Abstract:
A device for concentrating and/or purifying macromolecules in liquids is disclosed wherein the concentration/purification is accomplished by a membrane separating concentration and filtration chambers, the membrane and chambers being compressively held in fluid-tight relationship by sliding engagement with a sleeve-like housing. A method for assembling such a device is also disclosed.

Description:
This is a 371 of PCT/EP00/00976, filed Feb. 08, 2000, and claims priority of the Swedish Application No. 9900530-8, filed Feb. 15, 1999. 
    
    
     BACKGROUND OF THE INVENTION 
     Many biotechnology laboratories and health care institutions use micro- and ultrafiltration methods for processing biological solutions. Examples include the use of such filtration to remove bacteria, to remove suspended solids and contaminants, to concentrate proteins and other macromolecules and to eliminate unwanted micromolecules such as salts. 
     Centrifugal forces, gas or liquid pressure or vacuum are typically used as the driving force to push or pull solvent and small micromolecules through the membrane while solute components larger than the molecular weight cut-off of the membrane are retained on the feed side of the membrane as retentate or concentrate. In most applications, the higher the pressure or vacuum exerted, the higher is the rate of filtration relative to the membrane surface area used. In general, high speed and small surface area membranes are preferred. 
     Typical such devices include a chamber for the sample to be processed and a chamber for the capture of the permeate/filtrate, the two chambers being in fluid communication through at least one common aperture across which a membrane such as a microporous ultrafiltration or reverse osmosis membrane is arranged. The membrane is typically sealed fluid-tight at its periphery to the surface at the periphery of the aperture of either the processing/concentration chamber or the permeate chamber, or to both. 
     The membrane is usually reinforced on its permeate side to withstand transmembrane pressure. There is a feed inlet provided for introducing a liquid sample to the concentration chamber and an outlet for the filtrate from the filtrate chamber. In so-called tangential flow devices there is an additional outlet provided in the concentration chamber to permit recirculation of the liquid sample. 
     The sealing of the membrane to the filtration device must meet very high requirements so that the liquid to be processed is prevented from by-passing the membrane. The membrane may be sealed in a variety of ways such as by heat-sealing, adhesive- or solvent-bonding, ultrasonic welding or by an interference fit. In addition, material and wall thickness for the chambers which are also sealed together are chosen in order to withstand the operating pressure. Such devices are sometimes also provided with separate pressure holders or membrane assemblies placed between external pressure plates, which are typically bolted together to provide additional support. 
     One of the problems with such prior art devices is the difficulty in obtaining a satisfactory compromise between, on the one hand, seal reliability for both the membrane and/or for the joint between the chambers to provide adequate pressure containment and, on the other hand, low manufacturing cost. 
     For practicality and cost reasons, it is desirable to mold such filtration devices but it is not always possible to mold membrane support sections of sufficient thickness to withstand high operating pressures because it is difficult to mix thick and thin areas when molding. The alternative of using separate pressure holders or external pressure plates is both expensive and inconvenient for small devices. Also, although sealing systems that perform well at low to moderate pressures are frequently unreliable at higher pressures. 
     Another problem is achieving a sufficiently strong seal between the chambers, particularly with incompatible or non-sealing materials and in large surface area devices that must withstand higher overall pressures. 
     The design of one such centrifugal filtration device is described in U.S. Pat. No. 5,647,990, which discloses a half cylindrical sleeve to retain the filtration membrane in place over the aperture between the concentration and filtrate chambers. A principal drawback of that design is its potential for damaging the relatively fragile membrane when the retaining sleeve is pushed over the concentration chamber, since the resultant frictional forces tend to push the membrane out of alignment with the aperture. Another drawback of said design is that pressure containment and seal integrity are limited by the difficulty of molding a sufficiently thick membrane support plate and the incomplete support provided to the sealing area of the membrane due to permeate outlet passages that directly cross the seal area. The problems caused by frictional forces on the membrane and incomplete seal support are further exacerbated when the membrane is not first sealed to the concentration chamber, but sealing and assembly rather are effected in a single operation by compression during application of the perimeter of the membrane to the aperture of the concentration chamber. 
     UK Application No. 9819686.8 discloses a so-called tangential flow filtration device. The drawback of this device is the need to machine its component parts due to the difficulty of molding them sufficiently thick to contain high pressures. In addition, relatively expensive bolting mechanisms are required to hold the assembled device together sufficiently tightly to resist high operating pressures. 
