Blood collection tube, method for preparing cell-free plasma, and use thereof

Abstract

A blood collection tube, a method for preparing cell-free plasma, and use thereof. The blood collection tube (100) comprises: an outer tube (110), comprising an outer tube wall (230) extending in a first direction (A); and an inner tube (120), fixed in the outer tube (110) and comprising an inner tube wall (330) extending in the first direction (A). The inner tube wall (330) comprises a filter portion (340) and a receiving portion (331). The receiving portion (331) is located between the filter portion (340) and an inner tube bottom (310) of the inner tube (120). The filter portion (340) is spaced apart from the outer tube wall (230), and the filter portion (340) comprises a plurality of filter holes (341) for filtering cells.

Description

Blood collection tube, method for preparing cell-free plasma and use

[0001] Cross-reference to Related Applications

[0002] This application claims the benefit of Chinese Patent Application No. 202411171036.6, filed on August 23, 2024. The entire teachings of the above application are incorporated herein by reference. TECHNICAL FIELD

[0003] The present application relates to the technical field of clinical medical devices. In particular, the present application relates to a method for collecting and preparing cell-free plasma, and the use of the blood collection tube in preparing cell-free plasma. BACKGROUND

[0004] Circulating cell-free DNA (cfDNA) is DNA molecules of 50-300 base pairs in length released from cells into the blood, which usually exists in low concentration in the blood of healthy people, but has important significance in disease detection (such as tumor detection and monitoring). However, cells and organelles contain a large amount of DNA, the content of which is much higher than that of cfDNA. In order to avoid the contamination of cfDNA by the DNA in cells and organelles, it is necessary to prepare cell-free plasma first. SUMMARY

[0005] In a first aspect of the present disclosure, a blood collection tube is provided. The blood collection tube comprises: an outer tube comprising an outer tube wall extending along a first direction; an inner tube fixed within the outer tube and comprising an inner tube wall extending along the first direction, wherein the inner tube wall comprises a filter portion and a containing portion, wherein the containing portion is located between the filter portion and an inner tube bottom of the inner tube, the filter portion is spaced apart from the outer tube wall, and the filter portion comprises a plurality of filter holes for filtering cells

[0006] In a second aspect of the present disclosure, a method for preparing cell-free plasma is provided, comprising: placing whole blood of a subject in the inner tube of the blood collection tube according to any one of the preceding claims; performing a centrifugation operation on the blood collection tube to cause the plasma in the whole blood to pass through the filter portion of the inner tube and enter the outer tube of the blood collection tube; removing the inner tube from the blood collection tube; and obtaining the plasma in the outer tube.

[0007] In a third aspect of the present disclosure, a use is provided, which is the use of the blood collection tube according to the first aspect of the present disclosure in preparing cell-free plasma.

[0008] It should be understood that the content described in the summary section is not intended to limit or define key or important features of the embodiments of the present disclosure, nor is it used to limit the scope of the present disclosure. Other features of the present disclosure will become apparent from the following description. Attached Figure Description

[0009] To better understand the above and other objects, features, advantages, and functions of this disclosure, reference can be made to the preferred embodiments shown in the accompanying drawings. Like reference numerals in the drawings refer to like parts. Those skilled in the art should understand that the drawings are intended to schematically illustrate preferred embodiments of this disclosure and are not intended to limit the scope of this disclosure; the parts in the drawings are not drawn to scale.

