MXPA00004137A - Device and methods for determination of analyte in a solution - Google Patents

Abstract

Method and assay devices for the detection of the presence or amount of biological material, analyte(s), or microorganism(s) in a sample. The method includes the steps of liquefying the sample (if necessary) and distributing the liquefied sample over the surface of the assay device. The device may comprise an incubation plate, a dip stick device, or other devices. The devices have at least one reagent provided within the devices. Some devices have a generally flat horizontal surface which is divided into a plurality of recessed wells. Others have one or more surfaces with reagent island(s) immobilized thereon. Each well or reagent island is adapted to hold an aliquot of liquid. The wells or reagent islands are sized and shaped, and formed of a suitable material, to hold the aliquot within the well or reagent island by surface tension. Any excess liquid from the liquefied sample is drained from the surface of the device. The method then involves incubating the assay device until the presence or amount of the biological material, analyte, or microorganism is determined.

Description

DEVICE AND METHODS FOR THE DETERMINATION OF ANALYTS IN A SOLUTION RELATED REQUESTS This application is a continuation in part of the provisional application of E.U.A. 60 / 063,635, entitled METHOD FOR QUANTITATION OF BIOLOGICAL MATERIAL IN A SAMPLE USING A REAGENT-CONTAINING INCUBATION PLATE, (Method for the quantification of biological material in a sample using an incubation plate containing reagent), filed on October 27, 1997 BACKGROUND OF THE INVENTION This invention relates to the field of test technology, and in particular embodiments, to devices and methods for the quantification of analytes, for example, biological material, in a sample. Many industries need to detect and quantify the concentration and level of biological material or other analyte in a sample. For example, determining the concentration of bacteria in food and water is an essential part of quality testing of food and water. EPA regulations require that no coliform such as Escherichia coli be present in drinking water. The "presence / absence" format of a test medium, such as a Colilert® chemical mixture (IDEXX Laboratories, ME) that is used as a test medium for Escherichia coli and all coliform bacteria, is very useful to do this determination. The Colilert® chemical blend is based on the substrate technology described in Edberg, "Method and Medium for Use in Detecting Target Microbes in Situ n A Specimen Sample of A Possibly Contaminated Material," US patents. us. 4,925,789 and 5,492,933. However, there are areas in which quantification, not just detection, of bacteria concentration is important. Examples of such areas include waste water, inlet water in water purification systems, surface water and food testing. For example, many restaurant chains will only accept ground meat or poultry meat that contains less than a certain concentration of contamination by bacteria. Therefore, microbiological tests necessary to determine the bacterial concentration of these food products must be carried out in the food processing plants before they can be supplied to the customers. The classical methods of quantification of biological material are the standard plate counting method or the multiple tube fermentation (MTF) method. A sample quantity that is being tested for microbial contamination is first distributed in a Petri dish. Then 15 ml of the appropriate medium is emptied into the sample. The Petri dish is then subjected to swirling action to mix the sample in the medium and the petri dish is allowed to solidify at room temperature for approximately 20 minutes. The medium is then incubated at a specific temperature for a specific time and the resulting colonies are counted. The multi-tube fermentation method is described in Recles et al., "Most Probable Number Techniques" published in "Compendium of Methods for the Microbiological Examination of Foods", 3a. ed. 1992, pages 105-199, and Greenberg et al., "Standard Methods for the Examination of Water and Wastewater" 8a. ed. 1992). In this method, a sample volume is distributed in several tubes that represent this dilution scale. The tubes are then incubated at the appropriate temperature so the bacteria in each tube are allowed to grow. After incubation at a specific temperature for a specific time, the number of positive tubes is counted. The most probable number can be determined from the formula described in Recles et al., Supra. The water test in most cases is done by membrane filtration, where a certain volume of water is passed through the membrane and the membrane is incubated in a medium for a certain period. After the appropriate incubation, the colonies are counted. In many industries there is also a need to detect qualitatively and / or quantitatively the presence of an analyte in a liquid solution. For example, the detection of inorganic ions may be important in the manufacturing process using a test solution. Until now, the methods and devices on which it is generally based for the measurement of an analyte in solution have required the removal of an entire aliquot of the test solution or exposure of a dipstick to a test solution. Although these methods and devices can detect an analyte in solution, they suffer from many drawbacks. The methods related to the dipstick are not quantitative. For example, in the general modalities of immersion rods there is no ability to determine or quantify the presence of an analyte in a unit volume of that solution. Instead, the dipstick is simply brought into contact with the test solution. Other methods require a user to manually remove an aliquot of a test solution and transfer it to a separate device for the detection or quantification of the analyte. Therefore, despite the ability of these methods and devices to detect an analyte in solution, currently used methods and devices have been shown to have limited accuracy; and / or be expensive, complicated, time consuming; and / or have a limited scale of uses due to the particular testing technology. Thus, there is a need for a simple, accurate and low-cost method for the determination of an analyte in solution without the drawback known in the prior art. In particular, there is a need for devices and methods capable of providing a quantitative test of an analyte in a particular volume of a solution.

