Method of obtaining autogenic activated platelet-enriched plasma for dentistry

FIELD: medicine.

SUBSTANCE: sampling of blood from patient's ulnar vein with first and second syringe-test-tubes is carried out. First syringe-test-tube is centrifuged with acceleration 250 g for 10 minutes. Formed plasma and coagulated blood in second syringe-test-tube are centrifuged with acceleration 1000 g for 10 minutes. Platelet-poor plasma is separated from platelet-enriched plasma. Activator of platelet-enriched plasma is prepared from supernatant fluid after centrifugation of coagulated blood and platelet-poor plasma, with weight ratio in fractions: 1:1, and 10% calcium chloride solution. The latter - in drop manner, mixing until required mixture concentration is obtained. Platelet-enriched plasma is mixed with activator with weight ratio in fractions 1:3 respectively.

EFFECT: application of method allows to realise more complete isolation of platelets from sampled blood, which increases efficiency, physiologicity and safety of obtained plasma.

2 cl

 

The invention relates to the field of medicine, as well as drugs used for medical purposes containing materials from mammals, in particular containing components of human blood, and can be used in dentistry in the surgical treatment.

A method of obtaining autologous activated platelet-rich plasma for dentistry, in accordance with which withdraws blood from the cubital vein in vacuum plastic tubes with a volume of 9 ml, within which already contains the activator of clotting serum (plasma). In a laboratory centrifuge firm "Heltich" one-step centrifuged selected blood in the test tube for 12 minutes at a speed of 2600 rpm the Final product is a platelet-fibrin gel (Mourzenko, Avessi "Overview of equipment and methods of obtaining autologous platelet-rich plasma in dentistry" // journal "New in dentistry", 2003, №7, p.47).

The disadvantage of this method is that it is used for activation of the platelet activators of animal origin or artificially synthesized, which reduces the physiology of way. Furthermore, the method does not allow to retrieve from the selected material, the maximum number of platelets, i.e. the maximum use of the selected material, because of the moves goes not only sediment red blood cells, but also platelet-poor plasma. Besides this fact reduces the environmental well-known method.

The closest to the invention is a method of obtaining autologous activated platelet-rich plasma for dentistry, including drawing blood from the cubital vein by syringe-tube with a volume of 9 ml, which contains anticoagulant blood - citrate. Selected blood in the test tube is centrifuged in two stages in a laboratory centrifuge Heraeus Labofuge 300". In the first phase, centrifuged for 12 minutes at a speed of 4000 rpm Then the formed plasma supernatant layer from the first syringe tubes trying to enter into another syringe tube without citrate and processed in a centrifuge for 15 minutes at a speed of 3600 rpm After separation of platelet-poor plasma supernatant layer, get the final product - platelet-rich plasma in the amount of 0.3 ml of Activated platelet-rich plasma preparation "Cerasorb" - artificially synthesized beta-tricalcium phosphate. To do this, "Cerasorb" pre-mixed with blood taken from the wound, and then added to the mixture of platelet-rich plasma (Mourzenko, Avessi "Overview of equipment and methods of obtaining autologous platelet-rich plasma in dentistry" // journal "New in dentistry", 2003, №7, p.46).

The lack Izv the STN method is, first of all, in a small quantity of the finished product: from 9 ml of blood is obtained 0.3 ml of platelet-rich plasma, despite dvuhetapnogo processing blood by centrifugation, which reduces the efficiency of this method. This is due to the fact that in the known method at each stage uses a high speed centrifuge: 4000 and 3600 rpm, which is not consistently separate the components of blood with regard to their weight and the proportion of platelets goes unused residue. In addition, the waste goes and platelet-poor plasma. In the known method does not provide the maximum use of selected to receive platelet material, which, in turn, does not allow to obtain the maximum recovery of platelets from selected blood. Moreover, as in the known method in the waste leaving as a residue of red blood cells, formed after the first centrifugation, and platelet-poor plasma, it reduces the environmental well-known method.

In addition, in the preparation of the activator in the known method using the blood from the wound of the patient, which is not secure against the possibility of entering of an infection, as with the blood in the activator can get particles of dead tissue, pus, etc. are, as well as the use of artificially synthesized activate the RA clotting serum reduces the physiology of the method.

Thus, identified as a result of a patent search methods of obtaining autologous activated platelet-rich plasma for dentistry, similar and closest to the stated, when the implementation does not ensure the achievement of the technical result consists in the possibility of a more complete removal of platelets selected from blood, and consequently, to increase the efficiency of the method, in increasing physiology, less secure, more environmentally friendly way, due to the possibility of reducing the amount of waste after receipt of the final product according to the method.

