CN110394063B

CN110394063B - Centrifugal integrated molecular fractionation device

Info

Publication number: CN110394063B
Application number: CN201910752172.7A
Authority: CN
Inventors: 徐恩良; 徐高扬; 吴凤霞
Original assignee: Xiamen Xianming Biotechnology Co ltd
Current assignee: Xiamen Xianming Biotechnology Co ltd
Priority date: 2019-08-15
Filing date: 2019-08-15
Publication date: 2022-03-04
Anticipated expiration: 2039-08-15
Also published as: CN110394063A

Abstract

The invention relates to a centrifugal integrated molecular fractionation device, which comprises a separation tube and an integrated tube, wherein the separation tube comprises a tube body with a downward decreasing radius and a filter membrane arranged on the lower end part of the tube body, the tube body is divided into an upper receiving part and a lower filter part, at least two ribs which uniformly and axially extend are formed on the outer wall of the upper receiving part at intervals, a first matching part is formed on the upper end surface of each rib, a second matching part which is complementary with the first matching part is formed on the lower end surface of each rib, an upward opening notch corresponding to each rib is formed on the tube wall of the integrated tube, a third matching part which is complementary with the second matching part is formed at the bottom of each notch, a plurality of separation tubes with filter membranes with different apertures can be mutually inserted and fixed in the integrated tube in a nesting manner, and the aperture of the filter membrane of a superior separation tube is larger than that of a subordinate separation tube. The invention can realize one-time sample loading, obtain different target molecules by synchronous multi-stage separation, has simple and quick operation and saves samples.

Description

Centrifugal integrated molecular fractionation device

Technical Field

The invention relates to a molecular separation device, in particular to a grading separation device for synchronously and quickly separating and obtaining multi-target molecules in places such as life sciences, medicine, environment, agriculture, food safety and the like.

Background

In research and practical work in the fields of life science, medicine, environment, agriculture, food safety and the like, target biomolecules need to be rapidly separated and obtained from biological samples. The realization of rapid fractionation to obtain a plurality of target molecules has real requirements in both research and production.

At present, the technical means based on molecular separation and purification are rich, various, simple and various, such as salting-out method, centrifugal precipitation method, chromatography, filter membrane filtration method and the like, and the requirements of people for obtaining high-purity target molecules are met to a greater extent.

Among them, chromatography is a broad class of methods, and separation and analysis methods are numerous, such as adsorption chromatography, partition chromatography, ion exchange chromatography, affinity chromatography, size exclusion chromatography (molecular sieve), hydrophobic chromatography, and the like, and basically aim to separate a single molecular target. The chromatography greatly promotes the separation and analysis development of the biological molecules.

The method based on the membrane filtration comprises microfiltration, ultrafiltration, nanofiltration and reverse osmosis, and can be used for filtering or intercepting target particles or molecules with different molecular sizes for collection. At present, a filter membrane with a relatively determined pore size is adopted to separate and collect target molecules, and basically, the aim of separating and obtaining single-molecule target objects is achieved, so that the method is widely applied.

Microfiltration can retain particles larger than between 0.1 and 1 micron. Microfiltration membranes allow the passage of large molecules and dissolved solids (inorganic salts) but retain particulate matter such as suspended matter, bacteria and large molecular weight colloids.

Ultrafiltration is a process in which a microfiltration membrane having a nominal pore size of 0.01 microns or less is used and solute molecules having a pore size smaller than the pore size are separated by applying a suitable pressure to one side of the membrane to separate the molecules having a molecular weight greater than 500 daltons (atomic mass units). The ultrafiltration centrifugal tube is mostly used for concentrating and exchanging buffer solution of laboratory micro samples (less than 20 ml) and removing other interferents with small molecular weight.

Nanofiltration is a process of selectively separating different components of a solution by utilizing the charge property of a nanofiltration membrane body and the mechanical screening function, and is called membrane separation. The aperture range of the nanofiltration membrane is about a few nanometers, the separation process is a physical process, the phase change and the addition of an auxiliary agent for nanofiltration are not needed, and the nanofiltration membrane is mainly used for concentration of low-molecular organic matters such as polypeptide and the like. The nanofiltration membrane is a material with a charge selective separation function, so that the nanofiltration membrane still has high desalting performance under very low pressure and can remove inorganic salts by a membrane with the molecular weight cutoff of hundreds.

