Centrifugal treatment method for multiple times of centrifugation

Abstract

The invention discloses a multiple-centrifugation centrifugal treatment method, and relates to the technical field of platelet-rich plasma preparation, and the multiple-centrifugation centrifugal treatment method comprises the following operation steps: S1, model establishment; s2, weighing and classifying the blood samples; s3, classifying and centrifuging; s4, standing in a simulated human body temperature environment; and S5, extracting platelet-rich blood. According to the centrifugal treatment method of multiple centrifugation, vacuum blood collection tubes containing different blood sample volumes are classified by establishing an effect influence model of primary centrifugation and secondary centrifugation, and the vacuum blood collection tubes in the same volume interval are put into a centrifugal machine to be subjected to primary centrifugation and secondary centrifugation; and on the basis of the volume parameters, the centrifugal parameters with the best matching values are automatically selected by the model to centrifuge the vacuum blood collection tubes containing the blood samples, so that the time required by centrifugation is shortened as much as possible while the effect after centrifugation is ensured, and the operation parameters of the centrifugal machine are matched with the blood samples in all the volume intervals.

Description

Technical Field

[0001] The invention relates to the technical field of platelet-rich plasma preparation, in particular to a centrifugal treatment method of multiple centrifugation. Background Art

[0002] Platelet-rich plasma (PRP) is rich in growth factors, cytokines, antimicrobial peptides and other bioactive substances. It can promote cell proliferation, differentiation, matrix synthesis, tissue regeneration and repair, and plays an important role in regenerative medicine.

[0003] The existing preparation of PRP usually adopts the double centrifugation method, which includes performing two centrifugation steps after drawing venous blood. The first centrifugation is low-speed centrifugation to separate the blood into three layers, and the second centrifugation is high-speed centrifugation to concentrate platelets. However, it is difficult to accurately control the separation each time a blood sample is drawn, resulting in it being difficult to set the centrifugation speed and time that best matches the blood sample each time the blood is centrifuged. Ultimately, in order to ensure the centrifugation effect, it is often necessary to extend the centrifugation time, thereby extending the preparation time.

[0004] Therefore, in view of this, the existing structure and defects are studied and improved, and a centrifugal treatment method of multiple centrifugation is proposed. Summary of the invention

[0005] In view of the deficiencies in the prior art, the present invention provides a centrifugal treatment method of multiple centrifugation to solve the problems raised in the above background technology.

[0006] To achieve the above object, the present invention is implemented by the following technical scheme: a centrifugal treatment method of multiple centrifugation, the centrifugal treatment method of multiple centrifugation comprising the following operating steps:

[0007] S1. Model establishment:

[0008] Whole blood is collected from the patient's vein multiple times into vacuum blood collection tubes containing anticoagulants, and each portion of the blood sample is centrifuged once and twice using a centrifuge, and each parameter is recorded;

[0009] Then, the above parameters are used to establish a model of the effect of the centrifuge speed on the centrifugation of each component of venous blood, thereby establishing the centrifugation parameters that best match each component of blood sample when it is separated, and based on the effect after centrifugation, the blood sample volume with similar effect parameters is divided into the same interval;

[0010] S2. Blood sample weighing and classification:

[0011] When the model in the above steps is officially put into use, whole blood is first collected from the patient's vein into a vacuum blood collection tube containing an anticoagulant, and the volume of the vacuum blood collection tube after the blood sample is collected is weighed, and each vacuum blood collection tube is divided into different intervals based on the weighing result;

[0012] S3, classification centrifugation:

[0013] The vacuum blood collection tubes in the same interval are placed in a centrifuge for one centrifugation, and the blood sample volume parameters of the interval are input into the model in advance. The model automatically selects the best matching centrifugation parameters based on the blood sample volume of the interval. When the blood sample is centrifuged once, a layering liquid is added to the blood sample. The volume ratio of the blood sample volume to the layering liquid is 1:1 or 2:1. The layering liquid is a lymphocyte separation liquid. The effects of various amounts of the lymphocyte separation liquid added on the centrifugation effect of the blood sample are also input into the model and simulated.