The claimed method of obtaining autologous activated platelet-rich plasma for dentistry solves the problem of creating an appropriate way, which ensures the achievement of the technical result consists in the possibility of a more complete removal of platelets selected from blood, and consequently, to increase the efficiency of the method, in increasing physiology, less secure, more environmentally friendly way due to the possibility of reducing the amount of waste after receipt of the final product according to the method.

The invention consists in that in the method of obtaining autologous activated enriched trombi the Tami plasma for dentistry, includes drawing blood from the cubital vein by syringe-tube, centrifugation selected blood in the test tube, transfer the plasma (supernatant fraction separated after centrifugeuse,in an empty test tube, centrifuging the newly selected product, removal of the tubes formed nadeshiko (plasma poor in platelets,activation of the remaining in vitro platelet-rich plasma, what is new is the fact that additionally carry out blood sampling the second syringe-tube in number, constituting half of the total selected blood first syringe is a tube that is left in the upright position for blood clotting, the first syringe tube with selected blood is placed in a centrifuge and centrifuged at acceleration of 250 g for 10 minutes, after which the formed plasma is transferred into an empty test tube and centrifuged with an acceleration of 1000 g for 10 minutes, then remove the tube from the centrifuge, and then formed after centrifugation the plasma poor in platelets, separated from platelet-rich plasma and transferred to an empty tube, then prepare the activator of platelet-rich plasma, which make sure that the second syringe-tube blood clotted, after which the test tube with coagulating blood is placed in a centrifuge and centrifuged with accelerated is eating 1000 g for 10 minutes, then the test tube is then removed from the centrifuge and transfer the formed after centrifugation gross blood supernatant into a second empty tube, which then adds the plasma poor in platelets, the mass ratio at 1:1, and mix thoroughly, then drip, with stirring, to impose a 10% solution of calcium chloride to obtain the desired consistency of the mixture, mix thoroughly, after which platelet-rich plasma activated, for this is mixed with the received activator in the ratio of the masses in the 1:3 respectively. In addition, tubes with plasma and with coagulating blood centrifuged at the same time, having first ascertained that in the second syringe-tube blood clotted.

The technical result is achieved in the following way. Signs of claims: includes drawing blood from the cubital vein by syringe-tube, centrifugation selected blood in the test tube, transfer the plasma (supernatant fraction separated after the first centrifugeuse,in an empty test tube, centrifuging the newly selected product, removal of the tubes formed nadasdy, activating the remaining in vitro platelet-rich plasma, are key in obtaining platelet-rich plasma two-step method. This is rsnake ensure the operability of the inventive method, consequently, ensure the achievement of the stated technical result.

It is known that red blood cells are significantly heavier than platelets. The inventors have empirically found that to get in the sediment after the first centrifugation of the maximum number of red blood cells enough acceleration of 250 g. The optimal time of centrifugation, after which the residue appears the maximum number of red blood cells, also derived empirically by the authors of the claimed method and is 10 minutes. Since in the proposed method the first syringe tube with selected blood is placed in a centrifuge and centrifuged at acceleration of 250 g, the precipitate fall red blood cells as more severe, and platelets as more light remain in the supernatant layer in the plasma. Mode selection centrifugation at the first stage provides the maximum separation of the red cells from the platelets, which contributes to a more complete extraction of platelets from selected blood and increases the efficiency of the claimed method.

Signs of the formula: "...after which the formed plasma is transferred into an empty test tube and centrifuged..., ... formed after centrifugation the plasma poor in platelets, separated from platelet-rich plasma..." provide the opportunity for the implementation of the second final stage of obtaining platelet-rich plasma, and therefore, provide the ability to perform the claimed process and the achievement of the claimed technical result. The centrifugation parameters at the second stage: with the acceleration of 1000 g for 10 minutes, obtained by the authors of the claimed method empirically and are optimal for obtaining the maximum number of platelets in platelet-rich plasma, which increases the efficiency of the inventive method.

As experience has shown, through the optimal selection of modes centrifugation used in the claimed method of receiving a two-step centrifugation provides a more complete removal of platelets from selected blood, compared with the prototype, namely from 9 ml of blood is obtained 1.5 ml of platelet-rich plasma, while in the prototype from the same volume of selected material was obtained 0.3 ml of platelet-rich plasma. As a result, the optimal selection modes centrifugation provides the more complete recovery of platelets selected from blood, which increases the efficiency of the inventive method.