BRIEF DESCRIPTION OF THE INVENTION The present invention provides devices and methods for detecting and enumerating the presence or absence of biological materials, analytes and microorganisms in sample solutions. In one aspect, the invention provides a sterile incubation plate for determining the presence or amount of a biological material in a test sample. The plate is usually described by a flat horizontal surface containing depressed cavities. Each cavity is adapted to contain an aliquot of liquid, and has a size and sample, and is made of a material, suitable for containing the liquid aliquot within the cavity by the surface tension forces. At least one cavity contains at least one reagent for the detection of the biological material. No positive response is generated in the absence of the objective biological material. In a preferred embodiment, the reagent is deposited in the cavities by corona discharge treatment and drying. In another embodiment, the plate may also contain a lid. The cavity or cavities may contain a plurality of reagents, and different cavities may contain different reagents or different combinations of reagents, whereby numerous tests can be conducted on a single plate. The plate is preferably constructed of plastic, however, it can be constructed of other hydrophobic materials that are suitable for conducting the test. In a preferred embodiment, the plates of the present invention will be rectangular in shape, however, they may also be circular in shape, or in any way. In a preferred embodiment, the cavities are approximately 0.38 cm in diameter. In another preferred embodiment, the cavities in the plate contain a total of about 1 mm of solution. In another preferred embodiment, each cavity of the plate contains between 0.1 and 100 μl of liquid. The cavity or cavities of the plate can be bevelled to help remove excess fluid. The plate may further comprise a handle portion so that the operator can manipulate the plate and conduct the test without risk of making contact with the test sample. In another aspect, the invention provides a sterile incubation plate similar to the plate described above, and can be adapted with each of its modalities, with the additional feature that it comprises a cap that contains at least one projection (s) that fits (n) in the cavity (s). In this aspect, the reagent or combination of reagents is contained in the end of the projection (s) in such a way that the reagent (s) will dissolve in the test sample when closing or attaching the cap to the plate. In one embodiment, the projections may have a cavity. The reagent (s) or combination of reagents will dry on the end of each projection or on the surface of the cavity. The surface of the cavity will also be treated by corona discharge before depositing the reagent. In this aspect, also a reagent (s) or combination (s) of reagents can be dried on, or the projection (s) treated by corona discharge, and a combination of reagents can be used between multiple projections so that they can be used. conduct multiple tests on a single plate. The lower plate portion can be adapted to all the modalities with respect to the previously described plate. In another aspect, the invention describes a device for determining the presence or amount of analyte (s) or microorganism (s) in a test solution. The device has a substantially hydrophobic support structure with at least one islet of reagent immobilized on the support structure which is capable of absorbing a predetermined volume of test solution. The device also comprises means for determining the presence or amount of the analyte or microorganism, the media being placed on or within the reagent islet (s). This aspect of the invention can take the form of a plastic or polymer immersion rod with one or more reagent islets immobilized thereon. The islet of reagent can be made from any absorbent material, for example, cellulose. The support structure may have multiple islets of reagent immobilized thereon. The means for determining the presence or amount of microorganisms or analyte (s) may comprise a powder that is incorporated on or into the material for which the islet of reagent is prepared. The media can be a reagent or a combination of reagents that lead to the production of a detectable signal when the analyte (s) or target microorganism (s) is present. The device may contain multiple islets of reagent which may contain different reagents or combinations of reagents whereby numerous tests can be conducted in a single device. In a preferred embodiment, the reagent islets are composed of any absorbent material, for example, cellulose. The support structure can be made of plastic, polymer or any suitable material on which the islet of reagent can be mounted and which does not interfere with the test. In another embodiment, the device also comprises a container for containing the support structure before, during and after conducting the test. The container will typically comprise a test tube and may further comprise a cover for the type of test or further protect the device. In another aspect, the invention provides a test device for determining the presence or amount of an analyte (s) or microorganism (s) in a sample comprising a substantially hydrophobic solid support structure with at least one islet of immobilized discrete reagent. on it which is capable of absorbing and containing a predetermined volume of test solution. Placed on or within the reagent islet is a means to determine the presence or absence of an analyte or target microorganism. This aspect of the device further comprises a container that is open at both ends. The container can typically take the form of a laboratory pipette. This aspect of the invention may comprise any of the various embodiments described with respect to the device described above. The reagent islets may also be arranged in zones such that each zone provides a separate test. The islets of reagent can be adapted to retain aliquots of liquids by surface tension or by absorption. The device typically may comprise an elongated strip. The reagent comprised in the reagent islets may be a growth medium or an indicator growth medium. The reagent can also be an enzyme or an enzyme substrate. In another aspect, the invention provides a test device for determining the presence or amount of an analyte (s) or microorganism (s) in a sample comprising a solid support structure with at least one islet of reagent immobilized on the same Each islet of reagent is adapted to contain an aliquot of liquid and is of a size and shape and is made of suitable material to contain the aliquot within the islet of reagent. At least one of the reagent islets contains at least one reagent for the detection of the analyte (s) and / or microorganism (s) of interest. The device does not provide a positive response for the analyte (s) or microorganism (s) when the analyte (s) or microorganism (s) is not present in the test sample. In a preferred embodiment, the reagent islets of the device can absorb up to a preselected total volume. In one embodiment, the device may comprise a combination of reagents located in islets of separate reagents. In another embodiment, the islets of reagents are arranged in zones, where each zone provides a separate test so that multiple tests can be conducted in a single device. The reagent islets may be adapted to retain liquid sample aliquots by surface tension or by absorption. The reagent islets may be adapted to absorb liquid volumes of liquid. O well, reagent islets can exist in subsets of islets, where each subset consists of at least one islet, and each subset absorbs different volumes of the other subsets. The device can be constructed in such a way that the islet of reagent is immobilized on more than one surface. In different embodiments, the device can be a plate or an elongated strip. In a preferred embodiment, the support structure can be made of a hydrophobic material. In one embodiment, the reagent (at least one) can be a growth medium. In a specific modality, the growth medium can be a means of indicator growth. In various embodiments, the reagent (at least one) can be cells of at least one bacterial strain, an enzyme, or an enzyme substrate. The invention also provides methods for detecting the presence or amount of biological material in a sample. In one aspect, the method comprises the steps of: 1) providing an incubation plate with at least one cavity containing at least one reagent for detecting the biological material, the plate comprising a generally flat horizontal surface defining multiple cavities in depression, each of which is adapted to contain an aliquot of liquid, and being of a size and shape and made of a suitable material to contain the aliquot of liquid within the cavity by surface tension and wherein at least one cavity contains at least one reagent for the detection of biological material; 2) liquefying the test sample if necessary and distributing the sample on the surface of the incubation plate; 3) drain any excess liquid; and 4) incubating the plate until the presence or absence of the biological material in one or more cavities is determined so that the amount of biological materials can be determined. The dispensing step may comprise the immersion of the plate in the sample, or emptying and optionally may include swirling or tilting. This aspect of the invention can be adapted for all separate embodiments with respect to the previously described plate devices. In another aspect the invention provides a method for detecting the presence or amount of a biological material in a sample, comprising the steps of: 1) providing a sterile incubation plate with a lower portion that is a generally flat horizontal surface and contains multiple cavities in depression adapted to contain an aliquot of liquid and to be dimensioned and shaped and formed of a material suitable for containing the liquid within the cavity by surface tension, and a cover with at least one projection containing at least one reagent for the detection of biological materials, wherein the projection fits into individual cavities when the lid is closed on the lower plate portion; 2) liquefying the test sample if necessary and distributing the sample on the surface of the incubation plate; 3) drain any excess liquid from the plate; 4) closing the lid in the lower portion of the plate in such a way that the reagent (s) on at least one projection contacts aliquots of liquid in the cavities, thus allowing the distribution of the reagent (s); and 4) incubating the plate until the presence or absence of the biological material in one or more cavities is determined so that the amount of biological materials can be determined. This aspect of the invention can be adapted with all the modalities with respect to the methods described above, and all the modalities with respect to the above-described plate and lid device. In another aspect, the invention provides a method for making an incubation plate with at least one reagent, the method comprising the steps of 1) providing a plate with a generally flat horizontal surface with depressed cavities that are sized and shaped and formed of a suitable material to have aliquots of test sample within each cavity by surface tension; and 2) drying at least one reagent in the cavity (s). In preferred embodiments, at least one cavity can be treated by corona discharge prior to the drying step. In another embodiment, the portion treated by corona discharge may consist essentially of an internal surface of at least one cavity. In a preferred embodiment, the reagent can be a cell growth medium or a bacterial growth medium. The plate can be made so that it adapts to all the modalities of the plates previously described. In another aspect, the invention provides a method for making an incubation plate comprising the steps of 1) providing a sterile incubation plate with a lower portion that is a generally flat horizontal surface and containing depressed cavities, each cavity being adapted for containing an aliquot of liquid and being dimensioned and shaped and formed of a suitable material to contain an aliquot by surface tension, and a lid with at least one projection that fits within the individual cavities with the lid being closed on the portion of the bottom plate; and 2) drying the reagent (s) on the projection (at least one). This aspect may contain all the embodiments of the invention described above with respect to the other plate methods and devices. In another aspect, the invention provides a method for manufacturing a device useful for detecting an analyte (s) or microorganism (s) in a test solution, the method comprising the steps of 1) providing a material capable of absorbing a predetermined volume of liquid by amount of material; 2) prepare at least one reagent islet of the material; 3) combining the material with a medium to detect the presence or quantity of the analyte or microorganism; and 4) securing the islet of reagent to a substantially hydrophobic support structure.