     One object of the present invention is to provide a device of the type mentioned above which is simple to manufacture while providing increased overall reliability. 
     A further object is to provide a device for which it is possible to choose materials for the retentate and permeate compartments which do not require heat or ultrasonic seal compatibility with each other nor with the membrane used. 
     A further object is to provide a device which has a supported seal around the entire periphery of the membrane. 
     A further object of the invention is to provide a device which is possible to reopen and reclose, after processing in order to inspect and/or replace the membrane without damaging the retentate or permeate compartments or the membrane. 
     A still further object is to provide a method according to which the membrane will be sealed to the filtration device during assembly in a single operation. 
     The foregoing objects, uses and advantages of this invention will become apparent from the following description. 
     BRIEF SUMMARY OF THE INVENTION 
     The invention provides a device for concentrating and/or purifying macromolecules in solution and a method for manufacturing such a device. The device comprises a processing/concentration chamber for the sample to be processed and a chamber for filtrate that is separated by at least one membrane and more specifically provides a novel method of fixing the membrane in a pressure-resistant sealing relationship in the interface between the two chambers. Such devices may be used in centrifuges, as tangential flow modules, as gas-pressurized cells or with hand-operated syringes. 
    
    
     
       BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWINGS 
         FIG. 1  is a perspective view of an exemplary device of the present invention in the form of a tangential flow module. 
         FIG. 2  is a cross-sectional view of the filtration insert portion of the device shown in FIG.  1 . 
         FIG. 3  is a side view of the device shown in FIG.  1 . 
         FIG. 4  is a plan view of the device shown in  FIG. 1 , as assembled. 
         FIG. 5  is a cross-sectional view of the assembled device shown in FIG.  4 . 
         FIG. 6  is a plan view of an upper portion of the filtration insert portion of the device shown in FIG.  1 . 
         FIG. 7  is a plan view of a lower portion of the filtration insert portion of the device shown in FIG.  1 . 
         FIG. 8  is a plan view of a modification of the upper portion of the device shown in FIG.  6 . 
         FIG. 9  is an exploded perspective view of another exemplary embodiment of the present invention in the form of a centrifugal filtration device. 
         FIG. 10  is an exploded perspective view of the device shown in  FIG. 9  illustrating its assembly. 
         FIG. 11  is a perspective view of the device shown in  FIG. 9 , fully assembled. 
         FIG. 12   a  is a cross-sectional view of the assembled device shown in FIG.  11 . 
         FIG. 12   b  is a cross-sectional view of the assembled device shown in  FIG. 11  provided with a variant of the sleeve component of that device. 
         FIGS. 13   a  and  13   b  are two horizontal cross-sectional views taken through the assembled device shown in  FIG. 12   a  and  12   b , respectively. 
         FIG. 14  is an exploded perspective view of an exemplary device of the present invention adapted for the filtration of multiple, samples. 
         FIG. 15  is a cross-sectional view of the device shown in FIG.  14 . 
     
    
    
     DETAILED DESCRIPTION OF THE INVENTION 
     Referring now to the drawings, wherein the same numerals generally refer to like elements, there is shown in  FIG. 1  an exploded perspective view of a device according to the present invention in the form of a tangential flow filtration module or cell which can be used, for example, for concentrating and/or fractionating macromolecules in a liquid. In a filtration system this type of filtration module is connected to a pump that typically draws liquid from a reservoir of liquid sample material and pumps it through the module and recirculates liquid sample through a loop that includes the module. The module is incorporated into and in fluid communication with this loop via feed inlet  3  and permeate outlet  4  arranged on filtration insert  1 . The necessary system pressure is created by a flow restrictor (not shown) positioned at permeate outlet  4  of the cell. 
     As shown in FIGS.  3  and  6 - 7  filtration insert  1  comprises a concentration (retentate) chamber  10  in the exemplary form of a thin channel ( FIG. 6 ) situated in upper plate  12  ( FIG. 2 ) of insert  1 . Filtrate (permeate) chamber  11  ( FIG. 7 ) is situated in membrane support plate  6  ( FIG. 2 ) of filtration insert  1 . Concentration and filtrate chambers  10  and  11  are separated from each other by membrane  15  (see FIG.  5 ). 
     Due to the pressure difference between the feed side and the permeate side of membrane  15 , filtrate permeates through membrane  15  and filtrate chamber  11  and exits via discharge port  5  for collection outside the cell. 