In the proposed method for the activation of platelet-rich plasma use the activator, which is prepared from blood of the same patient, namely: to activate the platelet-rich plasma use the fibrinogen contained in the blood taken from this is E. the patient. This allows you to refuse the use to activate platelet-rich plasma, bovine thrombin or artificially synthesized activators. In addition, for cooking use only products centrifugation, which is sourced from the sterile test tubes. As a result, in the proposed method is improved in comparison with the prototype, the security method, and therefore, increases and physiology of the method.

For the manufacture of activator additionally carry out blood sampling the second syringe-tube in number, constituting half of the total selected blood first syringe-tube. The volume of blood required to obtain a sufficient amount of fibrinogen, determined by the inventors empirically and is optimal. This blood volume provides further obtaining fibrinogen in an amount sufficient to activate all volume of platelet-rich plasma, obtained from selected material - blood of the patient, in accordance with the claimed method that improves the efficiency of the inventive method. Due to the fact that the tube of blood to obtain activator leave for blood clotting, and in a vertical position, resulting in the coagulation of blood in the lower part of the tube erythrocytes accumulate as most Aulie. While the supernatant is concentrated fibrinogen as easier. For the final separation from fibrin, erythrocytes, and other heavier components of the blood previously convinced that the second syringe-tube blood clotted, after which the test tube with coagulating blood is placed in a centrifuge and centrifuged with an acceleration of 1000 g for 10 minutes. Mode centrifugation obtained by the authors empirically and is optimal for extraction from the blood of the patient the maximum amount of fibrinogen, which increases the efficiency of the inventive method. After centrifugation the second test tube is removed from the centrifuge and transfer the supernatant, which was formed after centrifugation thickening of the blood, the second, empty test tube, thereby separating from the resulting sludge supernatant containing fibrinogen. However, due to the proposed mode centrifugation, the supernatant fibrinogen contained in the maximum number.

To improve the efficiency of the method, by reducing the preparation time of the activator tube with plasma and with coagulating blood centrifuged at the same time, having first ascertained that in the second syringe-tube blood clotted.

Poor platelet plasma also contains the howling structure of fibrinogen. Due to the fact that in the proposed method platelet-poor plasma added to the supernatant, obtained after centrifugation of the clotted blood, the total number of fibrinogen is increased without increasing the total volume of blood withdrawn from the patient. The increase in the concentration of fibrinogen provides a complete, reliable activation of platelet-rich plasma. This provides the physiology of way. In addition, the use of platelet-poor plasma reduces the inventive method, in comparison with the prototype, the total amount of waste after processing the blood, which increases the sustainability of the claimed method.

Adding platelet-poor plasma in the supernatant obtained after centrifugation of the clotted blood, the mass ratio at 1:1 is the optimum obtained by the inventors empirically and provides the content of fibrinogen in the obtained liquid fraction of the amount needed to activate full platelet-rich plasma, obtained by the claimed method. This thorough mixing of the components ensures uniform distribution of fibrinogen in a liquid medium. The result is improved efficiency of the claimed method.

Introduction in the mixture of 10% solution of calcium chloride C which takes the reaction of polymerization of fibrinogen, as a result of which forms a fibrin clot. A drip of 10% solution of calcium chloride under stirring until you get the desired consistency of the mixture provides the ability to control the consistency of the activator (the density of the fibrin clot), which, in turn, helps to obtain the desired density of the clot-activated platelet-rich plasma. By mixing platelet-rich plasma obtained by the activator, the reaction begins platelet degranulation, i.e. their activation. Mixing platelet-rich plasma obtained by the activator in the ratio of 1:3 respectively provides the activation of the platelet-rich plasma in full, which increases the efficiency of the inventive method.

In addition, in the preparation of the activator of platelets contained in platelet-poor plasma, activated after activation of platelet-rich plasma complement the total number of activated platelets in the final product. In the claimed method provides the maximum recovery of platelets selected from the patient's blood, which increases the efficiency of the inventive method.

