process, involving inverting and then repositioning the syringe. A high-purity P-PRP preparation system includes a sterile syringe 80 for sampling, a centrifuge 90 for centrifugation, and centrifuge sleeves (93(B) is a repositioned centrifuge sleeve, and 93(A) is an inverted centrifuge sleeve) used in conjunction with the centrifuge 90, as well as a separator for purifying the sample from the centrifuged sterile syringe 80. The centrifuge sleeve is detachably filled with the sterile syringe to be centrifuged, and the injection port of the sterile syringe after centrifugation contains the sample component to be used. The separator is a three-way tube 60, with one end sealed during use, and the other two ends connected to the centrifuged sterile syringe 80 and the sterile syringe used for extraction, respectively. Two types of centrifuge sleeves are included: an inverted centrifuge sleeve 93(A) used for the first centrifugation and a repositioned centrifuge sleeve 93(B) used for the second centrifugation.

[0073] The centrifuge sleeve includes a barrel body and a cap 94 that can be screwed open and closed at the top.

[0074] The separator is a 60mm three-way connector, with a sealed end at one end and the other two ends connected to a sterile syringe after centrifugation and a sterile syringe for extraction, respectively. Figure 6 and Figure 9 The sterile syringe after centrifugation is located at the lower end, and its injection port 802 is connected to the second connector B of the three-way tube 60; the injection port 802 of another sterile syringe is connected to the first connector A. By pulling the plunger rod 801 of this sterile syringe, the upper layer liquid in the sterile syringe after the first centrifugation can be drawn into the new sterile syringe through the three-way tube 60.

[0075] Preferably, after the first centrifugation, the top of the sterile syringe placed upside down in the centrifuge sleeve is connected to a three-way valve, and the other opening of the three-way valve is connected to the injection port of another sterile syringe for extracting the upper plasma layer and the middle leukocyte and platelet layer (PRP).

[0076] Preferably, after the second centrifugation, the bottom of the sterile syringe 80, which is placed inside the centrifuge sleeve, is connected to a three-way valve. The other opening of the three-way valve is connected to the injection port of another sterile syringe for extracting white blood cells and platelet-rich plasma (PRP) from the lower end.

[0077] Preferably, the fixing device provided on the top inner side of the sealing cap of the inverted centrifuge sleeve is ring-shaped and is matched and coupled to the end seal of the syringe containing the test sample.

[0078] Preferably, the bottom of the receiving cavity of the upright centrifuge sleeve is provided with an inner support platform 932, and the center of the inner support platform 932 is provided with a support hole 933, the size of which matches the injection port of the sterile syringe 80.

[0079] Example 2:

[0080] Using the centrifuge system equipment in Example 1, see [link / reference] Figure 9 The present invention also discloses its preparation method: a method for preparing high-purity P-PRP, comprising the following steps:

[0081] (1) Sample preparation: Use a sterile syringe to draw whole blood samples to be centrifuged;

[0082] (2) First centrifugation: After removing the plunger from the end of the sterile syringe, place the sterile syringe upside down into the inverted centrifuge tube, and place the inverted centrifuge tube on the centrifuge for the first centrifugation operation. The centrifugation speed is 150g-210g and the duration is 10-15min. After centrifugation, the liquid in the sterile syringe will separate into 3 layers.

[0083] (3) Purification and sampling: Take another sterile syringe and connect one end of the three-way connector, seal the other end of the three-way connector, and connect the third end of the three-way connector to the injection port of the sterile syringe after the first centrifugation; at this time, the injection port of the sterile syringe after centrifugation is facing upward, and draw out its upper layer liquid by taking another sterile syringe. At this time, the upper layer liquid contains the plasma layer as well as the white blood cell and platelet layers, which are clearly separated from the red blood cell layer. Its volume is about 40%-45% of the whole blood volume.

[0084] (4) Second centrifugation: Place the sterile syringe containing the upper liquid extracted in (3) into the upright centrifuge tube, and place the upright centrifuge tube on the centrifuge for a second centrifugation operation. The centrifugation speed is 240g-350g and the duration is 12-18min.