[0010] Figure 1 shows a schematic diagram of an example blood collection tube according to an embodiment of the present disclosure;

[0011] Figure 2 shows a schematic diagram of the outer tube of an example blood collection tube according to an embodiment of the present disclosure;

[0012] Figure 3 shows a schematic diagram of the inner tube of an example blood collection tube according to an embodiment of the present disclosure;

[0013] Figure 4 shows a schematic diagram of the cap of an example blood collection tube according to an embodiment of the present disclosure;

[0014] Figure 5 shows a schematic diagram of the filter membrane of an example blood collection tube according to an embodiment of the present disclosure;

[0015] Figure 6 shows a flowchart of an example method for preparing decellularized plasma according to an embodiment of the present disclosure;

[0016] Figures 7A-7C illustrate schematic diagrams of various example states during the preparation of decellularized plasma according to embodiments of the present disclosure; and

[0017] Figure 8 shows a schematic diagram of the distribution of cfDNA fragments extracted from decellularized plasma obtained using blood collection tubes and different preparation methods. Detailed Implementation

[0018] Various embodiments will now be described with reference to the accompanying drawings, wherein similar reference numerals are used throughout to denote similar elements. In the following description, numerous specific details are set forth for purposes of explanation in order to facilitate a thorough understanding of one or more embodiments. However, it may be apparent in some or all cases that any of the embodiments described below can be practiced without employing the specific design details described below. In other instances, well-known structures and devices are illustrated in block diagram form to facilitate the description of one or more embodiments. A simplified overview of one or more embodiments is given below to provide a basic understanding of the embodiments. This overview is not an exhaustive summary of all contemplated embodiments, is not intended to identify key or essential elements of all embodiments, nor is it intended to define the scope of any or all embodiments.

[0019] Reference within the framework of this description to “an embodiment” or “one embodiment” is intended to indicate that a particular configuration, arrangement or characteristic described with respect to an embodiment is included in at least one embodiment. Thus, appearances of such phrases in various places within this description are not necessarily intended to refer to the same embodiment. Furthermore, in one or more embodiments, particular configurations, arrangements or characteristics can be combined in any suitable way.

[0020] In the following disclosure, unless otherwise indicated, when referring to absolute position modifiers such as the terms “front”, “back”, “top”, “bottom”, “left”, “right”, etc. or relative position modifiers such as the terms “above”, “below”, “higher”, “lower”, etc. or when referring to directional modifiers such as “horizontal”, “vertical”, etc., reference is made to the orientation shown in the figures.

[0021] As discussed above, there is a need in the related art for preparing acellular plasma. In the related art, acellular plasma preparation methods are generally two-step centrifugation, i.e. first removing cells by low-speed centrifugation (about 3000g), and then taking the supernatant to high-speed centrifugation (12000-16000g) to remove cell debris.

[0022] However, high-speed centrifugation generally requires operation by professional personnel using specific equipment, and most conventional hospitals or medical centers may not be equipped with a centrifuge that can perform high-speed centrifugation of 12000-16000g. In this case, it is necessary to transport whole blood to a specialized institution for processing, but the transportation process is easy to cause the rupture of white blood cells in the blood and release a large amount of gDNA, causing contamination of cfDNA. In addition, devices for transporting whole blood at room temperature, such as sampling tubes with preservatives, contain a variety of other components in addition to common anticoagulants that can cause cfDNA contamination, such as preservatives, and are relatively expensive.

[0023] In addition, in the related art, acellular plasma preparation can also be performed using an ultrafiltration tube. However, such methods also have some problems. For example, cell membranes are relatively fragile and can be sheared and broken during centrifugation with the pores of the filter membrane, resulting in gDNA contamination and clogging of the filter membrane. Secondly, the ultrafiltration tube is generally a single-layer filter membrane and cannot effectively separate cells and organelles of different sizes.

[0024] Therefore, the present disclosure provides a simple and low-cost method for collecting and preparing acellular plasma. In the method, a blood collection tube is provided, which includes an inner tube having a cell filtration function. The blood collection tube includes an outer tube and an inner tube located inside the outer tube. A filter for filtering cells is arranged on the tube wall of the inner tube, so that the inside of the inner tube is in communication with the inside of the outer tube. Thus, when centrifugation is performed using the blood collection tube, the filter allows the plasma in the inner tube to pass and thus enter the outer tube, while blocking cells and cell fragments and organelles after cell rupture.

