Abstract:
There is provided apparatuses and methods for eluting microorganisms from filter media. The apparatus includes a housing for receiving filer media suspected of containing microorganisms and means for exposing the filter media to a pressurized buffer solution. By passing the buffer solution through the filter media tinder pressure, microorganisms trapped in or on the filter media are eluted therefrom.

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS 
       [0001]    This application is a Continuation of U.S. patent application Ser. No. 11/303,531, filed Dec. 16, 2005, which claims the benefit of and priority to U.S. Provisional Application Ser. No. 60/636,678, filed on Dec. 16, 2004, the entire contents of which is being incorporated herein by reference. 
     
    
     BACKGROUND 
       [0002]    1. Technical Field 
         [0003]    The present disclosure relates to apparatuses and methods for eluting or otherwise removing microorganisms from filter media. 
         [0004]    2. Discussion of Related Art 
         [0005]    The determination and enumeration of microbial concentration is an essential part of microbiological analyses in many industries, including water, food, cosmetics, and pharmaceuticals. Microorganisms, of interest to water microbiology, such as  Cryptosporidium  spp. and  Giardia  spp, are often present in low concentrations. This generates a requirement to sample large volumes of water to generate meaningful data. In the water industry, typically, 1,000 liters of finished water or 10-50 liters of surface water (e.g. lake water, river water etc.) are filtered to test for the presence of  Cryptosporidium  spp. oocysts and  Giardia  spp. cysts. Following filtration, these organisms must be recovered for further identification and quantification. Two major commercial filtration devices and methods are approved in the United States and United Kingdom for this application. 
         [0006]    U.S. Pat. No. 5,690,825 disclose the use of an expansible, compressed, open cell, solid foam to capture and recover microorganisms such as  Cryptosporidium  spp. and  Giardia  spp. by filtering large volumes of liquid samples (e.g. water) through the filter. The contents of the &#39;825 patent are herein incorporated by reference. Captured organisms are released from the foam filter by removing the compression and washing the target organisms from the foam matrix. A compressed foam filter device and automated washing/eluting device is currently marketed by IDEXX Laboratories, Inc., Westbrook, Me. under the Filta-Max® trademark. The Filta-Max elution procedure and wash station includes steps to decompress the foam filter modules first followed by repeated strokes of compressing and decompressing the Filta-Max filter in the presence of a buffer solution using a reciprocating plunger. The buffer solution used in the Filta-Max method includes an aqueous solution of PBST (phosphate buffer saline—0.01% Tween 20). The current process of eluting microorganisms from the Filta-Max® device and methods requires a washing procedure that is significantly more labor intensive than the presently disclosed invention. 
         [0007]    Pall Gelman Sciences Inc. manufactures and sells membrane filters (available from Pall Corporation) for capture and recovery of microorganisms from large volume water samples. The filter devices are currently marketed under the Envirochek™ trademark (hydrophilic polyethersulfone filter media) and the Envirochek™ HV trademark (hydrophilic polyester membrane). The process of eluting microorganisms from either of these devices and methods requires a washing procedure that is significantly more labor intensive than the presently disclosed invention. 
         [0008]    It is therefore, an object of the present invention to provide an apparatus and method of eluting microorganisms from filter media that is faster, easier to use and more efficient than currently marketed devices and methods. 
       SUMMARY 
       [0009]    The present invention discloses a novel and efficient apparatus and method of eluting microorganisms from filter media. Generally, the apparatus includes a pressure chamber in which the filter media suspected of containing microorganisms is placed or to which the filter media is fluidly connected. A buffer solution is disposed in the pressure chamber on one side of the filter media. Following pressurization of the chamber, an outlet is opened on the other side of the filter media, allowing the pressure and buffer solution to rapidly pass, in a flow direction reversed to the sampling direction, through the filter media resulting in efficient elution of microorganisms from the filter media. The process may be repeated, depending on the desired elution efficiency and microorganism recovery rates. 
         [0010]    According to an aspect of the present disclosure, an apparatus for eluting microorganisms from filter media is provided. The apparatus includes a housing configured and dimensioned to receive filter media, the housing having an inlet and an outlet; filter media disposed in the housing, the filter media having been exposed to a liquid suspected of containing microorganisms; means for transporting a liquid buffer solution into the housing via the outlet; and means for causing the liquid buffer solution to pass through the filter media under pressure and to exit the housing via the inlet. 
         [0011]    The means for causing the fluid buffer solution to pass through the filter media may include a pressurizing assembly selectively connectable to the outlet of the housing. The pressurizing assembly may include a pressure chamber configured for pressurizing a quantity of a liquid buffer solution therein prior to transportation of the liquid buffer solution to the housing. The pressure chamber may be in selective fluid communication with a source of pressurizing gas. The pressurizing assembly may include an air valve fluidly disposed between the source of pressurizing gas and the pressure chamber and a non-return valve fluidly disposed between the air valve and the pressure chamber. 
         [0012]    The apparatus may further include a reservoir configured to store a quantity of a liquid buffer solution therein, and a first conduit in fluid communication with the reservoir. The first conduit may include a free end configured to selectively fluidly connect with the pressure chamber. 
         [0013]    The apparatus may further include a liquid buffer solution contained within the reservoir. 
         [0014]    The apparatus may further include a buffer inlet valve fluidly disposed between the reservoir and the pressure chamber. 
         [0015]    The apparatus may still further include an elution valve fluidly connected to the pressure chamber and fluidly connectable to the outlet of the housing. 
         [0016]    The apparatus may further include a venting valve fluidly connected to the pressure chamber. 
         [0017]    It is contemplated that the pressure chamber may be pressurizable to a pressure of between about 0 psi (0 Bars) to at least about 72.5 psi (5.0 Bars). 
         [0018]    It is envisioned that the filter media may include a plurality of discs stacked upon one another. The stack of discs may alternate between relatively large outer diameter discs and relatively small outer diameter discs. The stack of discs may be compressed in a linear direction. 
         [0019]    According to a further aspect of the present disclosure a method for eluting microorganisms from filter media is provided. The method includes the steps of providing filter media suspected of containing microorganisms; and forcing a pressurized liquid through the filter media to at least partially elute microorganisms from the filter media, if present. 
         [0020]    It is envisioned that step of forcing a pressurized liquid through the filter media may include forcing the pressurized liquid through the filter media in a direction opposite to a direction of filtration. 
         [0021]    The method may further include the step of forcing a fixed quantity of pressurized liquid at a known initial pressure through the filter media. 
         [0022]    The method may still further include the step of providing an apparatus for eluting the filter media, as described above. 
         [0023]    The method may further include the step of introducing a fixed quantity of liquid buffer solution to the pressure chamber. 
         [0024]    The method may further include the step of pressurizing the pressure chamber a pressure of between about 0 psi (0 Bars) to at least about 72.5 psi (5.0 Bars). 
         [0025]    The method may further include the step of forcing the pressurized liquid buffer solution through the filter media. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0026]    The foregoing advantages and features of the presently disclosed apparatus and methods for liquid sample testing will become more readily apparent and may be understood by referring to the following detailed descriptions of illustrative embodiments, taken in conjunction with the accompanying drawings, in which: 
           [0027]      FIG. 1  is a schematic illustration of an apparatus for eluting microorganisms from a filter, in accordance with an embodiment of the present disclosure; 
           [0028]      FIG. 2  is a schematic illustration of a pressurizing assembly of the eluting apparatus of  FIG. 1 ; 
           [0029]      FIG. 3  is a schematic illustration of a pressurizing assembly according to an alternate embodiment of the present disclosure; 
           [0030]      FIG. 4  is a schematic side elevational view of an exemplary prior art filter module or device which may be eluted with the eluting apparatus of the present disclosure; 
           [0031]      FIG. 5A  is a side elevation view of a filter element, according to an embodiment of the present disclosure, for use in filter device; 
           [0032]      FIG. 5B  is a top plan view of a first disc member of the filter element of  FIG. 5A ; 
           [0033]      FIG. 5C  is a top plan view of a second disc of the filter element of  FIG. 5A ; and 
           [0034]      FIG. 6  is a graph illustrating the recovery efficiencies of  Cryptosporidium parvum  oocysts and  Giardia lamblia  cysts using different pressure elution procedures. 
       