       FIG. 2  shows in section an exemplary filtration insert  1 . Membrane support plate  6  has a generally planar upper surface that functions as a membrane support  16 . Membrane support  16  is provided with a plurality of parallel grooves forming filtrate channels  9  for the collection of filtrate, also shown in FIG.  7 . Filtrate channels  9  are all in fluid communication with each other by means of transverse outlet channel  14  (shown in FIG.  5 ), that connects the end portions of filtrate channels  9  on one side of the filtration insert  1  in order to feed all the collected filtrate to discharge port  5 . 
     As shown in  FIG. 5 , membrane  15  is positioned on top of membrane support  6  of filtration insert  1  and extends to the vertical walls  13  of membrane support  6 . Along the inner side of vertical walls  13  a generally flat gasket seat  8  is arranged (see  FIGS. 2 ,  5 ,  7 ), making for a generally flat support for membrane  15  all around its periphery. Gasket  7 , in, the form of an O-ring in this embodiment, is arranged on top of membrane  15  over gasket seat  8  and upper plate  12  closes the filtration insert  1 . 
     A two-part sleeve  2 ,  2 ′, preferably manufactured by molding a suitable polymer, surrounds filtration insert  1 . In the embodiment shown in FIGS.  1  and  3 - 5  the two-part sleeve  2 , 2 ′ slidingly engages filtration insert  1  from opposing sides to form the complete sleeve housing  2 ″ and compresses insert  1  without any frictional forces on membrane  15 , thereby forming, in a single step, and without the use of any adhesives, bolts or tools, the concentration and filtrate chambers  10  and  11  by sealing the periphery of membrane  15 . An additional advantage of the module&#39;s construction is that it is easy to take apart for inspection, to change the membrane, for cleaning or for any other purpose without damaging any of its component parts. 
       FIG. 3  shows a side view of the embodiment shown in FIG.  1 . The sliding introduction of filtration insert  1  into the two sleeve parts  2  and  2 ′ starts very easily due to the tapered cross-section of the sleeve and insert, best seen in FIG.  5 . Greater force is only needed at the end of the assembly during compression of gasket  7 . Sleeve parts  2 ,  2 ′ and insert  1  may also be assembled by heating sleeve parts  2 ,  2 ′ before assembly and allowing them to cool. 
       FIG. 4  is a plan view of the assembled embodiment shown in FIG.  1 . In the lower portion of  FIG. 4  there is shown attached to sleeve  2  a longitudinal flange  17  adapted to matingly engage a corresponding slot at the upper portion of an adjacent module (not shown), thereby permitting a series of filtrating modules oriented on their edges to be stacked on top of each other. 
       FIG. 5  shows in section the assembled device shown in FIG.  4 . As noted above, the insides of the two-part sleeve  2 ,  2 ′ are tapered. In addition to being practical for manufacturing purposes, tapering permits the achievement of the necessary collaborating compressive forces for filtration insert  1 , the exterior of which has a corresponding taper. In order to achieve nearly the same compressive forces but with lower friction between sleeve housing  2 ″ and filtration insert  1  the top and bottom surfaces of filtration insert  1  could be made parallel and the tapered form could be realized by means of longitudinal tapered flanges on the top and bottom surfaces of at least one of them; such an arrangement is illustrated in  FIG. 8 , showing tapered flanges  18 . 
     An additional advantage to molding the sleeve housing  2 ″ in two parts  2 ,  2 ′ is that a standardized thread  180  may readily be formed during molding, the thread accommodating feed inlet  3  and permeate outlet  4 . 
       FIG. 9  shows an exploded perspective view of another embodiment of the device according to the invention in the form of a centrifugal filtration device. Concentration chamber  10  is provided for receiving the liquid sample to be processed. In this case the sample is not recirculated and so no outlet for the sample is provided therefrom, there being only a feed inlet  3 . Gasket  7  is positioned in gasket seat  8  around aperture  19  in the wall of concentration chamber  10 . A suitable membrane  15  is positioned over aperture  19 . As described above in connection with the embodiment shown in  FIG. 1 , a filtrate chamber is provided with a membrane support plate  6 , gasket seat  8  and a plurality of parallel grooves forming filtrate channels  9  for collection of the filtrate are created when support plate  6  is fixed against membrane  15 . Transverse outlet channel  14  is in fluid communication with filtrate channels  9  and with discharge ports  5 ′. Gas bleed  20  is provided to permit air or gas to escape from filtrate channels  9  prior to filtration. 
