GB2505223A - A filter retaining bucket - Google Patents

A filter retaining bucket Download PDF

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Publication number
GB2505223A
GB2505223A GB201215030A GB201215030A GB2505223A GB 2505223 A GB2505223 A GB 2505223A GB 201215030 A GB201215030 A GB 201215030A GB 201215030 A GB201215030 A GB 201215030A GB 2505223 A GB2505223 A GB 2505223A
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United Kingdom
Prior art keywords
filter
retaining bucket
open end
filter retaining
spin column
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
GB201215030A
Other versions
GB201215030D0 (en
GB2505223B (en
Inventor
Jolanta Vitkute
Greta Daujotaite
Jeffrey Leonard Coulling
Daniel Sims
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Thermo Fisher Scientific Lithuania Holding UAB
Original Assignee
Thermo Fisher Scientific Lithuania Holding UAB
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Thermo Fisher Scientific Lithuania Holding UAB filed Critical Thermo Fisher Scientific Lithuania Holding UAB
Priority to GB1215030.6A priority Critical patent/GB2505223B/en
Publication of GB201215030D0 publication Critical patent/GB201215030D0/en
Priority to DE201320006205 priority patent/DE202013006205U1/en
Priority to CN201320499619.2U priority patent/CN203803213U/en
Publication of GB2505223A publication Critical patent/GB2505223A/en
Application granted granted Critical
Publication of GB2505223B publication Critical patent/GB2505223B/en
Expired - Fee Related legal-status Critical Current
Anticipated expiration legal-status Critical

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M47/00—Means for after-treatment of the produced biomass or of the fermentation or metabolic products, e.g. storage of biomass
    • C12M47/06—Hydrolysis; Cell lysis; Extraction of intracellular or cell wall material
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5021—Test tubes specially adapted for centrifugation purposes
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5023—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures with a sample being transported to, and subsequently stored in an absorbent for analysis
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B04—CENTRIFUGAL APPARATUS OR MACHINES FOR CARRYING-OUT PHYSICAL OR CHEMICAL PROCESSES
    • B04B—CENTRIFUGES
    • B04B5/00—Other centrifuges
    • B04B5/04—Radial chamber apparatus for separating predominantly liquid mixtures, e.g. butyrometers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B04—CENTRIFUGAL APPARATUS OR MACHINES FOR CARRYING-OUT PHYSICAL OR CHEMICAL PROCESSES
    • B04B—CENTRIFUGES
    • B04B7/00—Elements of centrifuges
    • B04B7/08—Rotary bowls
    • B04B7/12—Inserts, e.g. armouring plates
    • B04B7/16—Sieves or filters
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/1003—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
    • C12N15/1017—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor by filtration, e.g. using filters, frits, membranes

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Organic Chemistry (AREA)
  • Biomedical Technology (AREA)
  • Genetics & Genomics (AREA)
  • Biotechnology (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • Microbiology (AREA)
  • Analytical Chemistry (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Sustainable Development (AREA)
  • Cell Biology (AREA)
  • Physics & Mathematics (AREA)
  • Biophysics (AREA)
  • Plant Pathology (AREA)
  • Hematology (AREA)
  • Clinical Laboratory Science (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)

Abstract

A filter retaining bucket 40 comprises a hollow body (42, fig 2), a first open end 44 and a second open end 50. The first open end 44 has a smaller diameter than the second open end 50 and the hollow body (42, fig 2) includes a tapered portion 48 extending from the second open end 50 towards the first open end 44 and a tubular portion 46 extending from the first open end 44 and towards the second open end 50 wherein the tubular portion 46 is configured to retain a filter 60, 62, 64, 66. The filter 60, 62, 64, 66 preferably comprises a silica membrane and the filter retaining bucket 40 is preferably for use in a spin column 10.