In the result of the execution of the claimed method the resulting output product for dentistry: autologous activated platelet-rich plasmas is in the form of a bunch of activated platelets with a high content of fibrin, completely made from the patient's blood. When this activated platelets are growth factors that are involved in regulating the growth and functioning of cells in the regeneration of periodontal tissue and fibrin is involved in the process of osteoconductive. In addition, since platelet-poor plasma, and in the supernatant after centrifugation of the clotted blood leukocytes are present, they are present in the final product, ensuring its immediate use topical antibacterial action. The main advantage of the method lies in the fact that activated platelet-rich plasma is prepared completely from the blood of the patient, which eliminates the risk of the introduction into the organism of the patient is incompatible with the blood of the patient biological bodies and minimizes the risk of infectious contamination. This increases the safety, physiology and effectiveness of the claimed method. The possibility of using platelet-poor plasma for the preparation of the activator reduces the amount of waste in obtaining platelet-rich plasma, which increases the sustainability of the claimed method.

Thus, from the above it follows that the claimed method of obtaining autologous activated platelet-rich plasma for dentistry domestic which ensures the achievement of the technical result consisting in the possibility of a more complete removal of platelets from selected blood and, consequently, to increase the efficiency of the method, in increasing physiology, less secure, more environmentally friendly way due to the possibility of reducing the amount of waste after receipt of the final product according to the method.

The claimed method of obtaining autologous activated platelet-rich plasma for dentistry performed as follows. Perform blood from the cubital vein of the patient first syringe-tube. Additionally carry out blood sampling the second syringe-tube in number, constituting half of the total selected blood first syringe-tube. The second syringe tube leave in a vertical position for blood clotting. The first syringe is a tube of blood is placed in a centrifuge and centrifuged at acceleration of 250 g for 10 minutes, after which the formed plasma is transferred to an empty tube. Make sure that the second syringe-tube blood clotted, after which the tubes with plasma and with coagulating blood is placed in a centrifuge and centrifuged with an acceleration of 1000 g for 10 minutes. Then the tube is removed from the centrifuge, and then formed after centrifugation plasma platelet-poor plasma is separated from platelet-rich plasma is transferred to an empty tube. After that, prepare the activator of platelet-rich plasma, which formed after centrifugation gross blood supernatant is transferred into a second empty tube, which then adds the plasma poor in platelets, the mass ratio at 1:1, and mix thoroughly, then drip, with stirring, to impose a 10% solution of calcium chloride to obtain the desired consistency of the mixture, mix thoroughly. After which platelet-rich plasma activated, for this purpose it is mixed with the received activator of the mass ratio to 1:3, respectively.

In the process used laboratory centrifuge, providing acceleration is not less than 1000 g. To draw blood from the cubital vein of the patient used the standard system with butterfly. All tubes in the method used sterile. For the main blood used syringe tube with a volume of 9 ml type EDTA/K2-Gel; for more blood used syringe tube type Serum Gel S/4,7; the test tube and the third tube with measuring marks and tightly closing the tube; the tube is fourth with a tight closing type tube Vacutainer volume 7 ml; the fifth test tube with tight-closing type tube Vacutainer volume 10 ml fluid was carried out by means of a sterile Pasteur pipet the K.

It is recommended for optimal use of centrifuges and to minimize vibration, which is important for complete sedimentation of the particles of the blood, to use an even number of tubes, for example, two in the main draw blood - the first tubes and two additional second tubes.

After centrifugation the plasma of the first tubes are transferred respectively to the third test tube. The amount of plasma in tubes equalize the transfusion of each other. Third tubes after centrifugation of the platelet-poor plasma is drained into the fourth tube, and the remaining third tubes platelet-rich plasma is poured into a third test tube. Then prepare the activator of platelet-rich plasma. The supernatant from the second tubes merge in the fifth test tube. To the content of the fifth test tube add in the plasma poor in platelets, the mass ratio at 1:1, and mix thoroughly. Then drip, with stirring, to impose a 10% solution of calcium chloride to obtain the desired consistency of the mixture, mix thoroughly. Then platelet-rich plasma to activate, which is mixed with the activator in the ratio of 1:3, respectively.

When using the inventive method of 9 ml of blood withdrawn from the patient received 1.5 ml of platelet-rich plasma.