    
    
     DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS 
       [0035]    The present disclosure will now be described more fully hereinafter with reference to the accompanying drawings, in which preferred embodiments of the disclosure are shown. Referring initially to  FIGS. 1 and 2 , an embodiment of an apparatus to elute microorganisms from a filter, filter module, filter device or the like, in accordance with the present disclosure, is generally designated as  100 . Although the presently disclosed elution apparatus  100  will be described and illustrated hereinafter in connection with specific embodiments and uses, such as, for example, the elution of  Cryptosporidium  and/or  Giardia  for filter modules/devices, it will be readily appreciated and understood by one skilled in the art that the presently disclosed elution apparatus  100  may be used in other applications equally as well or the elution apparatus  100  and methods disclosed herein may be adapted for use with a wide range of other filter modules/devices. 
         [0036]    With reference to  FIGS. 1 and 2 , elution apparatus  100  includes a reservoir or chamber  102 . Reservoir  102  is adapted to contain a quantity of a buffer solution “B” therein. As used herein, the buffer solution is any solution used to effect elution of the filter contained in the filter module housing. For example, the buffer solution may be a phosphate-buffered saline with 0.01% Tween 20. Alternatively, the buffer may comprise 0.1% Laureth 12, 10 mM Tris buffer. 1 mM di-sodium EDTA, and 0.015% antifoam A. It is further envisioned that the surfactant ingredients in the buffer solution may be selected from Tween 80, Igepal CA720, Niaproot, Laryl Sulphate, and Igepal CA630. A preferred buffer solution includes, for example, an aqueous solution of 0.02% (w/v) (or 0.45 mM) sodium pyrophosphate tetrabasic decahydrate, 0.03% (w/v) (or 0.84 mM) ethylenediaminetetraacetic acid trisodium salt and 0.01% (v/v) polyoxyethylenesorbitan monooleate (Tween 80), the complete disclosure of which is found in Inoue, M., Rai, S. K., Oda, T., Kimura, K., Nakanishi, M., Hotta, F., Uga, S., 2003, “A New Filter-eluting Solution that Facilitates Improved Recovery of  Cryptosporidium  Oocysts from Water,” J. Microbiol. Methods. 55, 679-686, the entire disclosure of which is incorporated herein by reference. An even further preferred buffer solution includes an aqueous solution of 0.01M Tris-HCL containing 0.02% (w/v) (or 0.45 mM) sodium pyrophosphate tetrabasic decahydrate, 0.03% (w/v) (or 0.84 mM) ethylenediaminetetraacetic acid trisodium salt and 0.01% (v/v) polyoxyethylenesorbitan monooleate (Tween 80). The reservoir  102  is envisioned to have at least 250 mL capacity; preferably, the reservoir will have a 10 L capacity for retaining buffer solution “B”. 
         [0037]    As seen in  FIGS. 1 and 2 , elution apparatus  100  further includes a pressurizing assembly  110  fluidly connected to reservoir  102  via a first conduit  104 . Pressurizing assembly  110  includes a pressure chamber  112  fluidly connected to reservoir  102 . In one preferred embodiment, the pressure chamber  112  has a 2.0 liter capacity and is capable of withstanding a pressure of at least 1 bar and preferably up to 12 bars. It is preferred that pressure chamber  112  includes a conical or frusto-conical lower portion  112   a  in order to facilitate the ejection of fluid therefrom. 
         [0038]    Pressurizing assembly  110  includes a first inlet or buffer inlet valve  114  fluidly connected between reservoir  102  and pressure chamber  112 . Buffer inlet valve  114  controls the inflow of buffer solution “B” into pressure chamber  112 . Pressurizing assembly  110  also includes a second inlet or compressed air inlet valve  116  fluidly connected between pressure chamber  112  and an air compressor, pump or the like  118 . Air inlet valve  116  controls the inflow of compressed air and/or other pressurizing gases into pressure chamber  112 . Preferably, a non-return valve  120  or the like may be fluidly connected between air inlet valve  116  and pressure chamber  112 . Non-return valve  120  prevents pressure loss from pressure chamber  112  back through air inlet valve  116 . 
         [0039]    Pressurizing assembly  110  may optionally include a third or venting valve  122  fluidly connected to pressure chamber  112 . The venting valve  122  allows air to exit pressure chamber  112  when pressure chamber  112  is being filled or charged with buffer solution “B”. 
         [0040]    Pressure assembly  110  further includes a fourth or elution valve  124  fluidly connected to pressure chamber  112 . Desirably, elution valve  124  is fluidly connected to lower portion  112   a  of pressure chamber  112 . Preferably, a fitting  126  is connected to a free end of elution valve  124 . The fitting  126  is configured and adapted to fluidly connect a filter housing or device  300  to elution valve  124 . 
         [0041]    Pressurizing assembly  110  further optionally includes a pressure gauge  130  operatively connected to pressure chamber  112  for measuring and displaying the pressure within pressure chamber  112 . 
         [0042]    Turning now to  FIG. 3 , an alternate embodiment of pressurizing assembly  110  is shown generally as  210 . Pressurizing assembly  210  is similar to pressurizing assembly  110  and will only be discussed in detail to the extent necessary to identify differences in construction and operation. 
         [0043]    As seen in  FIG. 3 , pressurizing assembly  210  includes a first inlet or buffer inlet valve  214  fluidly connected to pressure chamber  212  by a first union member  214   a . A first nipple  214   b  is operatively connected to buffer inlet valve  214  for connection with a first end of a tube or the like  215 . A second end of tube  215  may include a second nipple  214   c  for connection to reservoir  102  (see  FIG. 1 ). 
         [0044]    Pressurizing assembly  210  also includes a second inlet valve or compressed air inlet valve  216  fluidly connected between pressure chamber  212  and an air compressor, pump or the like  118  (see  FIG. 2 ). Preferably, a non-return valve  220  is fluidly connected between the compressed air inlet valve  216  and pressure chamber  212 . Non-return valve  220  prevents pressure loss from pressure chamber  212  back through the compressed air inlet valve  216 . Preferably, a first member  217   a  of a two-part quick-connect coupling  217  is connected to the compressed air inlet valve  216 . A second member  217   b  of the two-part quick-connect coupling  217  may be connected to a hose (not shown) extending from compressor  118  (see  FIG. 1 ) via a fitting  217   c.    