Description

Spin Column
Field of the Invention
The present invention relates generally to spin columns and, more particularly, to spin columns for use in DNA and/or RNA purification and concentration from enzymatic reaction mixtures and/or low concentration samples.
Background of the Invention
The isolation of nucleic acids from complex biological starting materials in the art is typically carried out under strongly denaturing and reducing conditions. During the isolation the starting materials containing the nucleic acids are partially solubilised using protein-degrading enzymes. This releases the nucleic acids, which can then be extracted into phenol and/or chloroform. The nucleic acids can then typically be recovered from the aqueous phase by using techniques such as dialysis or ethanol precipitation.
The problem with such isolation techniques is that they are time consuming, necessitate a considerable outlay of expense on apparatus and have been shown to be hazardous to health.
Alternative methods for the isolation of nucleic acids from different biological starting materials have been developed such as the use of agarose gels. In this method the agarose containing the nucleic acids to be isolated is dissolved in a solution of a chaotropic salt followed by bonding of the nucleic acids to particles of glass. The nucleic acids are then washed and eluted from the particles of glass.
A number of reagent kits have been developed for the purpose of isolating nucleic acids based on the principle of bonding nucleic acids to mineral carriers (such as a silica gel powder or glass fibre powder) under the presence of different chaotropic salts. Using such reagent kits small sample quantities can be used which is particularly relevant for the isolation of viral nucleic acids.
In order to provide the user with a more convenient method, a "single tube" method was developed in which the starting material combined with the chaotropic salt is placed in a spin column containing a firmly inserted glass fibre filter or silica layer allowing the nucleic acids to bond to the glass or silica in a single step.
The use of spin columns for nucleic acids purification is based on the general method; nucleic acids are first isolated from a certain volume of lysate, and are later concentrated via binding the nucleic acids onto a comparatively small volume of solid phase affinity resin. For example, when using traditional standard sized mini columns about SOOpI of initial solution (comprising lysed sample of about 200pl and the remainder being binding buffer) is loaded onto about 50-lOOp1 of a silica filter or any other suitable solid binding resin. Using this method the volume of binding resin determines the volume of elution buffer. The volume of elution buffer should be at least the same as the volume of the binding resin itself (in this particular case about lOO-200pl) as the use of a smaller volume of elution buffer negatively affects the nucleic acid yield.
When there is a need to concentrate nucleic acids from comparatively large volumes of initial biological material during purification process, smaller volumes of solid binding phase should be used in combination with a large volume capacity column to accommodate the larger volumes of initial sample and binding buffer.
The problem is that the spin columns can be quite large compared to some sample volumes to be isolated. However due to the standard sizes of spin columns it is not possible to simply make a smaller vessel as such vessels cannot be fitted into standard centrifuges.
One such solution to this problem involves the use of a plastic funnel structure that covers part of the filter thereby limiting the active surface area of the filter and thus its exposure to the lysed sample. However, the liquid is able to spread out sideways through the width of the filter and thus the active volume of the filter is not totally reduced.
At present when there is a need to decrease elution volume and, therefore, the filter volume, two approaches are currently used in the industry. In the first technique, the approximate 7mm diameter filter that is fitted into the mini spin column is covered by a funnel shaped ring, the inner diameter of which is significantly smaller than the outer diameter, thereby reducing the active surface area of the filter that comes into contact with the sample as mentioned above. A further disadvantage of such method is that part of sample material may diffuse throughout all the filter and become eliminated from further elution procedures. In the second technique a glass fibre filter is inserted into an adaptor portion, instead of into the cylindrical portion, of a spin column, which is used for inserting the spin column into a vacuum manifold.
As the diameter of such an adaptor portion is only about 2mm the active surface area of the filter is reduced, however, there is an increased risk of the sample becoming contaminated due to contact with the vacuum manifold.
Summary of the Invention
The present invention, namely, the use of an insertable plastic bucket with desired solid phase filter parameters solves the above mentioned problems and may be used for the whole range of standard spin columns (mini, midi, maxi) as it allows for both purification and concentration of nucleic acids from various amounts of starting materials using various volumes of reagents and binding agents to release the nucleic acid onto desired amount of solid affinity resin. Using such an insertable plastic bucket achieves the desired nucleic acid concentration within the required range for downstream application without the need to perform an additional concentration step.
Usually a certain ratio of the volume of the starting sample and the volume of the lysis and/or binding solutions is required to ensure good cell lysis and to improve the nucleic acid yield. Thus, it is not always possible to decrease the volume of lysed material prior to loading of the sample onto a column that matches the required degree of nucleic acid concentration after elution. In such cases an insertable bucket serves as convenient solution to this problem.