[0014] The lymphocyte separation fluid comprises the following ingredients by volume:

[0015] Ficoll: 10%; Sucrose: 9%; Sodium chloride (NaCl): 0.9%; Phosphate buffer (PBS): appropriate amount, the specific amount only needs to adjust the pH value to between 7.2-7.4;

[0016] Among them, Ficoll is a high molecular weight sugar substance;

[0017] The steps for preparing lymphocyte separation medium are as follows:

[0018] 1) Add Ficoll to deionized water and stir until completely dissolved;

[0019] 2) Add sucrose and continue stirring until completely dissolved;

[0020] 3) Add sodium chloride and stir until completely dissolved;

[0021] 4) Use phosphate buffer to adjust the pH value of the solution to 7.2-7.4;

[0022] 5) Use deionized water to make the solution to the required volume;

[0023] Similarly, when the vacuum blood collection tube in the same interval is centrifuged twice, the model automatically selects the centrifugation parameters that best match the blood sample volume parameters in the interval;

[0024] S4, standing still in an environment simulating human body temperature;

[0025] Simulate the human body temperature, place the centrifuge liquid for 2-3 hours at 37℃±1℃, and then centrifuge it for the second time;

[0026] S5. Extraction of platelet-rich blood:

[0027] After the blood sample is centrifuged twice, the plasma in the lower layer is extracted with a syringe, and depending on the needs, an activator is used to activate the platelet-rich blood to release the growth factors in the platelets.

[0028] Furthermore, in step S1, the amount of whole blood collected each time is different, and the collection range is 10-60 mL, and the recorded parameters include but are not limited to the blood sample volume, centrifugal speed and time of the centrifuge, temperature parameters and effect parameters after centrifugation.

[0029] Furthermore, in the step S1, when the centrifuge centrifuges the blood sample in the range of 10-60 mL once, the centrifugal speed is 1000-1500 rpm and the time is 5-10 minutes.

[0030] Furthermore, in step S1, when the centrifuge performs a second centrifugation on the plasma taken out after the first centrifugation, the centrifugation speed is 3000-4000 rpm and the time is 10-15 minutes.

[0031] Furthermore, in step S2, the specific operation of weighing the blood sample volume in each vacuum blood collection tube is as follows: first, each vacuum blood collection tube containing the blood sample is weighed to obtain the blood weight, and the blood density is 1.05 grams per milliliter, so the blood sample volume can be obtained by dividing the blood weight by 1.05, and the unit is milliliter.

[0032] Furthermore, in step S3, the centrifugation parameters that match the blood sample volume include but are not limited to centrifugation speed, centrifugation time, and temperature parameters.

[0033] Furthermore, in step S3, the centrifugation is performed at a low speed, and the blood sample is divided into three layers after centrifugation, the bottom layer is red blood cells, the middle thin layer is white blood cells and platelets, and the upper layer is plasma including platelets.

[0034] Furthermore, in step S3, during the second centrifugation, the plasma after the first centrifugation is taken out and the plasma is separated into two layers at a better centrifugal speed, the upper layer being the platelet-poor plasma and the lower layer being the platelet-rich plasma.

[0035] Furthermore, in step S5, the plasma in the lower layer extracted by the syringe should be prevented from mixing with the platelet-poor plasma in the upper layer.

[0036] Furthermore, in step S5, the activator is calcium chloride or thrombin.

[0037] The present invention provides a centrifugal treatment method with multiple centrifugations, which has the following beneficial effects:

[0038] 1. The centrifugal processing method of multiple centrifugation classifies vacuum blood collection tubes containing different blood sample volumes by establishing a model for the effects of primary and secondary centrifugation, and places vacuum blood collection tubes in the same volume range into a centrifuge for primary and secondary centrifugation. The model automatically selects the centrifugation parameters with the best matching values ​​based on the volume parameters to centrifuge the vacuum blood collection tubes containing the blood samples, thereby shortening the centrifugation time as much as possible while ensuring the effect after centrifugation, so that the centrifuge operating parameters match the blood samples in each volume range. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 The present invention is a schematic flow diagram of a centrifugal treatment method of multiple centrifugation. DETAILED DESCRIPTION

[0040] The following embodiments of the present invention are described in further detail in conjunction with the accompanying drawings and examples. The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention.