In various embodiments, the preparation step may comprise preparing multiple reagent islets. In another embodiment, the reagent islets may be capable of absorbing a predetermined volume of solution. In another aspect, the invention provides a method for detecting an analyte or microorganism in a test sample comprising the steps of 1) contacting the test sample with at least one islet of reagent capable of absorbing a predetermined volume of test solution and immobilized on a support structure, and comprising a means for determining the presence or absence of an analyte or microorganism located on or within the reagent pool (s); 2) separating the test sample from reagent islet (s) after the reagent islet (s) has absorbed a predetermined amount of sample; 3) subjecting the reagent islet (s) to reaction parameters that allow the development of the reagent and the generation of a sensitive signal; and 4) determining the presence or amount of an analyte or microorganism. In another aspect, the invention provides a method for detecting an analyte or microorganism in a test sample comprising the steps of 1) selecting a test solution for the detection of the analyte or microorganism; 2) providing a device comprising a substantially hydrophobic support structure and at least one reagent island (s) immobilized on the support structure, and which is capable of absorbing a predetermined volume of the test solution and comprises a medium to determine the presence or amount of the analyte (s) or microorganism (s); 3) contacting the device with the test solution for a sufficient time to allow the reagent cell (s) to absorb the predetermined volume and 4) allow the medium to determine the presence or amount of an analyte (s) or microorganism (s) to determine the presence or amount of the analyte (s) or microorganism. In one embodiment, the contact step may involve introducing the device into the test solution and removing it from the test solution. In another embodiment, the step of providing the device may comprise providing a means of determination comprising a reagent that produces a sensitive signal meaning the presence or amount of an analyte (s) or microorganism (s). In another embodiment, the step of allowing the means of determination may comprise subjecting the device to sufficient reaction parameters to allow the development of the reagent. Another step can be added to the method that includes observing the means of determination or the step of determining the presence or amount of an analyte (s) or microorganism (s), or a step of determining the amount of the analyte or microorganism.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1A is a plan view of a first embodiment of the device for the determination of analyte in solution of the present invention; Figure 1 B is an enlarged cross-section of Figure 1 A taken on lines 1 B-1 B; M Figure C is a side view of the first embodiment of the present invention of Figure 1A; Figure 2A is an enlarged view of Figure 1 B showing a cavity without a bevel; Figure 2B is an enlarged view of Figure 1 B showing a cavity with a bevel; Figure 3A is a plan view of a cover for the device of the first embodiment of Figure 1A; Figure 3A is an enlarged partial side view of the lid of Figure 3A; Figure 3C is a side view of the lid of Figure 3A; Figure 3D is a partially exposed perspective view of the lid of Figure 3A placed on the device of Figure 1A; Figure 4A is a plan view of a second embodiment of a lid having a plurality of projections on the internal surface thereof; Figure 4B is an enlarged section of the cap of Figure 4A taken on lines 4B-4B Figure 4C is a side view of the cap of Figure 4A; Figure 4D is an enlarged, partially exposed perspective view of a portion of the lid of Figure 4A showing the projections thereof; Figure 5A is a plan view showing the cover of Figure 4A placed on the device of Figure 1A; Figure 5B is an exposed, amplified view of Figure 5A; Figure 5C is a side view of Figure 5A; Figure 6A is a plan view of a second embodiment of the device of the present invention having a handle; Figure 6B is a cross-sectional view of Figure 6A taken on lines 6B-6B; Figure 6C is a side view of the second embodiment of the device of Figure 6A; Figure 7A is a plan view of the embodiment of Figure 6A having a lid placed thereon; Figure 7B is a cross-sectional view taken on line 7B-7B of Figure 7A; Figure 7C is a side view of Figure 7A; Figure 8 is a plan view of a third embodiment of the present invention that can be used as a single dip dipstick device, Figure 9 is a side view of Figure 8; Figure 10 is a fourth embodiment of the device of the present invention that can be employed as a dipstick test device having multiple reagent islets; Fig. 11 shows the device of Fig. 10 in which multiple tests have been performed; Figure 12 is a perspective view of the fifth embodiment of the device of the present invention in which a dipstick test device having two planes with multiple reagent islets thereon is removably insertable into a tube with a lid about it; Figure 13 is a perspective view of a sixth embodiment of the device of the present invention in which a plurality of reagent islets are located in more than one plane on a rod and are removably removable in a tube having a removable lid on it. the same; and Figure 14 is a perspective view of a seventh embodiment of the device of the present invention in which a laboratory pipette contains an elongated block having a plurality of reagent islet on one or more sides thereof.