According to a first aspect of the present invention there is provided a filter retaining bucket, comprising: a hollow body, a first open end and a second open end; the first open end having a smaller diameter than the second open end; the hollow body comprising a tapered portion extending from the second open end towards the first open end and a tubular portion extending from the first open end and towards the second open end wherein the tubular portion is configured to retain a filter.
Preferably the filter retaining bucket is configured to be used in conjunction with a spin column. The spin column may be a maxi spin column, a midi spin column or a mini spin column. A maxi spin column is configured to fit into a 50m1 collection tube and typically has about a 1 -3m1 elution volume when fitted with a standard about 26mm diameter and about 2-4mm deep filter. Glass fibre filters of various characteristics (pore size, thickness, number of filter layers) may be used to achieve the required active filter volume and liquid flow-through. To avoid loss of the bound nucleic acid the elution buffer volume used in such column has to be at least the same volume as the volume of the filter.
A midi spin column is configured to fit into a 15m1 collection tube and typically has about 0.5-imI elution volume when fitted with a standard about 12mm diameter and about 2-4mm deep filter. A mini spin column is configured to fit into a 2m1 collection tube and typically has about a 50-200pl elution volume when fitted with a standard about 7mm diameter and about 2-4mm deep filter.
Use of buckets of various size and design in combination with standard maxi, midi and mini columns disclosed in this invention provides for the user flexibility in varying diameter of glass fibre filter depending on their needs.
Preferably the first open end is provided with a cross bar extending across the opening.
Preferably the first open end is provided with two cross bars extending across the opening.
Preferably the internal diameter of the tubular portion is about 11.9mm and is configured to retain a filter of about 12mm in diameter, in the case of a maxi spin column filter retaining bucket.
Preferably the internal diameter of the tubular portion is about 6.9mm and is configured to retain a filter of about 7mm in diameter, in the case of a midi spin column filter retaining bucket.
Preferably the internal diameter of the tubular portion is about 5.9mm and is configured to retain a filter of about 6mm in diameter, in the case of a mini spin column filter retaining bucket.
Preferably the internal diameter of the tubular portion is about 2.24mm and is configured to retain a filter of about 2.35mm in diameter, in the case of a mini spin column filter retaining bucket for micro elution.
Preferably the tapered portion is angled at about 60 to 90° to the horizontal, more preferably the tapered portion is angled at about 70 to 8O°to the horizontal.
According to a second aspect of the invention there is provided a filter retaining bucket assembly comprising a filter retaining bucket as described above and a filter.
Preferably the filter has a volume of about 200-500pl, in the case of a maxi spin column filter retaining bucket.
Preferably the filter has a volume of about 50-150p1, in the case of a midi spin column filter retaining bucket.
Preferably the filter has a volume of about 50-lOOpI, in the case of a mini spin column filter retaining bucket.
Preferably the filter has a volume of about 8-l5pl, in the case of a mini spin column filter retaining bucket for micro elution.
Preferably the filter comprises a silica membrane.
According to a third aspect of the present invention there is provided a spin column assembly comprising a filter retaining bucket assembly as described above and a spin column.
According to a fourth aspect of the present invention there is provided a spin column assembly comprising a filter retaining bucket as described above and a spin column.
Preferably the spin column assembly further comprises a collection tube.
Spin columns with filter retaining buckets comprising silica filters enables the use of a minimal amount of silica in the range of about 200-500pl, in the case of a maxi spin column with a filter retaining bucket. Due to the minimal amount of silica used this results in a decreased surface area, capacity and volume. This is turn allows for smaller elution volumes in the range of about from equal to twice as large as the volume of glass fibre filter used, for example between about 200-l000pl in the case of a maxi spin column fitted with a filter retaining bucket. The filter retaining bucket can be used during DNA and RNA purification, concentration from enzymatic reaction mixtures, low concentration samples and agarose gels.
Spin columns with filter retaining buckets comprising silica filters enables the use of a minimal amount of silica in the range of about 50-i5Opl, in the case of a midi spin column with a filter retaining bucket. Due to the minimal amount of silica used this results in a decreased surface area, capacity and volume. This is turn allows for small elution volumes in the range of about 50-300pl in the case of a midi spin column with a filter retaining bucket. The filter retaining bucket can be used during DNA and RNA purification, concentration from enzymatic reaction mixtures, low concentration samples and agarose gels.
Spin columns with filter retaining buckets comprising silica filters enables the use of a minimal amount of silica in the range of about SO-lOOpI, in the case of a mini spin column with a filter retaining bucket. Due to the minimal amount of silica used this results in a decreased surface area, capacity and volume. This is turn allows for small elution volumes in the range of about 50-200pl in the case of a mini spin column with a filter retaining bucket. The filter retaining bucket can be used during DNA and RNA purification, concentration from enzymatic reaction mixtures, low concentration samples and agarose gels.