1. The method of obtaining autologous activated platelet-rich plasma for dentistry, including drawing blood from the cubital vein by syringe-tube, centrifugation selected blood in the test tube, transfer the plasma (supernatant fraction was separated after centrifugation) into an empty test tube, centrifuging the newly selected product, removal of the tubes formed nadeshiko (plasma, platelet-poor), activating the remaining in vitro platelet-rich plasma, characterized in that it further carry out the second blood sampling syringe-tube in number, constituting half of the total selected blood first syringe is a tube that is left in the upright position for blood clotting with the first syringe tube with selected blood is placed in a centrifuge and centrifuged at acceleration of 250 g for 10 min, after which the formed plasma is transferred into an empty test tube and centrifuged with an acceleration of 1000 g for 10 min, then remove the tube from the centrifuge, and then formed after centrifugation the plasma poor in platelets, separated from platelet-rich plasma and transferred to an empty tube, then prepare the activator of platelet-rich plasma, which make sure that the second syringe-tube blood clotted, the donkey which the tube coagulating blood is placed in a centrifuge and centrifuged with an acceleration of 1000 g for 10 min, then the test tube is then removed from the centrifuge and transfer the formed after centrifugation gross blood supernatant into a second empty tube, which then adds the plasma poor in platelets, the mass ratio at 1:1, and mix thoroughly, then drip under stirring impose a 10%solution of calcium chloride to obtain the desired consistency of the mixture, mix thoroughly, after which platelet-rich plasma activated, for this purpose it is mixed with the received activator of the mass ratio to 1:3, respectively.

2. The method according to claim 1, characterized in that the tubes with plasma and with coagulating blood centrifuged at the same time, having first ascertained that in the second syringe-tube blood clotted.



 

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3 ex

FIELD: sorbents in biology.

SUBSTANCE: invention relates to biological and medical areas and can be used for treatment of biological fluids. Sorbent contains nucleus of ferromagnetic with mono- or bilayer envelop or without thereof. Nucleus is made in the form of plate 500 to 5000 μm in size across its plane and with thickness from 0.1 to 1000 μm. Method for manufacture of sorbent is characterized by that powder of iron and/or nickel, and/or titanium, and/or tantalum is vaporized in low-temperature plasma to form product, which is cooled and condensed in gas flow, condensed product is transferred into stabilized dispersion medium. Resulting crystals and microingots are treated to form plates with desired thickness, which are rinsed with distilled water. Weak parts of plates are detached, affected by ultrasound, and dried. Dried plates are fractioned and sorbent nuclei with desired size are isolated and then are layer-by-layer enveloped.

EFFECT: achieved creation of magnet-governed sorbent characterized by essentially larger area of particles for the same mass of nucleus and providing efficient purification of biological fluids.

16 cl, 10 ex

FIELD: medicine, clinical oncology.

SUBSTANCE: one should perform blood exfusion at the quantity of 500-550 ml. Removed blood volume should be compensated with crystalloids at the ratio of 1:1.2 - 1: 1.3. Erythrocytic mass should be supplemented with 5-10 ml "Essentiale" solution. This mass should be introduced for a patient at simultaneous ultraviolet blood irradiation for 20-30 min. Plasmapheresis in combination with ultraviolet blood irradiation should be carried out every other day, about 2-4 seances/course, totally. The method provides normalized level of blood leukocytes and body detoxication, it, also, excludes prophylactic course of antibioticotherapy that enables to continue terms of therapy and not to decrease the dosages of chemoradiation therapy.

EFFECT: higher efficiency of therapy.

1 ex, 2 tbl

FIELD: medicine, oncology.

SUBSTANCE: the method deals with applying chemopreparations incubated with autolymph and radiation therapy. Lymph taken out of patient's thoracic duct should be centrifuged for 30 min at 2200 rot./min, lymphatic plasma should be taken and frozen at -40 C, lymphatic formic elements should be incubated in a thermostat at 37 C for 1 h together with chemopreparations by a certain scheme to be reinfused for a patient intravenously by drops according to the given scheme. Then, 2 wk later, one should perform therapy with a split course of distance gammatherapy at single focal dosage (SFD) of 4 Gy. At the first stage one should apply per 2 Gy twice daily at 4-5-h-long interval 5 times weekly, at achieving focal dosage of 28 Gy it is necessary to have a week-long interval. Then radiation therapy should be continued but SFD of 4 Gy should be applied at once. One should fulfill 3 fractions of irradiation per a week, there are 6 fractions during the second stage, totally. Total focal dosage (TFD) per the whole course of irradiation corresponds to 52 Gy. About 4 wk after radiation therapy one should defrost lymphatic plasma to incubate it with the same chemopreparations in a thermostat at 37 C for 1 h to be then reinfused for a patient intravenously by drops. The method enables to decrease tumor volume and tumor process metastasing without any operative interference.

EFFECT: higher efficiency of therapy.

1 ex

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