       FIG. 10  shows the device of  FIG. 9  partially assembled after the components  10 ,  17 ,  15  and  6  (shown in  FIG. 9 ) are assembled; a one-piece sleeve housing  2 ″ slidingly engages those assembled components in this embodiment, shown in FIG.  11 . Due to the tapered or conical form of the assembled filtrate chamber and the corresponding female portion of sleeve housing  2 ″ the filtrate chamber is forced against membrane  15  and compresses gasket  7  against concentration chamber  10 . It should be noted that membrane  15  will only be exposed to forces perpendicular to its surface when sleeve housing  2 ″ is in place. In this way a pressure sufficiently high to seal the membrane fluid-tight against concentration chamber  10  is created and will be maintained during centrifugation. 
       FIG. 12   a  shows in section the assembled device shown in FIG.  11 . At the upper part of concentration chamber  10  gas bleed  20  is in fluid communication with the outside, which may be utilized to evacuate air from filtrate channels  9  at the start of the filtration process. In this embodiment three discharge ports  5 ′ are provided for the discharge of filtrate, to be collected in a filtrate recovery tube (not shown), in which the device is inserted. Discharge ports  5 ′ are in fluid communication with transverse outlet channel  14  as in the embodiment shown in  FIG. 5 , connecting the end portions of filtrate channels  9  on one side of the filtrate compartment in order to feed all the filtrate collected to discharge ports  5 ′. The device is supported on flange  21  of sleeve housing  2 ″ against flange  22  of concentration chamber  10 , which means that the centrifugal forces acting on the device during centrifugation maintain sleeve housing  2 ″ in position with flange  21  abutting flange  22 . 
       FIG. 12   b  shows in section the assembled device shown in  FIG. 11  provided with a variant of sleeve housing  2 ″. In order to ensure sealing between gasket  7  and membrane  15  at the lower part of the device, sleeve housing  2 ″ has been made less flexible in this area by the addition of a stabilizing ring  23  of material around the opening in the lower portion of sleeve housing  2 ″. Stabilizing ring  23  can alternatively have a more flat configuration than the one shown in  FIG. 12   b . As filtrate channels  9  do not fall below the transverse outlet channel  14  in this embodiment it is possible to arrange a generally flat support for the permeate side of membrane  15  along its periphery, which will ensure a sealing effect all around. 
       FIGS. 13   a  and  13   b  show horizontal sections through the assembled device shown in  FIG. 12   a  at its upper and lower portions, respectively, whereby the tapered form of the membrane support plate  6  is clearly illustrated. 
     Yet another exemplary embodiment of the invention, for the simultaneous filtration of multiple liquid samples, is depicted in  FIGS. 14 and 15  wherein there is shown a filtration insert  1  housed in a two-part sleeve  2 ,  2 ′. Filtration insert  1  comprises membrane support plate  6 , gasket mat  7 , a plurality of flat circular membranes  15  and upper plate  12  with a plurality of tubular concentration chambers  10  and tubular filtrate chambers  11 . Tubular filtrate chambers  11  comprise tubular walls  30 , circumferential flange  31  and openings  28 . 
     Membrane support plate  6  includes a plurality of openings  24  therein which, in their number and arrangement, preferably correspond to the number and arrangement of receptacles  25  of a micro titer plate  26 . Openings  24  in support plate  6  can also be provided with a filter support, which in turn is in fluid communication with collection channels  8  and discharge ports  5 ′ for the discharge of filtrate through filtrate port  50 . It should be noted that collection chambers  8  are coextensive with the interior of filtrate chambers  11 . Gasket mat  7  is laid upon support plate  6  and has tubular filtrate chamber openings  28  aligned with and corresponding to the number of openings  24  in support plate  6 . For easy positioning of gasket mat  7  on support plate  6  and for a more secure sealing of membrane  15  against filtrate chambers  11 , partitions or webs  29  are installed surrounding openings  24  in support plate  6 . 