Spin columns with micro elution filter retaining buckets comprising silica filters enables the use of a minimal amount of silica in the range of about 8-isp1, in the case of a mini spin column with a micro elution filter retaining bucket. Due to the minimal amount of silica used this results in a decreased surface area, capacity and volume. This is turn allows for small elution volumes in the range of about 8-30pl in the case of a mini spin column with a micro elution filter retaining bucket. The filter retaining bucket can be used during DNA and RNA purification, concentration from enzymatic reaction mixtures, low concentration samples and agarose gels.
Brief Description of the Drawings
The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and, together with a general description of the invention given above, and the detailed description of the embodiments given below, serve to explain the principles of the invention.
Figure 1 is an exploded view of a mini spin column housing the micro elution filter retaining bucket of the present invention; Figure 2 is a top perspective view of the micro elution filter retaining bucket of the present invention; Figure 3 is a bottom perspective view of the micro elution filter retaining bucket of the present invention; Figure 4 is a side view of the micro elution filter retaining bucket of the present invention; Figure 5 is a cross sectional view of the micro elution filter retaining bucket about section line AA of Figure 4 without any filters, with approximate dimensions; Figure 6 is a cross sectional view of the micro elution filter retaining bucket about section line AA of Figure with 2 assembled filters, with approximate dimensions; Figure 7 is a flow diagram illustrating nucleic acid isolation using the micro elution filter retaining bucket of the present invention; Figure 8 illustrates an embodiment of the filter retaining bucket of the present invention fitted into a maxi spin column, with typical dimensions; Figure 9 illustrates an embodiment of the filter retaining bucket of the present invention fitted into a midi spin column, with typical dimensions; Figure 10 illustrates an embodiment of the filter retaining bucket of the present invention fitted into a mini spin column, with typical dimensions; Figure 11 illustrates a perspective view of an embodiment of the filter retaining bucket of the present invention configured for use with a mini spin column; Figure 12 illustrates a front view of an embodiment of the filter retaining bucket of the present invention configured for use with a mini spin column, with approximate dimensions; and Figure 13 illustrates a cross-sectional view of an embodiment of the filter retaining bucket of the present invention configured for use with a mini spin column, with approximate dimensions.
Detailed Description of the Invention
Turning now to the Figures. An exploded view of an exemplary micro spin column 10 housing an embodiment of the filter retaining bucket 40 is shown and described in detail in Figure 1. The micro spin column 10 includes a hollow body 12 having a first open end 14, a generally tubular side wall 16 extending from the first open end 14 to a second open end 18, and a cap 20 configured to close over the second open end 18 to seal the second open end 18 in agenerallyfluid tight manner. The cap 20 is formed integrally with the hollow body 12 during a moulding operation and connected thereto by a flexible strap or tab 21. The first open end 14 is provided with a tapered side wall 22 which reduces the diameter of the first open end 14 compared to the diameter of the second open end 18. The first open end 14 is also provided with a generally tubular side wall 24 which is of reduced diameter compared to the tubular side wall 16 of hollow body 12. The first open end 14 is further provided with an inward lip 26 which further reduces the diameter of the opening 28 of the first open end 14 compared to the diameter of the opening 50 of the second open end 18. The micro spin column 10 is formed by injection moulding from thermoplastic materials.
The filter retaining bucket 40 can be seen in more detail in Figures 2 to 6.
The filter retaining bucket 40 is configured to locate at the first open end 14 of the micro spin column 10 and rest on the inward lip 26 in use. The filter retaining bucket 40 includes a hollow body 42 having a first open end 44, a generally tubular sidewall 46 extending generally the same distance as generally tubular side wall 24 of micro spin column 10, a tapered internal side wall 48 extending the remaining distance to a second open end 50, a generally tubular external side wall 52 extending from the second open end 50 generally the same distance as the tapered internal sidewall 48. The second open end 50 having a sealing rim 54 configured to form a generally fluid tight seal with tubular side wall iSof hollow body 1201 micro spin column lOin use. The sealing rim 54 operates by being of slightly larger external diameter compared to the internal diameter of the tubular side wide 16 so that in use the tubular side wall 16 compresses the sealing rim 54 to hold the filter retaining bucket in place in the micro spin column 10. In an alternative the sealing rim 54 may form a generally fluid tight seal using an 0-ring or similar sealing mechanism.