Reverse osmosis is a membrane separation operation that uses a pressure differential as a driving force to separate a solvent from a solution. According to different osmotic pressures of various materials, a reverse osmosis pressure which is larger than the osmotic pressure is used, namely a reverse osmosis method, so that the purposes of separation, extraction, purification and concentration are achieved. Because of the small solvent molecules, the pore size of the reverse osmosis membrane is minimal.

The separation is to separate and collect the target molecules by a filter membrane with a relatively determined pore size. For a small amount of precious biological samples, the separation of a plurality of target objects takes a long time and the loss is large. If the sample can be loaded once, multistage filter membranes are adopted to sequentially and synchronously separate, and target molecules intercepted or filtered by filter membranes with different pore diameters are respectively collected, the device is not available at present. The device is invented based on the requirement of simultaneously separating multiple targets in the same sample.

Disclosure of Invention

The invention aims to provide a centrifugal integrated molecular fractionation device, which adopts a multistage filter membrane to perform one-time sample loading sequential synchronous separation, and respectively collects target molecules or particles intercepted or filtered by the filter membranes with different pore diameters, thereby improving the separation efficiency, shortening the separation time and saving the samples. Therefore, the invention adopts the following specific technical scheme:

a centrifugally integrated molecular fractionation apparatus comprising a separation tube and an integrated tube, wherein the separation tube comprises a tube body having a downward decreasing radius and a filter membrane mounted on a lower end portion of the tube body, the tube body is divided into an upper receiving portion and a lower filtering portion, at least two ribs extending in an axial direction are formed on an outer wall of the upper receiving portion at regular intervals, upper end surfaces of the ribs are formed with first fitting portions, lower end surfaces of the ribs are formed with second fitting portions complementary to the first fitting portions, an upwardly open notch corresponding to the ribs is formed on a tube wall of the integrated tube, a bottom of the notch is formed with third fitting portions complementary to the second fitting portions, and a plurality of separation tubes having different-diameter filter membranes are inserted and fixed in the integrated tube so as to be nested with each other, wherein an aperture of the filter membrane of an upper stage separation tube is larger than an aperture of the filter membrane of a lower stage separation tube and the lower stage separation tube passes through the filter membrane of a higher stage separation tube The filter portion is received in said upper receiving portion of the lower stage separator tube.

Further, the filter membrane is disposed at an inclination angle such that the filter membrane is upright and perpendicular to the centrifugal force when the centrifugally integrated molecular fractionation device is placed on an inclined corner of a centrifuge.

Further, the lower end portion is a slope.

Further, the first and third mating portions have a recessed structure and the second mating portion has a raised structure complementary to the recessed structure.

Further, the concave structure is rectangular, trapezoidal, triangular or zigzag.

Further, the filter membrane is a microfiltration membrane, an ultrafiltration membrane, a nanofiltration membrane, a reverse osmosis filter membrane or the like.

Further, the separating pipe and the integrated pipe are both integrally formed by injection molding of PP plastics.

Further, the bottom of the integrated pipe is formed with a cone, and a sleeve is sleeved outside the cone.

Further, the number of the ribs and the notches is 2-4.

By adopting the technical scheme, the invention has the beneficial effects that: the centrifugal integrated molecular fractionation device can realize one-time sample loading and synchronous multi-stage separation to obtain different target molecules, is simple and quick to operate, and saves samples.

Drawings

To further illustrate the various embodiments, the invention provides the accompanying drawings. The accompanying drawings, which are incorporated in and constitute a part of this disclosure, illustrate embodiments of the invention and, together with the description, serve to explain the principles of the embodiments. Those skilled in the art will appreciate still other possible embodiments and advantages of the present invention with reference to these figures. Elements in the figures are not drawn to scale and like reference numerals are generally used to indicate like elements.

FIG. 1 is a schematic view of a separation tube of a centrifugally integrated molecular fractionation apparatus of the present invention;

FIG. 2 is a schematic view of an integrated tube of the centrifugally integrated molecular fractionation apparatus of the present invention;

FIGS. 3a-3c are schematic views of a first mating portion of a separator tube and a third mating portion of an integration tube;

FIGS. 4a-4c are schematic views of a second mating portion of a separator tube;

FIG. 5 is a schematic view of a plurality of separator tubes of FIG. 1 nested within one another.