     The tubular walls  30  of filtrate chambers  11  have a larger inner diameter than that of openings  28  and are sized so that walls  30  may be provided with a circumferential sealant flange  31  constructed of sealant material. During assembly, circular membranes  15  are laid on the circumferential sealant flanges  31  of tubular walls  30  and tubular concentration chambers  10  are inserted within tubular walls  30 , with tubular concentration chambers  10  being open on both ends and connected to upper plate  12 . The outer circumferential surfaces of tubular concentration chambers  10  are congruent with the inner surfaces of tubular walls  30 . Tubular concentration chambers  10  serve in the first instance to receive liquid samples to be processed, and secondly to function in cooperation with gasket mat  7  to form a fluid-tight peripheral seal of membranes  15  between sealant flanges  31  and the lower open ends of tubular chambers  10 . Tubular walls  30  of tubular filtrate chambers  11  are pushed sufficiently far into tubular concentration chambers  10 , that their lowermost portions press against the periphery of membranes  15 . The two-part housing  2 ,  2 ′, is provided with feed inlets  3  on the upper side to receive the liquid samples, and on the underside with discharge ports  5 ′ for the filtrate, which outlets are aligned with openings  24  in support plate  6 . 
     During assembly of the apparatus, the sleeves  2 ,  2 ′ are pushed over the filtration insert  1  to form housing  2 ″. As this is done, sufficient compressive forces are created and maintained to hold membranes  15  fluid-tight against flanges  31 . To this end filtration insert  1  preferably has a conical outer profile, and housing  2 ″ has a corresponding conical inner profile, which two profiles upon fitting together, cooperate to create the necessary compressive forces. As shown in  FIG. 14 , support plate  6  and upper plate  12  are provided with lateral, wedge-shaped flanges  18 . 
     As shown in  FIG. 15 , when housing  2 ″ is closed, inlets  3  on the upper side of the housing align with the ends of tubular concentration chambers  10  through filtrate chamber openings  28 , with openings  24  in support plate  6  and with discharge ports  5 ′ of the underside of the housing  2 ″. 
     As also seen in  FIG. 15 , discharge ports  5 ′ on the underside of housing  2 ″ are equipped with filtrate discharge fittings  32 , which matingly engage with receptacles  25  of a micro titer plate  26 . 
     Many variants of the basic inventive idea are contemplated. For example, it could be advantageous in certain embodiments to instead situate gasket  7  between membrane  15  and filtrate chamber  11 . As mentioned above, devices according to the invention could be designed for use as a stackable tangential flow module useable singly or in series, as in the embodiments described with reference to  FIGS. 1-8 ; or for use in a centrifuge, as in the embodiments described with reference to  FIGS. 9-13 . Gas or liquid pressure or vacuum may be used to provide the driving force to push or pull solvent and small micromolecules through the membrane. The gasket means could be positioned in a suitable seat around one or both of the two apertures. 
     The present invention provides great flexibility as to the selection of materials for the different components of the filtration module. Bonding compatibility is not necessary between any of the components. This means, for example, that a relatively soft and flexible material could used for filtration insert  1  or parts thereof, while a relatively rigid and strong material could be used for the sleeve housing  2 ″. Gasket  7  could even be integrated into support plate  6  and/or upper plate  12  of the filtration insert. Gasket  7  could thus have the form of a ridge at the periphery of membrane support plate  6  and/or upper plate  12  and be formed from the same material as the rest of the parts  6  and  12 . 
     The chemical properties of the different components could also be chosen with greater flexibility. A material with higher solvent-resistance could, for example, be chosen for the internal structure in contact with the liquid flow path. 
     The two sleeve parts  2 ,  2 ′ have been shown with closed end portions. This design gives a very strong and rigid sleeve which is able to stand high pressures. But it should be understood that such closed end portions are not absolutely necessary in all applications. 
     A method for manufacturing the device according to the invention comprises the following steps: gasket means are first arranged around at least one of the apertures in the concentration chamber or the filtrate chamber. One of the apertures is then covered with a membrane with its feed side against the concentration chamber and its permeate side against the filtrate chamber. The chambers are then assembled with the apertures arranged over each other and a pressure-resistant sleeve is finally fitted on the outside and around the combination of the concentration chamber, the gasket means, the membrane and the filtration chamber. The sleeve will create and maintain a pressure sufficiently high to seal the membrane fluid-tight against at least one of the chambers during the assembly, while at the same time effecting an increase in the structural support of the entire assembly. 
     The terms and expressions which have been employed in the foregoing specification are used therein as terms of description and not of limitation, and there is no intention in the use of such terms and expressions of excluding equivalents of the features shown and described or portions thereof, it being recognized that the scope of the invention is defined and limited only by the claims which follow.