[0025] Compared with the related art method for collecting and preparing acellular plasma, the method for collecting and preparing acellular plasma according to the present disclosure simplifies the two-step centrifugation method in the conventional method for collecting and preparing acellular plasma, and acellular plasma can be obtained by one-step centrifugation. In addition, when the blood collection tube according to the present disclosure is used to collect and prepare acellular plasma, acellular plasma can be obtained by low-speed centrifugation, which reduces the high requirement for equipment (such as a high-speed centrifuge) in the conventional method for preparing acellular plasma. Thus, the time cost, labor cost, and material cost are reduced. Since the steps are simplified, the method according to the present disclosure is easy to operate and reduces the possibility of contamination in the preparation process of acellular plasma. At the same time, the method according to the present disclosure expands the application scenarios of the device for preparing acellular plasma. The blood collection tube according to the present disclosure can be used in physical examination centers and conventional hospitals to prepare acellular plasma, and is conducive to integration and automation.

[0026] The blood collection tube and the method for preparing acellular plasma according to the present disclosure will be described in detail below with reference to FIGS. 1 to 8. FIG. 1 shows a schematic diagram of an example blood collection tube 100 according to an embodiment of the present disclosure. As shown in FIG. 1, the blood collection tube 100 includes an outer tube 110 for containing plasma from which cells are removed by centrifugation. In the outer tube 110, an inner tube 120 for containing whole blood before centrifugation and containing cells after centrifugation is fixed. The blood collection tube 100 further includes a tube cover 130 for sealing the inner tube 120 in the outer tube 110. The structures of the outer tube 110, the inner tube 120, and the tube cover 130 will be described below with reference to FIGS. 2 to 5, respectively.

[0027] FIG. 2 shows a schematic diagram of an example outer tube 110 of a blood collection tube according to an embodiment of the present disclosure. As shown in FIG. 2, the outer tube 110 includes a circular outer tube bottom 210 at one end. The other end of the outer tube 110 includes an outer tube opening 220. The outer tube 110 further includes an outer tube wall 230 extending from the outer tube bottom 210 to the outer tube opening 220 in a first direction A pointing from the outer tube bottom 210 to the outer tube opening 220. The outer tube wall 230 has an annular cross-section.

[0028] In the illustrated embodiment, the outer tube mouth 220 is used to cooperate with the cap 130 to achieve sealing on one hand, and to secure the inner tube 120 on the other hand. To this end, the outer tube mouth 220 comprises an outer tube flange 221 extending outwardly, i.e. in a radial direction away from the outer tube wall 230. The outer tube flange 221 forms a circular ring around the outer tube wall 230. The outer tube flange 221 comprises a second inner surface facing the outer tube bottom 210, and a second outer surface 222 facing away from the outer tube bottom 210. The second outer surface 222 is used to support the inner tube 120. In addition, the outer tube mouth 220 further comprises an outer tube mouth peripheral wall 223 extending from an outer edge of the outer tube flange 221 in the first direction. An inner surface of the outer tube mouth peripheral wall 223 is capable of tightly abutting against the cap 130 to achieve sealing of the interior of the outer tube 110. It should be understood that the manner of securing the inner tube 120 as defined in the embodiments of the present disclosure is merely exemplary. For example, a bracket can also be provided inside the outer tube, so as to secure the inner tube on the bracket. The present disclosure is not intended to be limited in this regard.