Tubular external sidewall 52 is configured to generally have its external diameter correspond with the internal diameter of tubular sidewall 16 of hollow body 12 of micro spin column 10. Tubular sidewall 46 is configured to generally have its external diameter correspond with or be smaller than the internal diameter of tubular side wall 24 of the first open end 14 of hollow body 12 of micro spin column 10 to effectively reduce the internal volume of the first open end 14 of hollow body 12 of micro spin column 10. The first open end 44 of the filter retaining bucket 40 has an opening 56 having cross bars 58. The filter retaining bucket 40 is provided with at least one filter 60, 62, 64, 66 configured to have a diameter generally corresponding to the internal diameter of tubular side wall 46 of the first open end 44 of the filter retaining bucket 40, to cover opening 56 and locate in position resting on cross bars 58. Furthermore in the alternative where the sealing rim 54 is compressed this also acts to compress tubular side wall 46 which assists in retaining the at least one filter 60, 62, 64, 66 in position in the filter retaining bucket 40. The filter retaining bucket 40 is formed by injection moulding from thermoplastics materials.
The tapered internal side wall 48 is to be angled at generally about 700 to the horizontal.
The filter 60, 62, 64, 66 is preferably generally about 2.35-2.45mm in diameter, and formed from silica with a volume of generally 2-10 mm3 or a few p1. The filter 60, 62, 64, 66 may be formed from silica or in the alternative glass fibre or other suitable binding substrate.
Figures 8 to 10 illustrate alternative embodiments of the filter retaining bucket, 140, 240, 340 inserted into various sizes of spin column 110, 210, 10.
Figures 11 to 13 illustrate an alternative embodiment of the filter retaining bucket 440 configured for use with a mini spin column 10. In this embodiment the filter retaining bucket 440 is configured to retain a greater volume of filter compared with the filter retaining bucket 40 illustrated in Figures 1 to 6.
The filter retaining bucket 440 is configured to locate at the first open end 14 of the micro spin column 10 and rest on the inward lip 26 in use. The filter retaining bucket 440 includes a hollow body 442 having a first open end 444, a generally tubular sidewall 446 extending generally the same distance as generally tubular side wall 24 of micro spin column 10, and a tapered internal side wall 448 extending the remaining distance to a second open end 450. The second open end 450 having a sealing rim 454 configured to form a generally fluid tight seal with tubular side wall 16 of hollow body 12 of micro spin column 10 in use. The sealing rim 454 operates by being of slightly larger external diameter compared to the internal diameter of the tubular side wide 16 so that in use the tubular side wall 16 compresses the sealing rim 54 to hold the filter retaining bucket 440 in place in the micro spin column 10. In an alternative the sealing rim 454 may form a generally fluid tight seal using an o-ring or similar sealing mechanism. Tubular sidewall 446 is configured to generally have its external diameter correspond with or be smaller than the internal diameter of tubular side wall 24 of the first open end 14 of hollow body 12 of micro spin column 10 to effectively reduce the internal volume of the first open end 14 of hollow body 12 of micro spin column 10. The first open end 444 of the filter retaining bucket 440 has an opening 456 having cross bars 458. The filter retaining bucket 40 is provided with at least one filter (not shown) configured to have a diameter generally corresponding to the internal diameter of tubular side wall 446 of the first open end 444 of the filter retaining bucket 440, to cover opening 456 and locate in position resting on cross bars 458. Furthermore in the alternative where the sealing rim 454 is compressed this also acts to compress tubular side wall 446 which assists in retaining the at least one filter (not shown) in position in the filter retaining bucket 440. The filter retaining bucket 440 is formed by injection moulding from thermoplastics materials.
The tapered internal side wall 448 is to be angled at generally about 800 to the horizontal.
Example
Nucleic acid purification from enzymatic reaction mixtures and concentration of low concentration nucleic acid solutions using micro amounts of silica filter in the form of a membrane.
Figure 7 illustrates a flow diagram of a typical purification scheme utilizing the filter retaining bucket 40 of the present invention.
The enzymatic reaction, gel slice or low concentration nucleic acid sample is mixed with the binding buffer. It may be necessary to add isopropanol or ethanol to the binding buffers. The solution is then transferred to a micro spin column 10 with a filter retaining bucket 40 and silica filters 60, 62, 64, 66 housed within a collection tube to collect any flow-through, and centrifuged for 1510 30 seconds. During this process the nucleic acids become bound to the silica filters 60, 62, 64, 66. The f low-through collected in the collection tube is then discarded. The silica filters 60, 62, 64, 66 whilst still in situ are washed with a wash buffer and centrifuged for 15 to 30 seconds, again the flow-through is discarded. The micro spin column 10 complete with filter retaining bucket 40 and silica filters 60, 62, 64, 66 is then further centrifuged for another 15 to 30 seconds to remove again remains of the solution.
The complete micro spin column 10 is then transferred to a clean collection tube and the nucleic acids are eluted using 6-20F.il elution buffer or nuclease-free water which is applied in the centre of the column. The complete micro spin column 10 then undergoes centrifugation for one minute and the flow-through is collected and retained as it now comprises only pure and concentrated nucleic acids.
While the present invention has been illustrated by a description of an embodiment, and whilst this embodiment has been described in some detail, this is not intended in any way to restrict or in any way limit the scope of the appended claims to such detail. Additional advantages and modifications will readily appear to those skilled in the art. The various features of the invention may be used alone or in combination depending on the needs and preferences of the user. This has been a description of the present invention, along with methods of practising the invention as currently known. However, the invention itself should only be defined by the appended claims.