[0041] like Figure 1 As shown, the present invention provides a technical solution: a centrifugal treatment method of multiple centrifugation, the centrifugal treatment method of multiple centrifugation comprises the following operating steps:

[0042] S1. Model establishment:

[0043] Whole blood is collected from the patient's vein multiple times into vacuum blood collection tubes containing anticoagulants, and each portion of the blood sample is centrifuged once and twice using a centrifuge, and each parameter is recorded;

[0044] Then, the above parameters are used to establish a model of the effect of the centrifuge speed on the centrifugation of each component of venous blood, thereby establishing the centrifugation parameters that best match each component of blood sample when it is separated, and based on the effect after centrifugation, the blood sample volume with similar effect parameters is divided into the same interval;

[0045] The amount of whole blood collected each time is different, and the collection range is 10-60mL. The recorded parameters include but are not limited to the amount of blood sample, centrifugal speed and time of the centrifuge, temperature parameters and effect parameters after centrifugation. When the centrifuge centrifuges the blood sample in the range of 10-60mL once, the centrifugal speed is 1000-1500rpm, and the time is 5-10 minutes. When the centrifuge centrifuges the plasma taken out after the first centrifugation for the second time, the centrifugal speed is 3000-4000rpm, and the time is 10-15 minutes.

[0046] S2. Blood sample weighing and classification:

[0047] When the model in the above steps is officially put into use, whole blood is first collected from the patient's vein into a vacuum blood collection tube containing an anticoagulant, and the volume of the vacuum blood collection tube after the blood sample is collected is weighed, and each vacuum blood collection tube is divided into different intervals based on the weighing result. The specific operation of weighing the blood sample volume in each vacuum blood collection tube is as follows: first weigh each vacuum blood collection tube containing the blood sample to obtain the blood weight, and the density of blood is 1.05 grams per milliliter, thereby dividing the blood weight by 1.05 to obtain the blood sample volume in milliliters;

[0048] S3, classification centrifugation:

[0049] The vacuum blood collection tubes in the same interval are placed in a centrifuge for centrifugation, and the blood sample volume parameters of the interval are input into the model in advance. The model automatically selects the centrifugation parameters that best match the blood sample volume in the interval, and the centrifugation parameters that match the blood sample volume include but are not limited to centrifugation speed, centrifugation time, and temperature parameters;

[0050] Similarly, when the vacuum blood collection tube in the same interval is centrifuged twice, the model automatically selects the centrifugation parameters that best match the blood sample volume parameters in the interval;

[0051] Among them, the first centrifugation is low-speed centrifugation, and the blood sample is divided into three layers after centrifugation. The bottom layer is red blood cells, the middle thin layer is white blood cells and platelets, and the upper layer is plasma including platelets. The second centrifugation is to take out the plasma after the first centrifugation and separate the plasma into two layers at a better centrifugal speed. The upper layer is platelet-poor plasma, and the lower layer is platelet-rich plasma.

[0052] When the blood sample is centrifuged once, a layering liquid is added to the blood sample, and the volume ratio of the blood sample volume to the layering liquid is 1:1 or 2:1. The layering liquid is a lymphocyte separation liquid. The effect of each amount of the lymphocyte separation liquid added on the centrifugation effect of the blood sample is also input into the model and simulated.

[0053] The lymphocyte separation fluid comprises the following ingredients by volume:

[0054] Ficoll: 10%; Sucrose: 9%; Sodium chloride (NaCl): 0.9%; Phosphate buffer (PBS): appropriate amount, the specific amount only needs to adjust the pH value to between 7.2-7.4;

[0055] Among them, Ficoll is a high molecular weight sugar substance;

[0056] The steps for preparing lymphocyte separation medium are as follows:

[0057] 1) Add Ficoll to deionized water and stir until completely dissolved;

[0058] 2) Add sucrose and continue stirring until completely dissolved;

[0059] 3) Add sodium chloride and stir until completely dissolved;

[0060] 4) Use phosphate buffer to adjust the pH value of the solution to 7.2-7.4;

[0061] 5) Use deionized water to make the solution to the required volume;

[0062] Based on the above description, lymphocyte separation fluid is added to the blood sample during the secondary centrifugation. Since the density of the lymphocyte separation fluid is between that of red blood cells and platelets, the centrifugal stratification effect of the blood can be improved to speed up the stratification of the blood sample.