[0029] Fig. 3 shows a schematic view of the inner tube 120 of an example blood collection tube according to an embodiment of the present disclosure. As shown in Fig. 3, the inner tube 120 comprises a circular inner tube bottom 310 at one end. The other end of the inner tube 120 comprises an inner tube mouth 320. The inner tube 120 further comprises an inner tube wall 330 extending from the inner tube bottom 310 to the inner tube mouth 320 in the first direction A. The inner tube wall 330 has an annular cross-section. In the illustrated embodiment, the inner tube mouth 320 of the inner tube 120 comprises an inner tube flange 321 extending outwardly from the inner tube wall 330. The inner tube flange 321 comprises a first inner surface 322 facing the inner tube bottom 310, and a first outer surface facing away from the inner tube bottom 310. The outer diameter of the inner tube flange 321 is greater than the inner diameter of the outer tube 110, so that the inner tube 120 can be secured on the outer tube 110 by having the first outer surface rest on the second outer surface 222 of the outer tube flange 221. In addition, the outer diameter of the inner tube 120 is smaller than the inner diameter of the outer tube 110, and the length of the inner tube 120 is smaller than the length of the outer tube 110, so that after the inner tube 120 is secured on the outer tube 110, the inner tube bottom 310 is spaced apart from the outer tube bottom 210, and the inner tube wall 330 is spaced apart from the outer tube wall 230. It should be understood that the positional relationship in which the inner tube bottom 310 is spaced apart from the outer tube bottom 210 in this embodiment is merely exemplary, and the inner tube bottom 310 can be in partial contact with the outer tube bottom 210.

[0030] In the illustrated embodiment, the inner tube wall 330 further comprises a filter portion 340 adjacent to the inner tube mouth 320 in the first direction A and a containing portion 331 abutting the inner tube bottom 310, such that the containing portion 331 is located between the filter portion 340 and the inner tube bottom 310. The filter portion 340 comprises a plurality of filter pores 341 for filtering cells (such as cell debris and organelles after being broken up), and the containing portion 331 does not comprise pores, so as to contain the cells (i.e. cell precipitate and cell debris and organelles after being broken up) in the inner tube 120 blocked by the filter portion 340. Since the inner tube bottom 310 is spaced apart from the outer tube bottom 210 and the inner tube wall 330 is spaced apart from the outer tube wall 230 after the inner tube 120 is fixed on the outer tube 110, the filter portion 340 on the inner tube wall 330 allows the interior of the inner tube 120 to communicate with the interior of the outer tube 110, so that the liquid in the inner tube 120 can enter the outer tube 110 from the inner tube 120 through the filter portion 340. In embodiments for filtering cells, the average pore size of the plurality of filter pores 341 is in the range of 0.1 pm-1.5 pm. In some embodiments, the pore size of the filter pores 341 can be in the range of 0.2 pm-0.8 pm. In some embodiments, the ratio between the distance from the bottom of the filter portion 340 to the inner tube bottom 310 and the length of the inner tube wall 230 can be at least 45%. In some embodiments, the above-mentioned ratio can also be 45%-70%, 45%-65%, 45%-60%, 45%-55%, 45%-50% or 45%.

[0031] In some embodiments, the filter portion 340 comprises at least one opening opened on the inner tube wall 330, and a filter membrane 343 laid on the inner tube wall 330 and covering the at least one opening. The filter membrane 343 comprises a plurality of filter pores 341. In the embodiment of FIG. 3, the inner tube wall 330 comprises at least three openings, i.e. opening 342-1, opening 342-2 and opening 342-3. The filter membrane 343 is laid on the inner wall of the inner tube wall 330. The filter membrane 343 covers the opening 342-1, the opening 342-2 and the opening 342-3 in the inner tube wall 330.

[0032] In some embodiments, the filter membrane 343 can be a multi-layer porous membrane. For example, the filter membrane 343 can be a double-layer porous membrane. The filter membrane 343 implemented as a double-layer porous membrane will be described below with reference to FIG. 5. FIG. 5 shows a schematic diagram of a filter membrane 343 of an example blood collection tube according to an embodiment of the present disclosure. For the purpose of clarity, FIG. 5 only shows the filter membrane without showing other components. As shown in FIG. 5, the double-layer filter membrane includes a first filter membrane 510 and a second filter membrane 520. The first filter membrane 510 and the second filter membrane 520 are overlaid together in an unfolded state to form the double-layer filter membrane 343. When laid on the inner tube wall 330 of the inner tube 120, the first filter membrane 510 is located inside the second filter membrane 520, i.e., further away from the inner tube wall 330 than the second filter membrane 520.