         [0045]    Pressurizing assembly  210  further includes a third or venting valve  222  fluidly connected to pressure chamber  212 . The venting valve  222  allows air to exit pressure chamber  212  when pressure chamber  212  is being filled or charged with buffer solution “B”. 
         [0046]    Pressure assembly  210  further includes a fourth or elution valve  224  fluidly connected to pressure chamber  212  by a first union member  224   a . Preferably, a fitting  226  is connected to a free end of elution valve  224  for fluidly connecting a filter housing or device  300  to elution valve  224 . 
         [0047]    Pressurizing assembly  210  further optionally includes a pressure gauge  230  operatively connected to pressure chamber  212  for measuring and displaying the pressure within pressure chamber  112 . 
         [0048]    Turning now to  FIG. 4 , an exemplary filter device or module, for use to capture and recover target microbes such as  Cyptosporidium  spp. and  Giardia  spp. from the samples and for use with the elution apparatus  100 , is shown generally as  300 . 
         [0049]    By way of example only, filter device  300  includes a filter housing  310  having a generally cylindrical body provided with a fixed outlet end  312   a  having an axially extending outlet tube  314 . A cap  316  is provided at an inlet end  312   b  and includes an axially extending inlet tube  318 . Cap  316  is secured to inlet end  312   b  of cylindrical body  310  by a threaded connection and scaled by an O-ring  324 . The direction of flow, during the filtration process, though filter device  300  is indicated by arrow “A”. Within housing  310  is a filter element  326 . Filter device  300  includes an upstream compression member, in the form of an apertured end plate  328 , and a downstream compression member, in the form of an apertured end plate  330 , connected by a rod member, in the form of a bolt  332 , passing through a central aperture of each end plate  328 ,  330 . Between end plates  328 ,  330  are compressed approximately 60 circular discs  326  of reticulate foam each having an uncompressed thickness of approximately 1 cm and an uncompressed porosity of 90 ppi (36 pores per cm). Circular discs  326  have been stacked end-over-end plane  328  and bolt  332  and have been pushed down by end plate  330  to compress the foam layers to an overall thickness of from 2 to 3 cm. Reference may be made to U.S. Pat. No. 5,690,825, the entire contents of which are incorporated herein by reference, for a detailed discussion of filter device  300 . Exemplary filter devices  300  are marketed and available from IDEXX Laboratories, Inc., Westbrook, Me., under the Filta-Max® trademark. 
         [0050]    Turning now to  FIGS. 5A-5C , in accordance with the present disclosure, a filter element for use in filter device  300 , is shown generally as  350 . Filter element  350  is multi-tiered and includes a plurality of first filter members  352  and second filter members  354  stacked in alternating arrangement with one another. Preferably, filter element  350  includes forty (40) first filter members  352  and thirty-nine (39) second filter members  354 . While a filter element  350  having forty first filter members  352  and thirty-nine second filter members  354 , arranged in alternating relationship, has been described, it is envisioned and within the scope of the present disclosure that any number of first and second filter members  352 ,  354  may be used and may be arranged in any order. 
         [0051]    As seen in  FIG. 5B , desirably, first filter members  352  is circular having an outer diameter “D 1 ” and defining a central opening  352   a  having an inner diameter “D 3 ”. Preferably, outer diameter “D 1 ” of first filter member  352  is approximately 55 mm (˜2.17 inches) and inner diameter “D 3 ” of first filter member  352  is approximately 18 mm (˜0.71 inches). 
         [0052]    As seen in  FIG. 5C , preferably, second filter members  354  is circular having an outer diameter “D 2 ” and defining a central opening  354   a  having an inner diameter “D 3 ”. Preferably, outer diameter “D 2 ” of second filter member  354  is approximately 40 mm (˜1.57 inches) and inner diameter “D 3 ” of second filter member  354  is equal to the inner diameter of central opening  352   a  of first filter member  352 . 
         [0053]    Preferably, first and second filter members  352 ,  354  are fabricated from expansible, open cell reticulated foam or the like. The foam is compressed so as to reduce its effective pore size to a level sufficient to filter large volumes of liquid samples and capture small particles or microbes such as  Cryptosporidium  spp. and/or  Giardia  spp. in the sample. 
         [0054]    Preferably, filter element  350  may be placed in filter device  300  in lieu of circular discs  326  described above. Use of filter element  350  helps to maintain a flow rate through filter device  300  within acceptable limits as well as reducing the incidence of target organisms bypassing the filter element. More preferably, 
         [0055]    With reference to  FIGS. 1-4 , in accordance with the present disclosure, a method of using elution apparatus  100  to elute a filter device  300 , is shown and described. In accordance with the method, buffer solution “B” is transmitted to or introduced into pressure chamber  112 . In particular, with venting valve  122  open in order to vent air or gases from within pressure chamber  112  and air inlet valve  116  and elution valve  124  in a closed condition, buffer inlet valve  114  is manipulated to an open condition to open the passage between reservoir  102  of buffer solution “B” and pressure chamber  112 . Preferably, reservoir  102  is located above pressure chamber  112  so that buffer solution “B” is transmitted via a gravity feed, however, any method of introducing buffer solution “B” into pressure chamber  112  is contemplated, for example, by pouring into a sealable opening, using positive pressure to deliver buffer solution “B” to pressure chamber  112 , etc. Preferably, an effective amount or quantity of buffer solution “B” is introduced into pressure chamber  112 . For example, approximately 240 ml of buffer solution “B” is transferred from the reservoir  102  into the pressure chamber  112  for each elution process. 