DESCRIPTION OF THE PREFERRED MODALITIES Definitions "Microorganism" - microorganisms means any microbe including bacteria, fungi or protista. "Bacteria" - all organisms belonging to the Monera kingdom. "Biological material" - any material derived from a biological organism. "Target organisms" - any "microorganisms", preferably but not limited to E.coli, yeast and / or mo. "Cell" - any cell found in a plant, animal, bacteria or any other living organism. "Analyte" - any atom or molecule or ion that is involved in cellular metabolism in a living organism. "Target analyte" - any "analyte", preferably but not limited to metabolites and / or enzyme. "Proaceous material" - pros, peptides, enzymes or amino acids. "Hydrophobic" - having a sufficient degree of hydrophobicity to avoid "interference" or formation of bridges in liquids, such as samples or reagents, between adjacent cells, cavities or islets of incubation.

Structure The present invention provides a simpler method for the precise quantification of the number of microorganisms in a sample or for the quantification of any or other type of analyte, such as a biological material in discrete particles within a sample. Such biological materials include fungi or other living organisms, as well as pro aggregates, such as enzymes or even cofactors, using reaction mixtures well known to those skilled in the art. The invention generally uses a novel article that is designed to absorb and contain a preselected volume of a liquid sample and to provide a reagent or reagents for said sample volume. In the preferred embodiments, the preselected volume is divided into a plurality of sample aliquots. The plurality of sample aliquots may all be of the same volume or may be in series of aliquots, where each set contains aliquots of different volumes. The device used is generally in the form of a sterile incubation plate having a multitude of cavities capable of containing aliquots of separate liquids. The device is constructed to contain at least one reagent provided to at least one cavity and preferably to a plurality of or all of the individual cavities. Since the plate, which may include a lid, contains a reagent or reagents the reagent (s) is present in the plate before the introduction of a sample. For example, said reagent may be a growth medium specific for bacteria. The reagent (s) is provided prior to the addition of sample in a manner such that the reagent is not significantly washed or dissolved during the addition of the sample. The delivery of reagent or reagents into the plate eliminates the need for a test device to separately prepare the reagent and then add it to a sample or to the test plate. further, in many applications, the construction of the incubation plate is preferably arranged so that the use of a pipette is not required, the plate is simply immersed in the sample, or an approximate amount of the liquefied sample is emptied onto the surface of the sample. plate, and a designated volume of sample is retained within the cavities. In this way, the board provides a self-cleaning function. Each cavity to which the reagent or reagents is provided can receive the same reagent or different reagents. Thus, where a plurality of reagents are used, the separate cavities may receive different individual reagents, different combinations of reagents or combinations of these possibilities. The phrase "at least one cavity is supplied with at least one reagent" means that one or more reagents are contained in the plate before the addition of sample in a manner such that the reagent or reagents are present in a significant amount per at least in one of the individual cavities during incubation. In this way, the phrase distinguishes situations where the user adds the reagent together with the sample or in such a manner that the reagent is substantially diluted or dissolved in the sample before draining any excess liquid from the sample on the plate. In some embodiments, not all cavities will contain each reagent or may even contain no reagent. Thus, preferably the reagent or reagents are present in such a way that a discrete amount of reagent or reagents is supplied in or to each individually designated receiving cavity. Although it is possible for a user to prepare a plate by depositing a reagent in the cavities of a plate prior to the addition of the sample, preferably the plate is provided to the user of the reagent (s) already deposited and preferably dried in place. In general, the cavities are designed to form separate incubation chambers for each sample aliquot. The cavities may be of the same size or of different sizes and shapes to increase the count scales and / or to simulate dilution effects. Therefore, in a first aspect, the invention has a sterile incubation plate having a generally flat horizontal surface. The surface defines a plurality of depressed cavities (in preferred embodiments, at least 40, 60, 90 or even 200 depressed cavities are provided) and each cavity is adapted to contain aliquots of liquid by surface tension. Any excess liquid from the liquid sample drains from the surface of the plate out of the cavities due to the hydrophobic nature of the material used to form the plate. The plate can be constructed of plastic material or other hydrophobic material. In other embodiments, the plate may be generally circular in shape or have any shape. The plate is constructed to contain at least one reagent, for example, a growth medium, and to provide at least that reagent individually to at least one cavity. In this way, the reagent (s) is provided inside the plate instead of being added separately.