[0033] Here, the first filter membrane 510 includes a plurality of first filter holes 511 of the plurality of filter holes 341. The second filter membrane 520 includes a plurality of second filter holes 521 of the plurality of filter holes 341. The first filter hole diameter of the first filter holes 511 is greater than the second filter hole diameter of the plurality of second filter holes 521. In some embodiments, the first filter hole diameter can be in the range of 0.5 μm-1.5 μm, and the second filter hole diameter can be in the range of 0.1 μm-0.5 μm. In other embodiments, the first filter hole diameter can be in the range of 0.5 μm-0.8 μm, and the second filter hole diameter can be in the range of 0.2 μm-0.5 μm. In such embodiments, by setting a multi-layer filter membrane with different hole diameters, the filtering effect can be improved, and excessive impurities can be prevented from entering the outer tube.

[0034] FIG. 4 shows a schematic diagram of a tube cap 130 of an example blood collection tube according to an embodiment of the present disclosure. As shown in FIG. 4, the tube cap 130 includes a circular cap body 410. An annular wall 420 extending from a first end face 411 in a direction away from the first end face 411 is formed on the outer periphery of the cap body 410. In addition, a column 430 extending from a second end face 412 in a direction away from the second end face 412 is formed on the second end face 412 opposite the first end face 411 of the cap body 410. The inner wall of the annular wall 420 is spaced apart from the outer wall of the column 430 to form a space between the annular wall 420 and the column 430 for accommodating the outer tube mouth peripheral wall 223 of the outer tube 110. The cap body 410 further includes a first recess formed in the middle portion and extending downward from the first end face 411. Meanwhile, the column 430 is hollow, so that the thickness of the center of the tube cap 130 is small, which is conducive to the insertion of the needle tube. In some embodiments, the tube cap 130 can be made of an elastic material.

[0035] The structure of the different components of the blood collection tube 100 has been described in detail. Now returning to FIG. 1, in some embodiments, when assembling the blood collection tube 100, the outer tube 120 in the embodiment shown in FIG. 2 can be provided first, and then the inner tube 120 is fixed on the outer tube 110 by placing the inner tube flange 321 of the inner tube 120 on the outer tube flange 221 of the outer tube mouth 220. After the inner tube 120 is arranged, reagents 140 suitable for performing the preparation of acellular plasma, such as anticoagulants, are added in the inner tube 120. Finally, a negative pressure is formed in the tube and the tube cap 130 is capped on the outer tube mouth 320. By abutting the end of the cylinder 430 of the tube cap 130 against the inner tube flange 321, the inner tube flange 321 is pressed against the outer tube flange 221, and the cylinder 430 and the ring wall 420 are clamped on the outer tube mouth peripheral wall 223, so as to seal the space in the tube. In this way, the blood collection tube 100 in a marketable state is obtained.

[0036] In some embodiments, after the user obtains the blood collection tube 100, the user can insert one end of the blood collection needle into the blood collection tube 100 and the other end into the blood vessel of the subject. At this time, due to the negative pressure in the blood collection tube 100, the whole blood of the subject flows into the inner tube 120 of the blood collection tube 100 under the action of the pressure difference. Then, the user can use the blood collection tube 100 containing the whole blood of the subject to prepare acellular plasma. The preparation process of acellular plasma will be described below with reference to FIGS. 6-7C.

[0037] FIG. 6 shows a flowchart of an example method 600 of preparing acellular plasma according to embodiments of the present disclosure. As shown in FIG. 6, at 602, the whole blood of the subject is placed in the inner tube 120 of the blood collection tube 100 in the embodiment shown in FIG. 1. In some embodiments, the whole blood can be collected from the subject using the blood collection tube under a negative pressure. For example, after a channel is formed between the blood vessel of the subject and the blood collection tube, the whole blood can flow from the blood vessel into the inner tube 120 of the blood collection tube 100 via the tube cap 130 under the action of the pressure difference. At this time, the state in the blood collection tube 100 is as shown in FIG. 7A. As shown in FIG. 7A, the inner tube 120 of the blood collection tube 100 is fixed in the outer tube 110 by the tube cap 130. At this time, the whole blood 710 is contained in the inner tube 120.