         [0056]    With buffer solution “B” introduced into pressure chamber  112 , buffer inlet valve  114  is once again manipulated in order to close the passage between reservoir  102  of buffer solution “B” and pressure chamber  112 . Additionally, venting valve  122  is also manipulated to a closed position in order to prevent the escape of gas or buffer solution “B” from pressure chamber  112 . 
         [0057]    Once buffer solution “B” is contained in pressure chamber  112  and venting valve  122  is closed, air inlet valve  116  is manipulated to the open condition. By opening air inlet valve  116 , pressure chamber  112  is pressurized with air or the like from air compressor  118 . Air inlet valve  116  is maintained open until the pressure within pressure chamber  112  is about 1.0 bar (approximately 14.5 psi) to about 5.0 bars (approximately 72.5 psi), preferably about 4.0 bars (approximately 58 psi) at which time air inlet valve  116  is closed. The pressure within pressure chamber  112  is measured and visualized by pressure gauge  130 . 
         [0058]    At this point in the process, or, if desired, prior to this point, a filter device  300  is fluidly connected to elution valve  124 . In particular, the outlet tube  314  of filter device  300  is connected to elution valve  124 . Filter device  300  is preferably a filter device which has become at least partially saturated with microorganisms (e.g.,  Cryptosporidium  and  Giardia ) after performing numerous hours of filtering and/or after having filtered numerous gallons of fluid. In order to capture and/or contain the expurgated fluid or eluate (i.e., buffer solution “B” and the microorganisms from filter device  300 ) a collection container or the like is placed beneath inlet tube  318  of filter device  300 , or alternately, a fluid conduit (not shown) may be fluidly connected to inlet tube  318  of filter device  300 . 
         [0059]    With the pressure within pressure chamber  130  at or about the desired or required pressure, elution valve  124  is manipulated to the open condition thereby forcing pressurized buffer solution “B” through filter device  300 , in a direction opposite to arrow “A” of  FIG. 4 . In so doing, microorganisms captured and/or contained in filter device  300  are driven out of and/or forced out of filter element  326  of filter device  300 . 
         [0060]    Once the eluate is collected, elution valve  124  is manipulated to the closed condition. Filter device  300  may then be removed from elution valve  124  and discarded or reconditioned for further filtering operations. If required and/or desired, venting valve  122  may be re-opened to further vent pressure chamber  112 . The eluate may then be further processed and/or analyzed as known by those having ordinary skill in the art. It is envisioned and within the scope of the present disclosure that the filter device  300  may be maintained attached to or re-attached to elution valve  124  and additional pressurized buffer solution “B” forced therethrough in order to further expurgate and/or elute additional microorganisms. 
         [0061]    This invention and its benefit can be further illustrated by the following examples: 
       Example 1 
     Recovery Efficiencies of  Cryptosporidium  spp. oocysts and  Giardia  spp. Cysts from Drinking Water Samples 
       [0062]    Initially, 1,000 liters and 50 liters of drinking water samples from Newmarket, UK and Veolia Water Company, UK were spiked with 100  Cryptosporidium parvum  oocysts and 100  Giardia lamblia  cysts (Waterborne™, Inc. New Orleans, La., USA). The packed pellet sizes were &lt;0.5 mL for the Newmarket sample and 0.5 mL for the Veolia sample. Water samples containing the spiked  Cryptosporidium  spp. oocysts and  Giardia  spp. cysts were passed through each of the filter modules of the Filta-Max, and a 79-Disc filter according to the structure briefly described above in  FIG. 5 . The 79-Disc filter module consists of 79 open cell reticulated foam pad rings with two different sizes: 40 of the large foam pads have a 55 mm outer diameter and an 18 mm inner diameter and 39 of the small foam pads have a 40 mm outer diameter and an 18 mm inner diameter. All the roam rings of the 79-Disc filter are 10 mm thick. The two sizes of foam pads (i.e., the 55 mm and the 40 mm pads) are sandwiched in an alternating pattern into a stack. The stack is then compressed from about 790 mm to about 30 mm and is tightened by a retaining bolt. This construction resulted in a filter module with two filtration layers: the outer layer of the filter module (i.e., the region radially outward of the outer diameter of the 40 mm foam pads) is compressed  13  fold and acts as a pre-filter and the inner layer of the filter module (i.e., the region radially inward of the outer diameter of the 40 mm foam pads) is compressed  27  told and acts as a size exclusion filter. 
         [0063]    The Filta-Max method is the standard method in England and is approved by the Drinking Water Inspectorate (DWI). DWI is responsible for assessing the quality of drinking water in England and Wales, taking enforcement action if standards are not being met and appropriate action when water is unfit for human consumption. The filtered Filta-Max modules were processed and the captured organisms were eluted using the standard Filta-Max elution procedure as described in the DWI procedure. In this experiment, both minimally expanded (5 mm) and non-expanded 79-Disc filter were tested using one embodiment of this invention. The filters were eluted in a flow direction reversed to the sampling step only once with 240 mL pressurized buffer solution (0.45 mM sodium pyrophosphate, 0.84 mM tri-sodium EDTA, 0.01% Tween 80) at 5 bars pressure (i.e. 72.5 psi). The organisms in the eluted filtrates were purified using a standard immunomagnetic separation method (Dynal® Invitrogen Corporation, Carlsbad, Calif., USA), stained with a fluorescent antibody stain, and enumerated using a fluorescent microscope. As shown in the table below, these data indicated that, using the device and method of this invention, the recovery efficiencies were equivalent or better than the official method, Filta-Max. 
         [0000]    
       