In general, the reagent or reagents are supplied directly to individual cavities. In other embodiments, different cavities may contain different reagents or different combinations of reagents to provide a variety of applications. For example, providing different reagents or different combinations of reagents to different cavities can provide a plurality of different tests on a single plate. The reagent (s) is provided in such a way that it is not appreciably washed during the aliquot preparation or aliquot self-preparation procedure (as described below). In other preferred embodiments, a cap is also provided to prevent contamination of the liquid within the cavities; and the plate is provided in a sterile form so that positive aliquots are not noticed unless at least one particle of biological material is present in the sample. In embodiments in which a lid is present, the lid is considered as part of the incubation plate, whether fixed or detached from the plate portion containing the cavities. In this context, the lid is the "lid portion", and the plate portion containing the wells is the "lower plate portion". The lid may be a top cover, but may also be constructed as a container, such as a clam shell arrangement, so that the plate portion containing the cavities may be completely contained within the lid / container. In another preferred embodiment, the lid is designed as a rectangular tube, such that the plate portion is inserted into the open end of the tube, generally an enlarged portion of the plate will close the open end of the tube-shaped lid. In a preferred embodiment, the reagent is coated in individual cavities in a manner such that the reagent is not appreciably washed out or dissolved from the cavity during the process of distributing the sample in the cavities. As an example, the inner surface of the cavities can be treated by corona discharge so that the reagent will adhere tightly to the plate when it dries and will therefore dissolve only slowly when it comes into contact with the liquid sample. Additional agents can be added to the medium or the cavities can be coated on the medium for additional control of the rate of dissolution. In other preferred embodiments, the plate includes a cap having a structure designed and adapted to deliver the reagent (s) to the individual cavity (s). For example, the cap may be constructed with a projection or protrusion containing the reagent, and preferably a plurality of said projections or protuberances, such as a projection for each cavity. The lid can be fixed to the lower portion of the plate or it can be separated. When the lid is closed or placed on the lower portion of the plate after preparing the aliquot of the sample, the reagent will make contact with the liquid sample in the cavity and then dissolve in the liquid sample. The reagent can be applied as a coating on the projection, such as by the use of corona discharge treatment of the projection surface to adhere the reagent to the projection. The reagent can also be dried on the projection or projections of the lid. The projection may also have a cavity for containing the reagent, the surface of which may be treated to improve reagent adhesion. Different reagents or different combinations of reagents can be applied as a coating on each of the projections. In other embodiments, the plate has a portion of reagent assortment. The reagent assortment portion is constructed to contain a selected amount or selected amounts of one or more reagents and to deliver the reagent (s) individually to one or more cavities. For example, the reagent assortment portion may be a frame attached to the underside of a lid portion with rings or cylinders containing the reagent or reagents. When the lid closes after the distribution of the sample, the reagent or reagents will make contact with the individual sample aliquots and will disperse in the liquid. Other configurations can also be designed. The generally flat horizontal surface is designed to allow the liquid to be aliquoted easily between the cavities and then the excess liquid drains from the plate while retaining a liquid sample aliquot within each of the cavities. Those skilled in the art will recognize that the depth and shape of the cavities, as well as the material used to make the cavities and the plate, are chosen in such a way that the surface tension can be used to contain the aliquots against gravity within each cavity, depending on the type of liquid used in the liquefied sample; those skilled in the art will understand the factors for selecting the plate materials and the appropriate cavity sizes for the various liquids. Preferably the sample is an aqueous solution, most preferably a suspension or diluted aqueous solution. In a preferred embodiment, the cavity has a diameter of approximately 0.38 or 0.41 centimeters. Preferably, a cavity contains between 5 and 100 μl. Also in preferred embodiments, the plate may have a shape designed to increase retention of a sample aliquot and / or reagent retention within the cavity. An example of such an alternative cavity shape is a generally circular cavity with ribs projecting toward the center of the cavity, but other designs can also be used. In a preferred embodiment, the cavities are bevelled to allow the liquid that is above the horizontal plane to be easily emptied (see FIG. 2B). The dimensions of the cavities and the number of cavities on the plate can be selected so that a particular total volume is retained on a plate. Preferably a plate retains a total between about 0.1 and 10 ml, most preferably about 0.5, 1 or 2 ml, within the cavities. The incubation plate can be formed of any desired material, but in particular it is desirable that a plastic is used which allows separate aliquots of the liquefied sample to be contained by surface tension within each cavity without cross-contamination of the cavities. Preferably, the material is hydrophobic. The surface can be untreated or treated chemically or physically to increase the retention of liquid within the cavities, even when the plate is inverted or directed to any angle from the horizontal. The plate can also be treated to increase the adhesion of a reagent or reagents within a cavity or on another desired surface of the plate, for example it can be treated by corona discharge. Generally, the reagent will dry on the surface of the plate. In other preferred embodiments, the incubation plate is clear or colored, for example, white or yellow (to increase the appearance of color (eg, blue) within the incubation plate), and the plate has rectangular dimensions of approximately 3.81 by 6.35 or 1.54 by 10.16, or for circular plates, a diameter of approximately 7.62 or 12.7 centimeters. In related aspects, the invention relates to methods for the detection of a biological material in a sample using an incubation plate having at least one reagent as described above. The methods include the steps of liquefying the sample (if necessary) and distributing the liquefied sample on the surface of an incubation plate as described for the above aspect, preferably immersing the plate in the sample. In an alternative embodiment, the sample is emptied into the plate and the plate is tipped or swirled as necessary to distribute the sample; in this embodiment, the area containing the cavities of the plate is surrounded by a wall that retains the sample on the plate during the distribution of the sample. As described above, each cavity is adapted to contain an aliquot of liquid and is dimensioned and shaped, and formed of a suitable material, to contain the aliquot within the cavity by surface tension. Aliquots of liquid enter the individual cavities without being applied individually, and therefore the method incorporates an automatic aliquot preparation (or auto aliquot). Any excess liquid in the liquefied sample is drained from the surface of the plate to the outside of the cavities due to the hydrophobic nature of the material used to form the plate. In one method, the plate has at least one reagent in at least one cavity before the addition of the sample, so that the next aliquot preparation of the liquid sample in the cavities, the reagent or reagents dissolve in the liquid. In a related method, the reagent or reagents are contained in the cap, such as on or within projections on the cap projecting into the cavities when the cap is closed on the lower plate portion. In this way, when the lid is closed, the reagent (s) makes contact with the aliquot of the liquid in the individual cavity or cavities, and then disperses and dissolves in the liquid. In the context of this invention, the term "submerge" refers to a brief immersion of at least a portion of an incubation plate in a liquid. Preferably, the immersion period is less than about 3 seconds, most preferably less than about 2 seconds, and most preferably still less than about 1 second. As described, the plate is constructed to contain at least one reagent, such as a growth medium, and preferably all reagents necessary for the particular test where the reagent is on the plate prior to sample distribution. In other embodiments, different cavities may contain different reagents to provide a variety of applications, for example, to provide a plurality of different tests on a single plate. The reagent (s) is provided in such a way that it does not wash off considerably during the aliquot self-preparation procedure. Any necessary reagents not provided directly on the device can be provided in the sample or directly on the plate when used. The method then comprises incubating that incubation plate until the presence or absence of the biological material is determined. Also in preferred embodiments, the plate is provided with a handle portion by means of which the user can take the plate while avoiding contact with the sample application portion of the plate. For applications where the sample is distributed on the surface of the plate by immersing the plate in the sample, at least a portion of the handle is not immersed in the sample, thus helping to prevent contamination of the plate and / or the sample by contact with the user's hand.