[0038] At 604, centrifugation is performed on the blood collection tube 100 to cause the plasma in the whole blood to pass through the filter portion 340 of the inner tube 120 into the outer tube 110 of the blood collection tube 100. In some embodiments, after the whole blood 710 and the anticoagulant 140 in the inner tube 120 are uniformly mixed, the obtained blood collection tube 100 containing the whole blood 710 is placed in a centrifuge, and centrifugation is performed under suitable centrifugation conditions. The cells in the whole blood 710 are precipitated to the bottom 310 of the inner tube under the action of centrifugal force, forming a cell precipitate 711, and the upper layer of plasma is filtered through the filter membrane 343 covering the opening on the inner tube wall 330 of the inner tube 120 close to the inner tube opening 320 under the action of centrifugal force to remove cell debris and organelles. The cell debris and organelles are left in the inner tube 120, and the plasma flows into the outer tube 110, thereby obtaining cell-free plasma 712. As shown in FIG. 7B, the inner tube 120 of the blood collection tube 100 contains the cell precipitate 711, and the outer tube 110 contains the cell-free plasma 712.

[0039] In some embodiments, the centrifugation is a low-speed centrifugation. The conditions of the low-speed centrifugation can be: a centrifugal force of 1500-4500g, a centrifugation time of 5-20 minutes, and a centrifugation temperature of 2-8°C. In some embodiments, the conditions of the low-speed centrifugation can be: a centrifugal force of 2000-3000g, a centrifugation time of 10-15 minutes, and a centrifugation temperature of 2-8°C.

[0040] At 606, the inner tube 120 is removed from the blood collection tube 100, and at 608, the plasma in the outer tube 110 is obtained. In some embodiments, the tube cap 130 can be opened, the inner tube 120 is taken out and discarded, and the outer tube 110 is again sealed with the tube cap 130, thereby obtaining the cell-free plasma 711 placed inside the outer tube 110. As shown in FIG. 7C, the inner tube 120 of the blood collection tube 100 is taken out, and the tube cap 130 is fixed on the outer tube 110 to seal the cell-free plasma 711 in the outer tube 110.

[0041] FIG. 8 shows a schematic diagram of the distribution of cfDNA fragments extracted from cell-free plasma obtained using blood collection tubes and different preparation methods. Here, whole blood was collected from the same source using a Streck blood collection tube, a blood collection tube according to an embodiment of the present disclosure, and an EDTA blood collection tube commonly used in the art, in which the inner wall is coated with ethylenediaminetetraacetic acid (EDTA) anticoagulant, and divided into 4 groups, in which 1 tube of whole blood was collected using the blood collection tube according to an embodiment of the present disclosure (experimental group 1), 1 tube of whole blood was collected using the Streck blood collection tube (experimental group 3), and 2 tubes of whole blood were collected using the EDTA blood collection tube (control group and experimental group 2). The plasma of the 4 groups of whole blood was separated according to the following method:

[0042] Control group: whole blood collected by EDTA blood collection tube, prepared into acellular plasma by two-step centrifugation according to the method commonly used in the art, the specific steps are as follows:

[0043] 1. Collect whole blood by EDTA blood collection tube, centrifuge at 3000g for 10 minutes at 4°C;

[0044] 2. Move the upper plasma to a new centrifuge tube, and pay attention not to suck the white membrane layer containing leukocytes and the lower red blood cells;

[0045] 3. Centrifuge the centrifuge tube containing the plasma at 16000g for 10 minutes at 4°C;

[0046] 4. Save the upper plasma into a new test tube to obtain acellular plasma.