         
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                   
               
               
                 Filer &amp; Elution 
                 Sample 
                 
                   Cryptosporidium 
                 
                 
                   Giardia 
                 
               
             
          
           
               
                 Methods 
                 Sources 
                 Recovery 
                 Mean 
                 Recovery 
                 Mean 
               
               
                   
               
               
                 Filta-Max/DWI 
                 Newmarket, 
                 35.4% 
                 37.5% 
                 17.2% 
                 21.5% 
               
               
                   
                 UK 
               
               
                   
                 Veolia 
                 39.5% 
                   
                 25.8% 
               
               
                   
                 Water, UK 
               
               
                 79 Disc filter 
                 Newmarket, 
                 24.6% 
                 33.6% 
                 24.2% 
                 23.3% 
               
               
                 (0 mm)/PE 
                 UK 
               
               
                   
                 Veolia 
                 42.6% 
                   
                 22.4% 
               
               
                   
                 Water, UK 
               
               
                 79 Disc filter 
                 Newmarket, 
                 33.6% 
                 43.7% 
                 20.5% 
                 27.5% 
               
               
                 (5 mm)/PE 
                 UK 
               
               
                   
                 Veolia 
                 53.7% 
                   
                 34.4% 
               
               
                   
                 Water, UK 
               
               
                   
               
             
          
         
       
     