Detailed Description

The invention will now be further described with reference to the accompanying drawings and detailed description.

A centrifugally integrated molecular fractionation apparatus may include a plurality of separation tubes 1 and an integration tube 2. The separation tubes 1 and the integration tubes 2 are dimensioned such that each integration tube 2 can integrate 3, 4, 5 or 6 separation tubes 1. The specific structures of the separation tube 1 and the integration tube 2 will be described below with reference to fig. 1 to 5, respectively.

As shown in fig. 1, the separation tube 1 may include a tube body 11 having a radius decreasing downward and a filter 12 installed at a lower end portion of the tube body 11. The tubular body 11 may be integrally injection molded from pp plastic and divided into an upper receiving portion 111 and a lower filtering portion 112. The upper receiving portion 111 is a lumen for introducing a sample and is an area for receiving the lower filtering portion 112 of the upper-stage separation tube 1. The outer wall of the upper receiving portion 111 is formed with at least two ribs 113 extending uniformly in the axial direction. The rib 113 has a first engagement portion formed on an upper end surface thereof and a second engagement portion complementary to the first engagement portion formed on a lower end surface thereof. Wherein, the first matching part is a concave structure, and the shape of the first matching part can be rectangle, triangle, sawtooth or trapezoid, etc., as shown in fig. 3a-3 c; the second mating portion is a protrusion mechanism, as shown in fig. 4a-4 c. Thus, the separation tubes may be nested within each other by rectangular, trapezoidal, triangular or saw-tooth fits, as shown in FIG. 5, so that the separation tubes and the separation tubes are firmly joined without displacement during high speed centrifugation. The lower end of the lower filter portion 112 is a bevelled surface on which the filter membrane 12 can be mounted by means of a corresponding filter membrane carrier (not shown). The filter membrane carrier may be secured to the lower end of the lower filter portion 112 by snap-fitting or ultrasonic welding. Thus, the filter membrane 12 is also inclined. The inclination angle of the filter membrane 12 is set so that the filter membrane 12 stands upright and is perpendicular to the centrifugal force when the centrifugally integrated molecular fractionation apparatus is placed on an inclined rotor of a centrifuge. This maximizes the filtration area of the filter membrane and accelerates the passage of the sample through the filter membrane, thereby improving filtration efficiency. The construction of centrifuges is well known and will not be described here. The filter membrane 12 may be a microfiltration membrane, an ultrafiltration membrane, a nanofiltration membrane, a reverse osmosis membrane, or the like. Of course, in the case where the lower end portion of the lower filter portion 112 is a flat surface, the filter membrane 12 may be installed obliquely.

As shown in fig. 1-4, the integrated tube 2 is a circular cavity structure with a closed lower end and is integrally injection-molded by pp plastic. The integrated pipe 2 is divided into an upper part, a middle part and a lower part, wherein, the pipe wall of the upper part of the integrated pipe 2 is provided with a notch 21 which is opened upwards and corresponds to the rib 113 of the separation pipe 1. In the embodiment shown, the number of ribs 113 and indentations 21 is 4, but it could also be 2, 3 or more than 4. The width of the gap 21 corresponds to the width of the rib 113 so that the rib can be tightly engaged in the gap 21. The bottom of the indentation 21 is formed with a third mating portion, complementary to the second mating portion of the separator tube 1, and shaped as shown in fig. 3. Therefore, the separation tube 1 and the integrated tube 2 can be fixedly connected with each other through rectangular fitting, trapezoidal fitting, triangular fitting or zigzag fitting, so that the separation tube and the integrated tube can be firmly combined without displacement during high-speed centrifugation. The middle portion of the integrated pipe is adapted to receive the lower filtering portion 112 of the lowermost separation pipe 1. The bottom (lower part) of the header 2 is formed with a cone 22, the cone 22 being used to collect filtrate from the lowermost separator tube 2. A sleeve 23 is sleeved outside the cone 22, and the sleeve 23 is used for protecting the cone 22.

The integrated tube 2 is also provided with an upper cover to seal the upper opening of the uppermost separation tube 1 to prevent the sample from leaking. The structure of the upper cover is the same as that of the upper cover of the traditional centrifuge tube, such as a screw-top cover or the upper cover structure disclosed in CN206642738U, and will not be described here.