Claims (22)

  1. Claims 1. A filter retaining bucket, comprising: a hollow body, a first open end and a second open end; the first open end having a smaller diameter than the second open end; the hollow body comprising a tapered portion extending from the second open end towards the first open end and a tubular portion extending from the first open end and towards the second open end wherein the tubular portion is configured to retain a filter.
  2. 2. A filter retaining bucket as claimed in Claim 1 configured to be used in conjunction with a spin column.
  3. 3. A filter retaining bucket as claimed in Claim 1 or Claim 2 wherein the first open end is provided with a cross bar extending across the opening.
  4. 4. A filter retaining bucket as claimed in any preceding claim wherein the first open end is provided with two cross bars extending across the opening.
  5. 5. A filter retaining bucket as claimed in any preceding claim wherein the internal diameter of the tubular portion is about 11.9mm and is configured to retain a filter of about 12mm in diameter.
  6. 6. A filter retaining bucket as claimed in any of claims 1 to 4 wherein the internal diameter of the tubular portion is about 6.9mm and is configured to retain a filter of about 7mm in diameter.
  7. 7. A filter retaining bucket as claimed in any of claims 1 to 4wherein the internal diameter of the tubular portion is about 5.9mm and is configured to retain a filter of about 6mm in diameter.
  8. 8. A filter retaining bucket as claimed in any of claims 1 to 4 wherein the internal diameter of the tubular portion is about 2.24mm and is configured to retain a filter of about 2.35mm in diameter.
  9. 9. A filter retaining bucket as claimed in any preceding claim wherein the tapered portion is angled at about 6010 90° to the horizontal.
  10. 10. A filter retaining bucket assembly comprising a filter retaining bucket as claimed in any of claims ito 9 and a filter.
  11. 11. A filter retaining bucket assembly as claimed in Claim 10 wherein the filter has a volume of about 200-500pl.
  12. 12. A filter retaining bucket assembly as claimed in Claim 10 wherein the filter has a volume of about 50-l5Opl.
  13. 13. A filter retaining bucket assembly as claimed in Claim 10 wherein the filter has a volume of about 50-lOOp1.
  14. 14. A filter retaining bucket assembly as claimed in Claim 10 wherein the filter has a volume of about 8-15pl.
  15. 15. A filter retaining bucket assembly as claimed in Claim 10 wherein the filter has a volume of about 2-lOpl.
  16. 16. A filter retaining bucket assembly as claimed in any of claims 10 to 15 wherein the filter comprises a silica membrane.
  17. 17. A spin column assembly comprising a filter retaining bucket assembly of any of Claims 10 to 16 and a spin column.
  18. 18. A spin column assembly comprising a filter retaining bucket of any of Claims 1 to 9 and a spin column.
  19. 19. A spin column assembly of Claim 17 or Claim 18 further comprising a collection tube.
  20. 20. A filter retaining bucket substantially as hereinbefore described with reference to the accompanying figures.
  21. 21. A filter retaining bucket assembly substantially as hereinbefore described with reference to the accompanying figures.
  22. 22. A spin column substantially as hereinbefore described with reference to the accompanying figures
GB1215030.6A 2012-08-23 2012-08-23 Spin column Expired - Fee Related GB2505223B (en)