The shape of the incubation plate is not of fundamental importance, but in preferred embodiments it is a generally rectangular shape. Another example is a generally circular shape (such as that of a Petri dish). Actually, the incubation plate can be used to take the place of a Petri dish. Specifically, the method of this invention can be used to replace existing tests that are generally carried out in Petri dishes to record the number of microbial colonies. Because discrete aliquots of the sample are provided on the plate, one skilled in the art only needs to record the number of positive cavities in the plate to define the amount of biological material within the original sample, as with the MPN test that is described earlier. In preferred embodiments, the incubation plates of these methods are constructed of plastic, or other hydrophobic materials. As mentioned above, the biological material that can be detected is any material that forms a discrete particle, such as a microorganism, which can be quantified by determining the presence or absence of said biological material within each cavity of the incubation plate. The sample can be any biological or environmental sample such as waste water, feed, swab surface sample, or sample from other swab surfaces, such as pharyngeal swab, or other samples well known to those skilled in the art. This sample can be a liquid sample, or it can be dissolved in a liquid to form the liquefied sample. Thus, the term "liquefaction" refers to providing the sample in a liquid with which aliquots can be prepared rapidly within the incubation plate. The liquefied sample may remain as a liquid or may solidify, for example, gel, in the cavities after the excess liquid has been removed. This invention provides a very useful device and method that allows non-expert personnel to quickly determine the amount of biological material within a sample. Because the sample is easily liquefied by people without significant training in microbiology, and the materials for any specific tests can be provided by the manufacturer, such people can easily carry out the tests accurately. The incubation plate is usually provided in sterile form so that inappropriate detection of biological material can not occur. Although it is known to provide plastic containers that can contain liquid within a plurality of depressions, this device and method provide an automatic aliquot preparation method (or self-preparation of aliquots) in which the steps that the person going to make will perform. The test is reduced and simplified by eliminating the requirement of manual addition of a reagent and preferably also eliminating the need to pipette a sample or other liquid into the plate. This is an improvement over the existing products used to detect and quantify microorganisms as they reduce the potential for reagent contamination or errors in dilution. The present device can be used particularly in the analysis of foods and in tests of clinical samples. The separation of the cavities of the present device prevents interference or contamination between each aliquot. Because of this, many of the tests can be performed by observing the fluorescence (which is not easily done in a Petri dish containing agar). The device is particularly useful when there is a large number of microorganisms present in a sample, such as more than one organism per one ml or per ten ml. Other features and advantages of the invention will be apparent from the following description of the preferred embodiments thereof, and from the claims. Referring to Figures 1A-1 D, there is shown an incubation plate 10 having a plurality of cavities 12 each having a diameter of about 0.40 cm. Incubation plate 10 has rectangular dimensions of approximately 6.85 x 3.55 cm. The incubation plate is made of molded plastic. The cavities 12 are sufficiently separated to prevent interference between the cavities. These cavities may have a bevel (Fig. 2B) if desired to prevent the liquid from remaining at the upper edge of the cavity. The internal surfaces of the cavities can be treated by corona discharge and a reagent, such as a growth medium, can be dried on those surfaces. The reagent can be either a single reagent or a combination of reagents. Those skilled in the art will recognize that the incubation plate 10 can be easily formed by standard procedures and fabricated with or without a boundary wall, and with or without a cover 14 (Figures 3A-3C). This lid is provided with depressions 16 to prevent contact of the lid with the plate 10. Referring to Figures 4A-4D and 5A-5C, a rectangular incubation plate 10 is shown with a corresponding lid 20. The lid is constructed with a series of projections or protrusions 32 which project into the cavities 12 of the plate when the lid closes on the plate. The reagent can be applied to the ends of these protrusions so that when the cap is placed on the plate, the protrusions project into the cavity and the reagent dissolves in the test sample. The lid can be separated from the bottom plate or it can be formed as an integral part. For example, the lid portion can be attached to the lower plate portion with a thin plastic "hinge portion", which will allow the lid to be closed in the lower plate portion. In this embodiment example, each of the lugs of the lid has a recess 34 in the tip, in which a reagent is deposited. The inner surface of the cavity can be treated by corona discharge to increase reagent adhesion, and then the reagent can be dried in the treated region by corona discharge. In use, aliquots would be prepared with the sample in the cavities, then closing the lid would put the tips of the protrusions in contact with the liquid in the cavities. This contact will result in the dissolution of the reagent contained in the cavities at the tips of the projections in each of the sample aliquots. In the case when the reagent is a bacterial growth medium and the sample contains bacteria that can grow in that medium, then bacterial growth can occur in the cavities in which there are viable bacteria. Referring to Figures 6A-6C, a rectangular incubation plate 10 is shown which is similar in construction to the plate of Figures 1A-1C, except that it has a handle portion 18. A user can take the portion of grip, thus avoiding possible accidental contact with the sample application area of the plate. Furthermore, in applications in which the plate is immersed in a liquid sample, by taking the handle portion, contact of the user's fingers or a gripping device with the overall sample can be avoided, thus reducing the possibility of accidental contamination of the sample. the sample. The plate 10 of Figures 6A-6C is also shown in Figures 7A-7C, together with a lid 20. This example of a lid is formed to completely enclose the plate, and therefore can prevent contamination of the underside of the a submerged plate, and prevent contamination of the surfaces of the work table or other objects with the liquefied sample from the underside of a submerged plate. Accordingly, in use, the cavity-containing portion of the plate is immersed in a liquefied sample, excess liquid is drained from the plate, and the plate is inserted into the lid / container for incubation. The lid / container 20 can be kept closed by using friction coupling pressures 22 in the upper and lower portions of the lid. The cavities of the plate can be bevelled.