[0085] (5) Final sampling: The upper liquid is sampled again using the method in (3). At this time, the injection port of the sterile syringe after centrifugation is facing down. The lower liquid is extracted by taking another sterile syringe to obtain the final pure L-PRP. Its volume is about 20% of the liquid volume in the sterile syringe after the second centrifugation.

[0086] Preferably, when purifying and re-sampling in step (3), when pulling the plunger of the new sterile syringe, the liquid is drawn until the red blood cell layer is clearly separated and the bottom plunger plane of the sterile syringe from the first centrifugation is close to the plane of the first centrifugation. The volume drawn is about 40%-45% of the whole blood volume.

[0087] Preferably, in step (5), when performing the final sampling, the liquid is drawn until the plasma layer is clearly separated and the stratification line is close to the bottom plunger plane of the sterile syringe after the second centrifugation, and the volume drawn is about 20% of the liquid volume in the sterile syringe after the second centrifugation.

[0088] Preferably, in step (2), placing the sterile syringe upside down into the inverted centrifuge tube means that the plunger of the sterile syringe is facing down and the injection port is facing up.

[0089] Preferably, in step (4), placing the sterile syringe upright into the upright centrifuge tube means that the plunger of the sterile syringe faces upward and the injection port faces downward.

[0090] Example 3: See Figure 7 The present invention provides a method for preparing P-PRP by secondary centrifugation in a positive orientation. The specific implementation method can be referred to in Example 2. However, each sterile syringe 80 used for sampling is always placed in the positive orientation centrifugation sleeve for centrifugation.

[0091] After the first centrifugation, the chromatogram inside the tube is shown below. Figure 7 The first image on the left shows the material to be selected after centrifugation, consisting of plasma, white blood cells, and platelets. In this case, the only way to collect the sample is to first remove the red blood cell layer fluid from the syringe tip before sampling. Some of the red blood cell layer fluid will adhere to the syringe wall during the initial discharge process. Figure 7 (See the second figure on the left in the middle). In this way, subsequent sampling will flow out through the discharge pipeline, resulting in a large amount of red blood cells remaining in the sample, leading to sample impurity.

[0092] Example 4: See Figure 8 The present invention provides a method for preparing P-PRP by inverted secondary centrifugation. The specific implementation method can be referred to in Example 2. However, each sterile syringe 80 used for sampling is always placed upside down in the inverted centrifugation sleeve for centrifugation.

[0093] In this case, after the first centrifugation, the chromatogram inside the tube is shown below. Figure 8The first image on the left shows the materials to be used after centrifugation: plasma layer, white blood cell layer, and platelet layer. In this case, a second sterile syringe is used to extract approximately 40%-45% of the liquid from the top of the first sterile syringe. Then, a second centrifugation is performed using a new sterile syringe, leaving the red blood cell layer inside the first sterile syringe. At this point, the useful liquid adhering to the syringe wall does not affect purity.

[0094] After the second centrifugation, the chromatogram inside the tube is shown below. Figure 8 The third image on the left shows the plasma layer at the top of the sterile syringe nozzle. The liquid to be collected is the L-PRP layer drawn from the sterile syringe after secondary centrifugation; its volume is approximately 20% of the liquid volume inside the syringe after the second centrifugation. In this case, the plasma layer at the syringe nozzle must be discarded first, but this will cause the plasma layer to adhere to the syringe wall, such as... Figure 8 The first image on the right in the middle shows that subsequent sampling will flow out through the discharge pipe, resulting in a significant amount of red blood cells remaining in the sample, leading to sample impurity.

[0095] Traditional centrifugation methods, limited by established thinking, typically employ only one approach, either Example 3 or Example 4, which involves either placing the centrifuge upright or inverted. Most operators wouldn't consider using the method described in Example 2, where the centrifuge is first inverted and then upright.

[0096] P-PRP was extracted using the method described in Example 2, and the components of P-PRP obtained before and after centrifugation were analyzed by comparing whole blood before and after centrifugation.

[0097] The purity of P-PRP depends mainly on the number of platelets (PLT) in the blood sample. If the ratio of platelet concentration to whole blood platelet concentration after centrifugation is greater than 3, i.e., W = PRP(PLT) / whole blood (PLT) > 3, then the P-PRP preparation is successful.