[0063] S4, standing still in an environment simulating human body temperature;

[0064] Simulate the human body temperature, place the centrifuge liquid for 2-3 hours at 37℃±1℃, and then centrifuge it for the second time;

[0065] S5. Extraction of platelet-rich blood:

[0066] After the blood sample is centrifuged twice, the plasma in the lower layer is extracted with a syringe, and the activator is selected according to the needs to activate the platelet-rich blood to release the growth factors in the platelets. The plasma in the lower layer extracted with the syringe should be prevented from mixing with the platelet-poor plasma in the upper layer, and the activator is calcium chloride or thrombin;

[0067] Based on the above description, the present invention classifies vacuum blood collection tubes containing different blood sample volumes by establishing a model that affects the effects of primary and secondary centrifugation, and places vacuum blood collection tubes in the same volume range into a centrifuge for primary and secondary centrifugation. The model automatically selects the centrifugation parameters with the best matching values ​​based on the volume parameters to centrifuge the vacuum blood collection tubes containing the blood samples, thereby ensuring the effect after centrifugation while shortening the centrifugation time as much as possible, so that the centrifuge operating parameters match the blood samples in each volume range.

[0068] In summary, when the centrifugal treatment method of multiple centrifugation is used, whole blood is first collected from the patient's vein multiple times into a vacuum blood collection tube containing an anticoagulant, and each component of the blood sample is centrifuged once and twice using a centrifuge, and each parameter is recorded, and then the above parameters are used to establish a model of the effect of the centrifuge speed on the centrifugation of each component of venous blood, thereby establishing the centrifugal parameters that best match each component of the blood sample when it is separated, and based on the effect after centrifugation, the blood sample amount with similar effect parameters is divided into the same interval;

[0069] When the model in the above steps is officially put into use, whole blood is first collected from the patient's vein into a vacuum blood collection tube containing an anticoagulant, and the volume of the vacuum blood collection tube after the blood sample is collected is weighed, and each vacuum blood collection tube is divided into different intervals based on the weighing result. The specific operation of weighing the blood sample volume in each vacuum blood collection tube is as follows: first weigh each vacuum blood collection tube containing the blood sample to obtain the blood weight, and the density of blood is 1.05 grams per milliliter, thereby dividing the blood weight by 1.05 to obtain the blood sample volume in milliliters;

[0070] The vacuum blood collection tubes in the same interval are placed in a centrifuge for a centrifugation, and the blood sample volume parameters of the interval are input into the model in advance. The model automatically selects the best matching centrifugation parameters based on the blood sample volume of the interval. Similarly, when the vacuum blood collection tubes in the same interval are centrifuged for a second time, the model automatically selects the best matching centrifugation parameters based on the blood sample volume parameters of the interval.

[0071] Among them, the first centrifugation is low-speed centrifugation, and the blood sample is divided into three layers after centrifugation. The bottom layer is red blood cells, the middle thin layer is white blood cells and platelets, and the upper layer is plasma including platelets. The second centrifugation is to take out the plasma after the first centrifugation and separate the plasma into two layers at a better centrifugal speed. The upper layer is platelet-poor plasma, and the lower layer is platelet-rich plasma.

[0072] After the blood sample is centrifuged twice, the plasma in the lower layer is extracted with a syringe, and depending on the needs, an activator is used to activate the platelet-rich blood to release the growth factors in the platelets.

[0073] The embodiments of the present invention are given for the purpose of illustration and description, and are not intended to be exhaustive or to limit the invention to the disclosed forms. Many modifications and variations will be apparent to those of ordinary skill in the art. The embodiments are selected and described in order to better illustrate the principles and practical applications of the present invention and to enable those of ordinary skill in the art to understand the present invention and thereby design various embodiments with various modifications suitable for specific uses.