Coating the surfaces of the plate with reagent A variety of different methods can be used to provide a reagent or reagents within an incubation plate. For example, a reagent could be provided so that the reagent would dissolve easily and quickly upon contact with a pH regulator or solution, preferably an aqueous solution. In this embodiment, the amount of liquid added must be controlled so that the resulting concentration of reagent distributed in the cavities is appropriate. Therefore, in this case, the reagent could be placed inside the incubation plate in any manner and location that retains the reagent during handling but allows rapid dissolution when adding the liquid. In this invention, however, the reagent is provided in a manner and / or location on the plate so that it is not necessary to measure the volume of liquefied sample with which the plate is contacted in order to obtain an appropriate concentration of reagent within the sample aliquots retained in the cavities. This is achieved by providing a discrete amount of the reagent directly to each cavity in which the presence of the reagent is desired. Said discrete quantities may be provided in a variety of ways, including the particular embodiments described herein. These modalities either use a reagent adhered to the surface of the cavity in a manner such that the amount of reagent that dissolves during the aliquot self-preparation procedure is negligible, or provide a reagent at a location so that it does not enter contact with the liquid until after the aliquot self-preparation procedure is complete. For example, the inner surface of one or more cavities can be treated by corona discharge and the reagent can be dried on the treated surface, such as a highly concentrated reagent solution. The reagent deposited in this manner will dissolve slowly, so that the amount lost during the aliquot self-preparation process is negligible. Most of the reagent will then dissolve during the initial part of the incubation period. The rate of dissolution can also be controlled by selecting the components of the media. In another example, the reagent can be deposited on projections on the surface of the plate cap. The projections are positioned so that the tips thereof project into the individual cavities when the lid is closed on the plate. The reagent can be deposited on the external surface of the projections, but is preferably deposited in the cavities at the tip of the projections. The reagent can be deposited in any manner by which the reagent is substantially retained during handling prior to the use of the plate. Thus, for example, the reagent can be retained by physical barrier means, but preferably adheres to the surface of the projection and / or the surface of a cavity at the tip of the projection, such as by a process that understand treatment by corona discharge and drying of the reagent on the treated surface.