[0098] After centrifugation, the platelets in the extract are activated, and the α-granules on their surface open, releasing a large number of various growth factors. This promotes the regeneration of blood vessels and collagen at the local injection site. Moreover, the activated PRP is mostly gel-like, which can also play a role in local tissue filling.

[0099] Example 5:

[0100] Figures 10-11 This is a sample composition analysis table of Wang XX's whole blood and PRP prepared for laboratory testing.

[0101] This is laboratory test data;

[0102] The sample was taken from Wang XX on March 12, 2025. 10ml of whole blood was collected. The data are shown in Table 1. The platelet (PLT) count before centrifugation was 156. After a second centrifugation, about 2ml of extract liquid was obtained (about 20% of the original whole blood). The data are shown in Table 2. The pre-platelet (PLT) count after centrifugation was 532. The ratio W = 532 / 156 ≈ 3.41 > 3, indicating that the P-PRP was successfully prepared.

[0103] Example 6:

[0104] Figures 12-13 This is the test report for whole blood and PRP samples prepared from Li XX during hospital testing. This is the hospital's trial test data 1:

[0105] The sample was obtained from Li XX on June 23, 2025. 10ml of whole blood was collected. The data are shown in Table 3. The platelet (PLT) count before centrifugation was 296. After a second centrifugation, approximately 2ml of extract liquid was obtained (about 20% of the original whole blood). The data are shown in Table 4. The pre-platelet (PLT) count after centrifugation was 1297. The ratio W = 1297 / 296 ≈ 4.38 > 3, indicating that the P-PRP preparation was successful.

[0106] Example 7:

[0107] Figures 14-15 This is the test report for whole blood and PRP samples prepared for hospital testing of Lei XX. This is hospital trial test data 2:

[0108] The sample was obtained from Lei XX, and the examination time was June 23, 2025. 10ml of whole blood was collected. The data is shown in Table 5. The platelet (PLT) count before centrifugation was 118. After a second centrifugation, about 2ml of extract liquid was obtained (about 20% of the original whole blood). The test data is shown in Table 6. The pre-platelet (PLT) count after centrifugation was 612. The ratio W = 612 / 118 ≈ 5.19 > 3, indicating that the P-PRP was successfully prepared.

[0109] Example 8:

[0110] Figures 16-17 This is the test report for whole blood and PRP samples prepared from Jiang XX during hospital testing. This is test data 3 from the hospital's trial use.

[0111] The sample was taken from Wang XX on June 23, 2025. 10ml of whole blood was collected. The data are shown in Table 7. The platelet (PLT) count before centrifugation was 139. After a second centrifugation, about 2ml of extract liquid was obtained (about 20% of the original whole blood). The data are shown in Table 8. The pre-platelet (PLT) count after centrifugation was 601. The ratio W = 532 / 156 ≈ 3.11 > 3, indicating that the P-PRP was successfully prepared.

[0112] The experimental data from Examples 5-8 demonstrate that the centrifugation system and the centrifugation separation method of inverting and then uprighting the centrifuge used in this invention effectively reduce the risk of contamination and improve the purity of P-PRP preparation. This also avoids the impact of liquid adhering to the walls on purification.

[0113] It should be noted that, in this document, the terms “comprising,” “including,” or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such process, method, article, or apparatus.

[0114] This article uses specific examples to illustrate the principles and implementation methods of the present invention. The above examples are only for the purpose of helping to understand the method and core ideas of the present invention. The above descriptions are only preferred embodiments of the present invention. It should be noted that due to the limitations of textual expression, while there are objectively infinite specific structures, those skilled in the art can make several improvements, modifications, or changes without departing from the principles of the present invention, and can also combine the above technical features in an appropriate manner. These improvements, modifications, changes, or combinations, or the direct application of the inventive concept and technical solution to other situations without modification, should all be considered within the scope of protection of the present invention.

Claims

1. A high-purity P-PRP preparation system, characterized in that, It includes a sterile syringe for sampling, a centrifuge for providing centrifugation, a centrifuge sleeve for use with the centrifuge, and a separator for purifying the sample from the centrifuged sterile syringe; the centrifuge sleeve is detachably filled with the sterile syringe to be centrifuged, and the injection port of the sterile syringe after centrifugation contains the sample component to be used. The separator is a three-way tube, which has an end seal at one end and the other two ends are connected to the sterile syringe after centrifugation and the sterile syringe for extraction, respectively. There are two types of centrifuge tubes: an inverted centrifuge tube used for the first centrifugation and an upright centrifuge tube used for the second centrifugation.

