Method for purifying and testing biomolecules from biological samples

Abstract

The present disclosure relates to, inter alia, an easy-to-operate, fully closed component, which can be part of an instrument, for purification of biomolecules from biological samples, and subsequent transfer, and testing of the biomolecules, as well as an instrument comprising the component, and a method for using the component.

1 - 17 . (canceled) 
     
     
         18 . A method for purifying a biomolecule from a biological sample, comprising:
 (a) providing a magnetic field and a fully-closed component for purifying one or more biomolecule(s) from a biological sample,   said component comprising: a plurality of mixing chambers; a plurality of connection tubes; a plurality of storage containers; a plurality of pierceable sealing membranes; a plurality of pressure seals or switches; purified sample collection tubes or testing chambers; magnetic beads; lysis buffer and/or binding buffer; washing solution; and eluent;   wherein:   each mixing chamber is a plastic container, the mixing chambers being connected to each other by connection tubes, which are switches or sealable tubes; each mixing chamber is located adjacently and horizontally or adjacently and vertically to the next mixing chamber;   the pressure seals or switches are adapted for opening and/or closing the connection tubes, to enable or disable liquid exchange between adjacent mixing chambers, and are located on and around the middle of each connection tube;   one or more of the mixing chambers are connected with one or more storage containers physically but not in liquid contact, with a pierceable sealing membrane between the connected mixing chamber and the storage container, each said storage container is located above or below the mixing chamber the storage container is connected to;   the pierceable sealing membrane of a storage container is adapted to prevent the transfer of a liquid, solid, or solid-liquid suspension in the storage container to the mixing chamber before the membrane is pierced;   when there is more than one storage container mounted on a mixing chamber or a testing chamber, they are located next to each other;   each storage container comprises a puncture device;   one or more storage containers comprise washing solution;   one or more storage containers optionally comprise eluent;   one or more storage containers comprise lysis buffer and/or binding buffer;   one or more storage containers comprise magnetic beads;   one or more mixing chambers are connected to one or more testing chambers or purified sample collection tubes, located above, below or adjacent to said one or more mixing chambers, each of said one or more mixing chambers is connected to each of said one or more testing chambers or purified sample collection tubes by a connection tube;   one or more of said plurality of storage containers are attached to one or more of said plurality of testing chambers or one or more of said plurality of mixing chambers comprising testing solution or dry powder of claim  1 , either by itself or housed in a machine;   (b) Introducing a biological sample to the component, by placing the biological sample in a first storage container, compressing the first storage container such that the puncture device damage the structural integrity of the sealing membrane, thereby allowing the biological sample, magnetic beads, and lysis/binding buffer to be mixed in the connected mixing chamber (first mixing chamber), which is the leftmost or the rightmost chamber of the component, wherein the magnetic beads and/or lysis/binding buffer, which can be either a binding buffer or a lysis buffer, or both, are introduced into the first mixing chamber either from the first storage container with the sample or another storage container in physical contact with the first mixing chamber and upon compressing that storage container, the puncture device damage the structural integrity of the sealing membrane, allowing the magnetic beads and lysis/binding buffer to enter into the first mixing chamber;   (c) mixing the biological sample, magnetic beads and lysis/binding buffer in the solution by rotation or vibration;   (d) opening pressure seal (first pressure seal) between the first and an adjacent mixing chamber (second mixing chamber) thereby connecting the two mixing chambers through the connection tube in between them (first connection tube);   (e) placing the magnetic field below the second mixing chamber comprising the sample and the solution to capture and thus enrich for the magnetic beads;   (f) tilting the sealed component, so that the liquid flows into the first mixing chamber; the magnetic beads remaining in the second mixing chamber due to the magnetic field;   (g) Closing the first pressure seal, thus closing the first connection tube, and stopping the liquid flow between the first and the second mixing chambers;   (h) Compressing a storage container (the second storage container) connected to the second mixing chamber, so the puncture device damages the integrity of sealing membrane, thereby introducing the washing solution in the second storage container (the second storage container comprises a washing solution) into the second mixing chamber such that the magnetic beads and washing solution are allowed to be mixed in the second mixing chamber and the magnetic beads are washed by rotation or vibration;   (i) Opening a pressure seal between the second mixing chamber and an adjacent mixing chamber (third mixing chamber) that is not the first mixing chamber, and connecting the second and third mixing chambers through a connection tube between them (second connection tube);   (j) Moving the magnetic field to below the third mixing chamber to capture and enrich for the magnetic beads;   (k) Tilting the sealed component, so that the liquid flows into the second mixing chamber;   (l) Resetting the second pressure seal, and closing the second connection tube to stop the liquid flow between the second and third mixing chambers;   (m) Optionally repeating washing steps (d)-(l) to remove impurities in additional mixing chamber(s) adjacent to the third mixing chamber but distally to the second mixing chamber;   (n) After washing, the magnetic beads are enriched in the second to last mixing chamber in which the final washing was performed and closing the connection tube between this mixing chamber and the one adjacent to it (third to last mixing chamber) from which the magnetic beads resided prior to residing in the second to last mixing chamber, the second to last mixing chamber is more distal to the first mixing chamber than the third to last mixing chamber;   (o) Compressing a storage container above the second to last mixing chamber (second to last storage container), which container comprises the eluent, so the puncture device damages the structural integrity of sealing membrane, thereby the magnetic beads and eluent are mixed and the bound materials on the magnetic beads are eluted from magnetic beads by rotation or vibration;   (p) Moving the magnetic field to the second to last mixing chamber to capture and enrich for the magnetic beads;   (q) Tilting the sealed component, opening pressure seal (second to last pressure seal) between the second to last mixing chamber and the last mixing chamber, and connecting the second to last mixing chamber and the last mixing chamber with the opened connection tube between these two mixing chambers (second to last connecting tube), so that the liquid is allowed to flow from the second to last mixing chamber into the last mixing chamber;   (r) Resetting the second to last pressure seal, and closing the second to last connection tube to stop the liquid flow between the second to last mixing chamber and the last mixing chamber;   (s) Compressing a storage container (last storage container) comprising a test solution/dry powder connected to the last mixing chamber, so the puncture device damages the structural integrity of sealing membrane, thereby allowing the purified sample to be mixed with test solution/dry powder by rotation or vibration;   (t) Opening pressure seal (last pressure seal), and connecting the last mixing chamber and testing chamber through the last connection tube, the test chamber being the most distal chamber from the first mixing chamber, then keep the sealed component tilt, so that the liquid flows into the testing chamber for testing.   (u) The purified sample can also be collected in a purified sample collection tube for storage after biomolecules are eluted from magnetic beads (step p), without the addition of test solution/dry powder or testing;   (v) wherein some aforementioned steps may be omitted, such as no elution step or testing step; the method can comprise more steps, such as addition of enzymes, antibody solution, substrate solution and/or additional incubations; and   wherein there can be more or less numbers of mixing chambers, connection tubes, storage containers, sealing membranes, pressure seals or switches, and purified sample collection tubes or testing chambers, more biological samples; more magnetic beads, lysis/binding buffer, washing solution, optionally eluent, and optionally test solution can be added, optionally a sample collection device with a sealing cap can be added, optionally a heater/cooler can be added, and optionally a detection sensor can be added.   
     
     
         19 . The method of  claim 18 , wherein the component is housed inside a machine. 
     
     
         20 . The method of  claim 18 , further comprising conducting testing of said biomolecule in said component.