[0047] Experimental group 1: whole blood collected by blood collection tube according to the present disclosure, prepared into acellular plasma by one-step centrifugation according to the method described in the present disclosure, the specific steps are as follows:

[0048] 1. Collect whole blood by blood collection tube according to the present disclosure, mix well;

[0049] 2. Centrifuge at 3000g for 10 minutes at 4°C;

[0050] 3. Take out and discard the tube holder (200), close the tube body (400) with the tube cap (100) to obtain the acellular plasma placed inside the tube body (400).

[0051] The filter membrane used in the blood collection tube in this embodiment is a double-layer filter membrane made of polyether sulfone, the average pore size of the inner filter membrane is about 0.8 μm, and the average pore size of the outer filter membrane is about 0.4 μm.

[0052] Experimental group 2: whole blood collected by EDTA blood collection tube, prepared into acellular plasma by one-step centrifugation in experimental group 1, the specific steps are as follows:

[0053] 1. Collect whole blood by EDTA blood collection tube, centrifuge at 3000g for 10 minutes at 4°C;

[0054] 2. Move the upper plasma to a new centrifuge tube, and pay attention not to suck the white membrane layer containing leukocytes and the lower red blood cells to obtain acellular plasma.

[0055] Experimental group 3: whole blood preserved by Streck blood collection tube, prepared into acellular plasma according to the method of experimental group 2, the specific steps are as follows:

[0056] 1. Collect whole blood by Streck blood collection tube, centrifuge at 3000g for 10 minutes at 4°C;

[0057] 2. The upper plasma is moved to a new centrifuge tube, and the white membrane layer containing leukocytes and the lower red blood cells are not sucked to obtain the cell-free plasma.

[0058] The cell-free plasma collected from the control group and experimental groups 1, 2 and 3 is subjected to cfDNA extraction using a QIAamp Circulating Nucleic Acid Kit (50) kit, and the extracted cfDNA is subjected to fragment size analysis using a labchip.

[0059] The main peak of cfDNA is theoretically located at about 166 bp, and the two peaks are located at about 332 bp. There should be no obvious peak in the longer fragment region. If a peak appears in the longer fragment region, such as the region above 500 bp, it represents that the cfDNA is contaminated by large fragment DNA, such as gDNA, in cells or organelles.

[0060] After extraction, the results are shown in FIG. 8: the cell-free plasma separated from the experimental group 1 (blood collection tube according to the present disclosure) after one-step centrifugation is subjected to cfDNA extraction, and the fragment distribution of the cfDNA is consistent with that of the control group, specifically: the main peak is consistent, and is located at about 166 bp; there are two peaks, and no large fragment DNA above 500 bp is contaminated. The experimental groups 2 and 3 only undergo one-step centrifugation, and are contaminated by large fragment DNA above 500 bp.

[0061] Therefore, the blood collection tube for collecting and preparing cell-free plasma according to the present disclosure and the use method thereof do not need to undergo a complicated two-step centrifugation, but can complete the pretreatment of cfDNA plasma, the centrifugal separation of plasma and blood cells, and the separation of organelles and cell fragments in plasma by one-step centrifugation. According to the scheme of the present disclosure, high-speed centrifugation at 12000-16000g is not needed, but low-speed centrifugation can complete the pretreatment of cfDNA plasma.

[0062] The above has described various embodiments of the present disclosure, and the above description is exemplary, not exhaustive, and is not limited to the disclosed embodiments. Many modifications and changes are obvious to those skilled in the art without departing from the scope and spirit of the described embodiments. The selection of terms used herein is intended to best explain the principles, practical applications, or improvements to the technology in the market of the embodiments, or to enable other ordinary skilled persons in the art to understand the various implementations disclosed herein.

Claims

1. A blood collection tube (100), comprising: an outer tube (110) comprising an outer tube wall (230) extending along a first direction (A); an inner tube (120) fixed within the outer tube (210) and comprising an inner tube wall (330) extending along the first direction (A), wherein the inner tube wall (330) comprises a filter portion (340) and a receiving portion (331), wherein the receiving portion (331) is located between the filter portion (340) and an inner tube bottom (310) of the inner tube (120), the filter portion (340) is spaced apart from the outer tube wall (230), and the filter portion (340) comprises a plurality of filter holes (341) for filtering cells.