2. The high-purity P-PRP preparation system according to claim 1, characterized in that, After the first centrifugation, the top of the sterile syringe placed upside down in the centrifuge sleeve is connected to the three-way valve. The other opening of the three-way valve is connected to the injection port of another sterile syringe, which is used to extract the upper plasma layer and the middle white blood cell and platelet layer (PRP).

3. The high-purity P-PRP preparation system according to claim 1, characterized in that, After the second centrifugation, the bottom of the sterile syringe placed inside the centrifuge sleeve is connected to a three-way valve. The other opening of the three-way valve is connected to the injection port of another sterile syringe, which is used to extract white blood cells and platelet-rich plasma (PRP) from the lower end.

4. The high-purity P-PRP preparation system according to claim 1, characterized in that, The fixing device on the top inner side of the sealing cap of the inverted centrifuge sleeve is ring-shaped and is matched and coupled to the end seal of the syringe containing the test sample.

5. The high-purity P-PRP preparation system according to claim 1, characterized in that, The bottom of the receiving cavity of the upright centrifuge tube is provided with an inner support platform, and the center of the inner support platform is provided with a support hole, the size of which matches the injection port of the sterile syringe.

6. A method for preparing high-purity P-PRP using the preparation system as described in claims 1-5, characterized in that, Includes the following steps: (1) Sample preparation: Use a sterile syringe to draw whole blood samples to be centrifuged; (2) First centrifugation: After removing the plunger from the end of the sterile syringe, place the sterile syringe upside down into the inverted centrifuge tube, and place the inverted centrifuge tube on the centrifuge for the first centrifugation operation. The centrifugation speed is 150g-210g and the duration is 10-15min. After centrifugation, the liquid in the sterile syringe will separate into 3 layers. (3) Purification and sampling: Take another sterile syringe and connect one end of the three-way connector, close the other end of the three-way connector, and connect the third end of the three-way connector to the injection port of the sterile syringe after the first centrifugation; at this time, the injection port of the sterile syringe after centrifugation is facing upward, and the upper layer of liquid is drawn out by taking another sterile syringe, the volume of which is about 40%-45% of the whole blood volume. (4) Second centrifugation: Place the sterile syringe containing the upper liquid extracted in (3) into the upright centrifuge tube, and place the upright centrifuge tube on the centrifuge for a second centrifugation operation. The centrifugation speed is 240g-350g and the duration is 12-18min. (5) Final sampling: The upper liquid is sampled again using the method in (3). At this time, the injection port of the sterile syringe after centrifugation is facing down. The lower liquid is extracted by taking another sterile syringe to obtain the final pure L-PRP. Its volume is about 20% of the liquid volume in the sterile syringe after the second centrifugation.

7. The method for preparing high-purity P-PRP according to claim 6, characterized in that, When purifying and re-sampling in step (3), when pulling the plunger of the new sterile syringe, stop drawing liquid when the red blood cell layer is clearly separated and the bottom plunger plane of the sterile syringe from the first centrifugation is close. The volume drawn is about 40%-45% of the whole blood volume.

8. The method for preparing high-purity P-PRP according to claim 6, characterized in that, When performing the final sampling in step (5), when pulling the plunger of the new sterile syringe, the liquid is drawn until the plasma layer is clearly separated and the bottom plunger plane of the sterile syringe after the second centrifugation is close to the plane. The volume drawn is about 20% of the liquid volume in the sterile syringe after the second centrifugation.

9. The method for preparing high-purity P-PRP according to claim 6, characterized in that, In step (2), placing the sterile syringe upside down into the inverted centrifuge tube means that the plunger of the sterile syringe is facing down and the injection port is facing up.

10. The method for preparing high-purity P-PRP according to claim 6, characterized in that, In step (4), placing the sterile syringe upright into the upright centrifuge tube means that the plunger of the sterile syringe is facing upward and the injection port is facing downward.

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