Claims

1. A centrifugal treatment method with multiple centrifugations, characterized in that: The centrifugal treatment method of multiple centrifugation comprises the following steps: S1. Model establishment: Whole blood is collected from the patient's vein multiple times into vacuum blood collection tubes containing anticoagulants, and each portion of the blood sample is centrifuged once and twice using a centrifuge, and each parameter is recorded; Then, the above parameters are used to establish a model of the effect of the centrifuge speed on the centrifugation of each component of venous blood, thereby establishing the centrifugation parameters that best match each component of blood sample when it is separated, and based on the effect after centrifugation, the blood sample volume with similar effect parameters is divided into the same interval; S2. Blood sample weighing and classification: When the model in the above steps is officially put into use, whole blood is first collected from the patient's vein into a vacuum blood collection tube containing an anticoagulant, and the volume of the vacuum blood collection tube after the blood sample is collected is weighed, and each vacuum blood collection tube is divided into different intervals based on the weighing result; S3, classification centrifugation: The vacuum blood collection tubes in the same interval are placed in a centrifuge for one centrifugation, and the blood sample volume parameters of the interval are input into the model in advance. The model automatically selects the best matching centrifugation parameters based on the blood sample volume of the interval. When the blood sample is centrifuged once, a layering liquid is added to the blood sample. The volume ratio of the blood sample volume to the layering liquid is 1:1 or 2:

1. The layering liquid is a lymphocyte separation liquid. The effects of various amounts of the lymphocyte separation liquid added on the centrifugation effect of the blood sample are also input into the model and simulated. The lymphocyte separation fluid comprises the following ingredients by volume: Ficoll: 10%; Sucrose: 9%; Sodium chloride (NaCl): 0.9%; Phosphate buffer (PBS): The specific dosage only needs to adjust the pH value to between 7.2-7.4; Among them, Ficoll is a high molecular weight sugar substance; The steps for preparing lymphocyte separation medium are as follows: 1) Add Ficoll to deionized water and stir until completely dissolved; 2) Add sucrose and continue stirring until completely dissolved; 3) Add sodium chloride and stir until completely dissolved; 4) Use phosphate buffer to adjust the pH value of the solution to 7.2-7.4; 5) Use deionized water to make the solution to the required volume; Similarly, when the vacuum blood collection tube in the same interval is centrifuged twice, the model automatically selects the centrifugation parameters that best match the blood sample volume parameters in the interval; S4, standing still in an environment simulating human body temperature; Simulate the human body temperature, place the centrifuge liquid for 2-3 hours at 37℃±1℃, and then centrifuge it for the second time; S5. Extraction of platelet-rich blood: After the blood sample is centrifuged twice, the plasma in the lower layer is extracted with a syringe, and depending on the needs, an activator is used to activate the platelet-rich blood to release the growth factors in the platelets.

2. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In step S1, the amount of whole blood collected each time is different, and the collection range is 10-60 mL, and the recorded parameters include but are not limited to the blood sample volume, centrifugal speed and time of the centrifuge, temperature parameters and effect parameters after centrifugation.

3. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In the step S1, when the centrifuge centrifuges the blood sample in the range of 10-60 mL, the centrifugal speed is 1000-1500 rpm and the time is 5-10 minutes.

4. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In the step S1, when the centrifuge performs a second centrifugation on the plasma taken out after the first centrifugation, the centrifugation speed is 3000-4000 rpm and the time is 10-15 minutes.

5. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In step S2, the specific operation of weighing the blood sample volume in each vacuum blood collection tube is as follows: first, each vacuum blood collection tube containing the blood sample is weighed to obtain the blood weight, and the blood density is 1.05 grams per milliliter, so the blood sample volume can be obtained by dividing the blood weight by 1.05, and the unit is milliliter.

6. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In step S3, the centrifugation parameters that match the blood sample volume include but are not limited to centrifugation speed, centrifugation time, and temperature parameters.

7. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In step S3, the first centrifugation is low-speed centrifugation. After centrifugation, the blood sample is divided into three layers, the bottom layer is red blood cells, the middle thin layer is white blood cells and platelets, and the upper layer is plasma including platelets.

8. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In step S3, during the second centrifugation, the plasma after the first centrifugation is taken out and the plasma is separated into two layers at a better centrifugal speed, the upper layer is the platelet-poor plasma, and the lower layer is the platelet-rich plasma.

9. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In step S5, the plasma in the lower layer should be extracted by the syringe to avoid mixing with the platelet-poor plasma in the upper layer.

10. The centrifugal treatment method of multiple centrifugation according to claim 1, characterized in that: In step S5, the activator is calcium chloride or thrombin.

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