The working principle of the present invention is explained in detail below.

When the separating tube is used, firstly, the separating tube is selected according to the size of molecules to be separated and intercepted from large to small according to the aperture of a filter membrane or according to the size of molecules to be intercepted from large to small (according to the estimated size of molecular weight), then the rib of the separating tube with the smallest aperture of the filter membrane is aligned and inserted into the notch of the integrated tube, the second matching part of the separating tube is matched and embedded with the third matching part of the integrated tube, and the separating tube is pressed to the bottommost part; and then the ribs of the grading separation tube with larger aperture of the filter membrane are aligned and inserted into the gap of the integrated tube, and the second matching part of the grading separation tube is matched and embedded with the first matching part of the next-stage separation tube and is firmly fixed, so that the required grading separation tubes are sequentially placed. And a sample with a specified volume is added into the last-stage separation tube, and finally, the screw top cover of the integrated tube is screwed on, so that the tightness is moderate.

The integrated tube containing the separation tube is placed at an oblique angle turning head of a centrifuge, and the integrated tube is placed in the outward direction of the separation tube filter membrane, so that the separation tube filter membrane is in a vertical state. Proper rotating speed and centrifugal time are set according to the membrane tolerance and the radius of the rotor, and the membrane is centrifuged at 4 ℃ for specified time. During the washing, buffer solution in certain volume may be added to wash the separating tubes several times and the volume of the cone of the integrating tube is limited.

After the washing separation is completed, the target in each layer of separation tube is washed out with a suitable buffer. Adding buffer solution containing glycerol into the separation tube for storage.

The invention can realize one-time sample loading, obtain different target molecules by synchronous multi-stage separation, has simple and quick operation and saves samples.

While the invention has been particularly shown and described with reference to a preferred embodiment, it will be understood by those skilled in the art that various changes in form and detail may be made therein without departing from the spirit and scope of the invention as defined by the appended claims.

Claims

1. A centrifugally integrated molecular fractionation device, comprising a separation tube and an integration tube, wherein the separation tube comprises a tube body with a downward decreasing radius and a filter membrane mounted at the lower end of the tube body, the lower end is an inclined surface, the filter membrane is arranged in an inclined manner at an inclination angle such that the filter membrane is upright and perpendicular to a centrifugal force when the centrifugally integrated molecular fractionation device is placed on an inclined corner of a centrifuge, the tube body is divided into an upper receiving portion and a lower filtering portion, at least two ribs extending in an axial direction are formed at regular intervals on the outer wall of the upper receiving portion, first engagement portions are formed on the upper end surfaces of the ribs, second engagement portions complementary to the first engagement portions are formed on the lower end surfaces of the ribs, and upwardly open notches corresponding to the ribs are formed on the tube wall of the integration tube, the bottom of the notch is formed with a third fitting portion complementary to the second fitting portion, a plurality of the separation tubes having filter membranes of different diameters are inserted and fixed in the integrated tube so as to be able to be nested with each other, wherein the diameter of the filter membrane of the upper separation tube is larger than that of the filter membrane of the lower separation tube and the lower filtration portion of the upper separation tube is received in the upper reception portion of the lower separation tube, and the bottom of the integrated tube is formed with a cone, the outer portion of which is sleeved with a sleeve.

2. The centrifugally integrated molecular fractionation apparatus of claim 1, wherein the first engagement and the third engagement have a recessed structure, and the second engagement has a raised structure complementary to the recessed structure.

3. The centrifugally integrated molecular fractionation apparatus of claim 2, wherein the depressed structure has a rectangular, trapezoidal, triangular or saw-toothed shape.

4. The centrifugally integrated molecular fractionation apparatus of claim 1, wherein the filtration membrane is a microfiltration membrane, an ultrafiltration membrane, a nanofiltration membrane or a reverse osmosis membrane.

5. The centrifugally integrated molecular fractionation apparatus according to claim 1, wherein the separator tube and the integration tube are each integrally formed by injection molding of PP plastic.

6. A centrifugally integrated molecular fractionation apparatus as claimed in claim 1, wherein said ribs and said notches are 2 to 4 in number.