       Example 2 
     Recovery Efficiencies of  Cryptospodium  spp. Oocysts and  Giardia  spp. Cysts from Raw Water Samples 
       [0064]    Initially, 50 liters of surface water samples from Iowa, North Dakota, California, and Pennsylvania were spiked with 100  Cryptosporidium parvum  oocysts and 100  Giardia lamblia  cysts (Waterborne™, Inc. New Orleans, La., USA). The packed pellet size for all these water samples was 0.5 mL. Water samples containing the spiked  Cryptosporidium  oocysts and  Giardia  cysts were collected using the filter modules of Gelman HV, Filta-Max. ID filter and 79-Disc filter. The 79-Disc tilter module consists of 79 open cell reticulated foam pad rings with two different sizes: 40 of the large foam pads have a 55 mm outer diameter and an 18 mm inner diameter and 39 of the small foam pads have a 40 mm outer diameter and an 18 mm inner diameter. All the foam rings are 10 mm thick. The two sizes of foam pads (i.e., the 55 mm and the 40 mm pads) are sandwiched in an alternating pattern into a stack. The stack of foam pads is then compressed from about 790 mm to about 30 mm and is tightened by a retaining bolt. This construction resulted in a filter module with two filtration layers: the outer layer of the filter module (i.e. the region radially outward of the outer diameter of the 40 mm foam pads) is compressed  13  fold and acts as a pre-filter and the inner layer of the filter module (i.e., the region radially inward of the outer diameter of the 40 mm foam pads) is compressed  27  fold and acts as a size exclusion filter. The ID-filter (increased-depth) module is constructed from 67 rings of open cell reticulated polyester foam. 51 of the rings are 84 mm in diameter and 16 of the rings are 55 mm in diameter. All of the rings are 10 mm thick and have an 18 mm central hole. The rings are layered in an alternating pattern with the larger rings grouped in stacks of three interspaced by a smaller ring. The stack is compressed from about 670 mm to about 30 mm. This construction results in a filter module with two filtration layers. The outer later of the filter module (i.e. the region radially outward of the outer diameter of the 40 mm foam pads) is compressed  17  fold and acts as a pre-filter. The central core of the filter module (i.e., the region radially inward of the outer diameter of the 40 mm foam pads) is compressed  22  fold and acts as an efficient size exclusion filter. 
         [0065]    Filta-Max and Gelman HV methods are the standard method accepted by the United Stated Environmental Protection Agency (USEPA) and are included as the USEPA Method 1623 for concentrating and recovering the  Cryptosporidium  spp. oocysts and  Giardia  spp. cysts in surface water samples. The Filta-Max module and Gelman HV were processed and the captured organisms in these filters were eluted using the standard Filta-Max and Gelman HV procedures as described in the USEPA Method 1623. Both ID-filters and 79-Disc filters were processed to elute the captured organisms using one embodiment of this invention, respectively. In this experiment, both minimally expanded (5 mm) and non-expanded filter modules of the ID-filters and 79-Disc filters were evaluated. The filters were eluted in a flow direction reversed to the sampling step only once with 240 mL pressurized buffer solution at 5 bars pressure (i.e. 72.5 psi). The organisms in the eluted filtrates were purified using a standard immuno-magnetic separation method (Dynal® Invitrogen Corporation, Carlsbad, Calif., USA), stained with a fluorescent antibody stain, and enumerated using a fluorescent microscope. As shown in the table below, these data indicated that, using the device and method of this invention, the recovery efficiencies were equivalent or better than those of the official methods, Filta-Max and Gelman HV. 
         [0000]    
       
         
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                   
               
               
                 Filter/Elution 
                 Sample 
                 
                   Cryptosporidium 
                 
                 
                   Giardia 
                 
               
             
          
           
               
                 Methods 
                 Sources 
                 Recovery 
                 Mean 
                 Recovery 
                 Mean 
               
               
                   
               
               
                 Gelman HV 
                 Iowa 
                 33.4 
                 37.0% 
                 46.2 
                 49.4% 
               
               
                 Filter 
                 North Dakota 
                 31.1 
                   
                 43.7 
               
               
                   
                 California 
                 55.4 
                   
                 52.2 
               
               
                   
                 Pennsylvania 
                 27.9 
                   
                 55.6 
               
               
                 Filta-Max 
                 Iowa 
                 43.5 
                 37.1% 
                 43.1 
                 37.8% 
               
               
                   
                 North Dakota 
                 30.5 
                   
                 39.4 
               
               
                   
                 California 
                 35.7 
                   
                 39.6 
               
               
                   
                 Pennsylvania 
                 38.5 
                   
                 29.2 
               
               
                 ID Filter 
                 Iowa 
                 29.2 
                 33.8% 
                 38.5 
                 43.7% 
               
               
                 (0 mm) 
                 North Dakota 
                 23.0 
                   
                 23.2 
               
               
                   
                 California 
                 36.2 
                   
                 51.1 
               
               
                   
                 Pennsylvania 
                 42.6 
                   
                 62.1 
               
               
                 ID Filter 
                 Iowa 
                 23.8 
                 37.9% 
                 39.2 
                 42.9% 
               
               
                 (5 mm) 
                 North Dakota 
                 46.6 
                   
                 39.4 
               
               
                   
                 California 
                 38.6 
                   
                 37.3 
               
               
                   
                 Pennsylvania 
                 42.6 
                   
                 55.6 
               
               
                 79 Disc 
                 Iowa 
                 44.7 
                 52.0% 
                 47.7 
                 48.2% 
               
               
                 (0 mm) 
                 North Dakota 
                 69.7 
                   
                 57.0 
               
               
                   
                 California 
                 52.1 
                   
                 44.2 
               
               
                   
                 Pennsylvania 
                 41.6 
                   
                 43.9 
               
               
                 79 Disc 
                 Iowa 
                 45.3 
                 57.0% 
                 45.4 
                 51.5% 
               
               
                 (5 mm) 
                 North Dakota 
                 72.8 
                   
                 61.3 
               
               
                   