Priority Applications (3)

Application Number Priority Date Filing Date Title
GB1215030.6A GB2505223B (en) 2012-08-23 2012-08-23 Spin column
DE201320006205 DE202013006205U1 (en) 2012-08-23 2013-07-09 Spin column
CN201320499619.2U CN203803213U (en) 2012-08-23 2013-08-15 Filter holding barrel, filter holding barrel component and rotating column component

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
GB1215030.6A GB2505223B (en) 2012-08-23 2012-08-23 Spin column

Publications (3)

Publication Number Publication Date
GB201215030D0 GB201215030D0 (en) 2012-10-10
GB2505223A true GB2505223A (en) 2014-02-26
GB2505223B GB2505223B (en) 2014-11-19

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GB1215030.6A Expired - Fee Related GB2505223B (en) 2012-08-23 2012-08-23 Spin column

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CN105482994B (en) * 2016-01-19 2018-02-16 浙江硕华医用塑料有限公司 A kind of cell filter
CN111283423B (en) * 2020-03-23 2020-12-15 杭州博日科技股份有限公司 Automatic assembling equipment for centrifugal column

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US20020192656A1 (en) * 2001-06-15 2002-12-19 Richardson Derrick A. Filtration column devices and methods of filtering therewith
US20050026159A1 (en) * 2003-07-31 2005-02-03 Robbins Claudia A. Devices and methods for isolating RNA
US20080020446A1 (en) * 2006-07-22 2008-01-24 Xiyu Jia Plasmid DNA isolation
US20100009832A1 (en) * 2005-09-26 2010-01-14 Andreas Schaefer Method for Processing a Fluid and Fluid Processing Device

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US20020192656A1 (en) * 2001-06-15 2002-12-19 Richardson Derrick A. Filtration column devices and methods of filtering therewith
US20050026159A1 (en) * 2003-07-31 2005-02-03 Robbins Claudia A. Devices and methods for isolating RNA
US20100009832A1 (en) * 2005-09-26 2010-01-14 Andreas Schaefer Method for Processing a Fluid and Fluid Processing Device
US20080020446A1 (en) * 2006-07-22 2008-01-24 Xiyu Jia Plasmid DNA isolation

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