2. The blood collection tube (100) according to claim 1, wherein a hole diameter of the filter holes (341) is in a range of 0.1 pm - 1.5 pm.

3. The blood collection tube (100) according to claim 2, wherein the hole diameter of the filter holes (341) is in a range of 0.2 pm - 0.8 pm.

4. The blood collection tube (100) according to claim 1, wherein the filter portion (340) comprises: at least one opening (342-1, 342-2, 342-3) opened on the inner tube wall (330); and a filter membrane (343) laid on the inner tube wall (330) and covering the at least one opening (342-1, 342-2, 342-3).

5. The blood collection tube (100) according to claim 4, wherein the filter membrane (343) comprises: a first filter membrane (510) comprising a plurality of first filter holes (511) of the plurality of filter holes (341), the first filter holes (511) having a first filter hole diameter; and a second filter membrane (520) comprising a plurality of second filter holes (521) of the plurality of filter holes (341), the plurality of second filter holes (521) having a second filter hole diameter, wherein the first filter membrane (510) is located inside the second filter membrane (520), and the first filter hole diameter is larger than the second filter hole diameter.

6. The blood collection tube (100) according to claim 5, wherein the first filter hole diameter is in a range of 0.5 pm - 1.5 pm, and the second filter hole diameter is in a range of 0.1 pm - 0.5 pm.

7. The blood collection tube (100) according to claim 6, wherein the first filter hole diameter is in a range of 0.5 pm - 0.8 pm, and wherein the second filter hole diameter is in a range of 0.2 pm - 0.5 pm.

8. The blood collection tube (100) according to claim 1, wherein the outer tube wall (230) extends from an outer tube bottom (210) to an outer tube mouth (220), the outer tube mouth (220) comprises an outer tube flange (221) extending outwardly from the outer tube wall (230), and an inner tube mouth (320) of the inner tube (120) comprises an inner tube flange (321) extending outwardly from the inner tube wall (330), ​ ​ wherein a first inner surface (322) of the inner tube flange (321) facing the inner tube bottom (310) abuts against a second outer surface (222) of the outer tube flange (221) facing away from the outer tube bottom (210).

9. The blood collection tube (100) according to claim 8, wherein the outer tube mouth (220) further comprises an outer tube mouth peripheral wall (223) extending from an outer edge of the outer tube flange (221) in the first direction, and the blood collection tube (100) further comprises a tube cap (130) disposed in the outer tube mouth (220), wherein an outer wall of the tube cap (130) abuts against the outer tube mouth peripheral wall (223), and an end of the tube cap (130) abuts against a first outer surface of the inner tube flange (321) facing away from the inner tube bottom (310).

10. The blood collection tube according to claim 9, wherein the tube cap (130) is made of an elastic material.

11. A method (600) of preparing a decellularized plasma, comprising: placing whole blood of a subject in an inner tube (120) of a blood collection tube (100) according to any one of the preceding claims; performing a centrifugation operation on the blood collection tube (100) to cause plasma in the whole blood to pass through a filter portion (340) of the inner tube (120) into an outer tube (110) of the blood collection tube (100); removing the inner tube (120) from the blood collection tube (100); and obtaining the plasma in the outer tube (110).

12. The method according to claim 11, wherein the centrifugation operation is a low speed centrifugation operation, wherein the low speed centrifugation operation is under conditions of a centrifugal force of 1500-4500 g, a centrifugation time of 5-20 minutes, and a centrifugation temperature of 2-8 °C.

13. The method according to claim 12, wherein the low speed centrifugation operation is under conditions of a centrifugal force of 2000-3000 g, a centrifugation time of 10-15 minutes, and a centrifugation temperature of 2-8 °C.

14. Use of the blood collection tube according to any one of claims 1-10 in preparing a decellularized plasma.

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