                 California 
                 65.6 
                   
                 51.7 
               
               
                   
                 Pennsylvania 
                 44.2 
                   
                 47.5 
               
               
                   
               
             
          
         
       
     
       Example 3 
     Recovery Efficiencies of  Cryptosporidium  spp. oocysts and  Giardia  spp. Cysts from 50 L Surface Water Samples Between Two Methods 
       [0066]    Initially, seven (7) surface water samples including California River #1 US; Massachusetts Lake, US; Alabama River, US; an unknown River, US; Georgia Reservoir, US and River Cambridge, UK were used. With the exception of River Cambridge sample which had a packed pellet size of 0.4 mL, the pellet sizes for all other samples were 0.5 mL. 50 liters of the indicated water samples were spiked with 100  Cryptosporidium  oocyst and 100  Giardia  cysts (Easyseed™, BTF Pty Ltd., North Ryde Australia). Water samples containing the spiked  Cryptosporidium  oocysts and  Giardia  cysts passed through the filter modules of Filta-Max and a 79-Disc filter with the structure described in  FIG. 5 . The 79-Disc filter module consists of 79 open cell reticulated foam pad rings with two different sizes: 40 of the large foam pads have a 55 mm outer diameter and an 18 mm inner diameter and 39 of the small foam pads have a 40 mm outer diameter and an 18 mm inner diameter. All the foam rings are 10 mm thick. The two sizes of foam pads are sandwiched in an alternating pattern into a stack. The stack is then compressed from about 790 mm to about 30 mm and is tightened by a retaining bolt. This construction resulted in a filter module with two filtration layers: the outer layer of the filter module (i.e., the region radially outward of the outer diameter of the 40 mm foam pads) is compressed  13  fold and acts as a pre-filter and the inner layer of the filter module (i.e., the region radially inward of the outer diameter of the 40 mm foam pads) is compressed  27  fold and acts as a size exclusion filter. 
         [0067]    The filtered Filta-Max modules were processed and the captured organisms were eluted according to the standard Filta-Max elution procedure as described in the USEPA Method 1623 for the concentration and recovery of  Cryptosporidium  and  Giardia  in surface water samples. The 79-Disc filters were processed to elute the captured organisms using one embodiment of this invention. This elution embodiment used a 4-step elution sequence: (1) air purge with 4 bars (i.e. 58 psi) of compressed air, (2) 240 mL pressurized buffer elution at 4 bars pressure, (3) air purge with 4 bars (i.e. 58 psi) of compressed air, and (4) 150 mL pressurized buffer elution at 4 bars pressure. The buffer solution used for this elution procedure contained Sodium pyrophosphate tetra-basic decahydrate (0.2 gram/Liter), EDTA tri-sodium salt (0.3 gram/Liter), Tris-HCl (0.01M), and Tween-80 (0.1 mL/Liter). The organisms in the eluted filtrates were purified using a standard immuno-magnetic separation method (Dynal® Invitrogen Corporation, Carlsbad, Calif., USA), stained with a fluorescent antibody stain, and enumerated using a fluorescent microscope. As seen in the table below, these data indicated that, using the device and method of this invention, the mean recovery efficiencies for Cryptosporidium was 31.5% and for Giardia was 41.5%, which were about 115% for  Cryptosporidium  and about 128% for  Giardia  relative to those of the official methods, Filta-Max. 
         [0000]    
       
         
               
               
               
               
             
               
               
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                   
                   
               
               
                   
                 pack 
                 
                   Cryptosporidium 
                 
                 
                   Giardia 
                 
               
             
          
           
               
                   
                 pellet 
                 Filta- 
                   
                 Filta- 
                   
               
               
                 Surface Water Samples 
                 size 
                 Max 
                 79-Disc 
                 Max 
                 79-Disc 
               
               
                   
               
             
          
           
               
                 California River #1, US 
                 0.5 mL 
                 31.6% 
                 37.9% 
                 42.6% 
                 44.4% 
               
               
                 Massachusetts Lake, US 
                 0.5 mL 
                 40.0% 
                 27.1% 
                 28.5% 
                 60.0% 
               
               
                 California River #2, US 
                 0.5 mL 
                 41.2% 
                 69.4% 
                 39.2% 
                 66.9% 
               
               
                 Alabama River, US 
                 0.5 mL 
                 22.4% 
                 20.6% 
                 27.7% 
                 25.4% 
               
               
                 Unknown River, US 
                 0.5 mL 
                 11.2% 
                 8.8% 
                 5.4% 
                 7.7% 
               
               
                 Georgia Reservoir, US 
                 0.5 mL 
                 16.5% 
                 22.4% 
                 37.7% 
                 30.0% 
               
               
                 Cambridge River, UK 
                 0.4 mL 
                 28.8% 
                 34.4% 
                 46.2% 
                 56.2% 
               
               
                 Overall Mean Recovery 
                   
                 27.4% 
                 31.5% 
                 32.5% 
                 41.5% 
               
               
                   
               
             
          
         
       
     
       Example 4 
       [0068]    Recovery Efficiencies of  Cryptosporidium  spp. Oocysts and  Giardia  spp. Cysts Using Different Pressure Elution Procedures 
         [0069]    Initially, 10 liters of RO water samples were spiked with 100  Cryptosporidium parvum  oocysts and 100  Giardia lamblia  cysts (Waterborne™, Inc. New Orleans, La., USA). Water samples containing the spiked  Cryptosporidium  oocysts and Giardia cysts passed through the filter modules of a 79-Disc filter with the structure described in  FIG. 5 . The 79-Disc filter module consists of 79 open cell reticulated foam pad rings with two different sizes: 40 of the large foam pads have a 55 mm outer diameter and an 18 mm inner diameter and 39 of the small foam pads have a 40 mm outer diameter and an 18 mm inner diameter. All the foam rings are 10 mm thick. The two sizes of foam pads are sandwiched in an alternating pattern into a stack. The stack is then compressed from about 790 mm to about 30 mm and is tightened by a retaining bolt. This construction resulted in a filter module with two filtration layers: the outer layer of the filter module (i.e., the region radially outward of the outer diameter of the 40 mm foam pads) is compressed  13  fold and acts as a pre-filter and the inner layer of the filter module (i.e., the region radially inward of the outer diameter of the 40 mm foam pads) is compressed  27  fold and acts as a size exclusion filter. The 79-Disc filters were processed to elute the captured organisms using different embodiments of this invention. These included: (1) 2 sequential pressurized buffer elution (1×240 mL+1×150 mL); (2) one time compressed air purge followed by 2 sequential pressurized buffer elution (i.e. AP+1×240 mL+1×50 mL); (3) one time compressed air purge, one time 240 mL pressurized buffer elution, one time air purge, followed by one time 150 mL pressurized buffer elution (i.e. AP+1×240 ml, +AP+1×150 mL); (4) one time compressed air purge followed by 3 times 130 mL pressurized buffer elution; (5) one time compressed air purge followed by 4 times 100 mL pressurized buffer elution; (6) one time compressed air purge followed by 5 times 80 mL pressurized buffer elution; and (7) one time compressed air purge followed by 5 times pressurized buffer elution with the buffer pre-warmed to 37° C. All pressure elution steps were carried out in a flow direction reversed to the sampling step at 4 bars pressure. The buffer solution used for this elution procedure contained Sodium pyrophosphate tetra-basic decahydrate (0.2 gram/Liter), EDTA tri-sodium salt (0.3 gram/Liter), Tris-HCl (0.01M), and Tween-80 (00.1 mL/Liter). The organisms in the eluted filtrates were purified using a standard immunomagnetic separation method (Dynal® Invitrogen Corporation, Carlsbad, Calif., USA), stained with a fluorescent antibody stain, and enumerated using a fluorescent microscope. As seen in  FIG. 6 , these data indicated that, using the device of this invention, the recovery efficiencies were essentially similar to one another among different embodiments of this invention. 
       Example 5 
     Procedural Time Difference Between Filta-Max and the Methods of the Present Invention 
       [0070]    In the present example, 5 water samples including 1 reagent water sample (representing clean water sample) and 4 raw water samples with different turbidities were used in this experiment. Water samples passed through the filter modules of a 79-Disc filter with the structure described in  FIG. 5 . The 79-Disc filter module consists of 79 open cell reticulated foam pad rings with two different sizes: 40 of the large foam pads have a 55 mm outer diameter and an 18 mm inner diameter and 39 of the small foam pads have a 40 mm outer diameter and an 18 mm inner diameter. All the foam rings are 10 mm thick. The two sizes of foam pads are sandwiched in an alternating pattern into a stack. The stack is then compressed from about 790 mm to about 30 mm and is tightened by a retaining bolt. This construction resulted in a filter module with two filtration layers: the outer layer of the filter module (i.e., the region radially outward of the outer diameter of the 40 mm foam pads) is compressed  13  fold and acts as a pre-tilter and the inner layer of the filter module (i.e., the region radially inward of the outer diameter of the 40 mm foam pads) is compressed  27  fold and acts as a size exclusion filter. 
         [0071]    The Filta-Max modules were processed according to the standard Filta-Max procedures as described in the USEPA Method 1623. The 79-Disc filters were processed using the device and method of this invention (i.e. Pressure Elution). Filta-Max&#39;s sample processing time ranged from II minutes and 25 seconds to twenty six minutes and forty five seconds depending on the nature of water samples. When the device and method of this invention (i.e. pressure elution) was used to perform the sample elution, the time required to process the elution step only took 2 minutes and five seconds irregardless of the nature of the water samples. As seen in the table below, there is therefore significant benefit in the reduction of sample processing time requirement and labor saving using the device and method of this invention. 
         [0000]    
       
         
               
               
               
               
             
               
               
               
               
               
             
           
               
                   
                   
               
               
                   
                 Procedural 
                 Added 
                 Total 
               
               
                   
                 Time 
                 Time 
                 Time 
               
               
                   
                   
               
             
             
               
                   
               
             
          
           
               
                 Filta-Max 
                 Reagent Water 
                 11:25 
                 00:00 
                 11:25 
               
               
                 Elution 
                 Samples 
               
               
                   
                 Average of 4 Raw 
                 11:25 
                 15:20 
                 26:45 
               
               
                   
                 Water Samples 
               
               
                 Pressure 
                 Reagent Water 
                  2:05 
                 00:00 
                  2:05 
               
               
                 Elution 
                 Samples 
               
               
                   
                 Average of 4 Raw 
                  2:05 
                 00:00 
                  2:05 
               
               
                   
                 Water Samples 
               
               
                   
               
             
          
         
       
     
         [0072]    While the invention has been particularly shown and described with reference to the attached sheets of schematics and drawings, it will be understood by those skilled in the art that various modifications, including without limitation of having a fully automatic device and method to process the sample elution, in form and detail may be made therein without departing from the scope and spirit of the invention. Accordingly, modifications such as those suggested above, but not limited thereto, are to be considered within the scope of the invention.