Treatment by corona discharge and reagent deposit Typical plastics are formed by long chains of linked subunits, for example, polyethylene is composed of long chains of ethylene subunits. In general, aqueous solutions and solutes in aqueous solutions are not attracted to interior chain subunits and therefore will not bind effectively to such locations. In contrast, there is often much more affinity between said solutes and chain ends, particularly where the chain end has a relatively polar character. As understood by those skilled in the art, corona discharge treatment is a method that increases the hydrophilic character of a treated plastic surface. The corona discharge treatment procedure generally involves passing the plastic through an electric arc. The energy imparted by the electric arc introduces a significant number of chain breaks, thus increasing the number of relatively hydrophilic ends available for interaction. In addition, the electric arc also generates ozone, which, as a strong oxidizing agent, generates, however, additional chain breaks and polar side groups by oxidizing bonds within the polymer chains.

As indicated, this treatment procedure increases the binding of polar compounds, such as components of aqueous solution. Therefore, for the use of corona discharge treatment in preparing plates of this invention, after treatment by corona discharge of the cavity surfaces, a concentrated reagent solution is dispensed into the cavities. As the surfaces treated by corona discharge can now be moistened (eg, hydrophilic) the liquid reagent forms a uniform coating on the surface of the cavity. The plate is then dried, generally in a drying oven, removing the water and leaving a hard coating of solid reagent in the cavities. As mentioned before, different reagents can be provided to different cavities, and / or some cavities may not receive any reagents. Cavities that do not receive reagent would not usually be treated by corona discharge. Other methods known in the art can also be used to increase the adhesion of the reagent to the appropriate plate surfaces.

Use In use, a test sample can be liquefied or diluted with an appropriate sterile pH regulator or saline solution. Then a quantity of the liquefied sample can be distributed on the surface of the plate. In a preferred embodiment of use, the liquefied sample is expected to contain or dilute to contain about 1-40 units of the biological material to be quantified, for example, viable bacterial cells. An incubation plate 10 of for example Figures IA-IC is immersed in the liquid sample sufficiently to fill the cavities, is removed, and allowed to drain briefly holding it at approximately 90 ° with respect to the horizontal. In this use mode, the board provides both a self-prepared aliquot function and a self-dispensing function. Alternatively, in the case of a plate having a delimiting wall, an excess of the liquefied sample is placed in the incubation plate 10 and that liquid is spun into the incubation plate 10 to distribute the inoculated liquid to the plate. each of the cavities 12. The incubation plate 10 is then held at an angle of approximately 90 ° to allow excess liquid to drain from the plate. As shown in Figures 3A-3C, a lid 14 can then be placed on the incubation plate and that plate kept in an incubator for the appropriate time, for example 18-48 hours. After that time, the presence or absence of a positive result can be recorded in each cavity 12 of the plate.

EXAMPLE 1 Use of incubation plate for global test For the total plate count, a plate as described above is used for the detection and quantification of the total bacterial concentration of food. It can be based on a multiple enzyme technology that correlates the activity of enzymes with the presence of viable bacteria in food. It uses multiple enzyme substrates that produce a blue fluorescent color when metabolized by bacteria. The multiple enzyme reagent is applied as a coating on the interior surfaces of the cavities of the plate. When a sample of liquefied prepared food is distributed in the cavities of a plate as described herein, the total viable bacterial concentration of this food product can be determined after 24 hours of incubation. The actual medium used here is not of fundamental importance for the invention, but is provided solely for purposes of illustration.

Storage and disposal Unused test plates are stored at room temperature (4o to 25 ° C) away from light. After use, the incubation plate device will contain viable bacteria which must be handled and disposed of properly.

Test procedure - sample application of immersion method 1.- A sample is obtained by removing an appropriate quantity of a bulk material to be tested. If necessary, the sample is liquefied or diluted in sufficient volume to allow a plate to be immersed in the sample. 2. - An incubation plate is taken without making contact with the sample application surface (preferably the plate is taken by the handle portion if the plate includes said portion). The lower portion of an open incubation plate is immersed in the test, the plate is removed immediately, and maintained at approximately 90 ° to the horizontal to allow excess liquid to drain from the plate. Preferably the excess liquid is allowed to drain into a waste container instead of allowing it to drain back into the sample container in order to reduce the possible deposit of the reagent in the sample container. Be sure to eliminate all liquid "cross bridges" between the cavities by lightly hitting the plate. The excess liquid is removed appropriately. 3.- Put the lid back on the plate (or close a lid attached). 4.- Place the plate in an incubator for 24 hours. The plates can be inverted if desired. 5.- The number of fluorescent cavities is counted after 24 hours by placing a UV light of 6 wats and 365 nm to 12.7 cm of the plate. The plate is not read before 24 hours. The results are stable at 48 hours. 6.- The number of fluorescent cavities is compared to an MPN table to determine the most probable number of bacteria present in the plate.

EXAMPLE 2 Use of incubation plate for unit dose test The plate containing media described in example 1 is used for this test.

Test procedure 1.- Add 10 ml of sterile pH regulator (or saline) to a tube. If a sample of food greater than 0.1 ml is to be inoculated into the test, the volume of the sterile pH regulator is reduced appropriately to achieve a final volume of 10 ml in the tube. 2.- The pH regulator is inoculated with the food sample that is being tested. 3.- The tube is shaken several times to completely mix the pH regulator and the inoculated food sample. 4. If the tube is of sufficient size, immerse the plate in the sample or alternatively pour the pH / sample regulator suspension onto the sample application surface of an incubation plate as appropriate, taking care that all the cavities are filled. After the sample application, immediately hold the plate at approximately 90 ° to the horizontal to allow excess liquid to drain from the plate. Preferably the excess liquid is allowed to drain into a waste container. Be sure to eliminate all liquid "cross bridges" between the cavities by tapping the plate lightly. The excess of liquid is eliminated in an adequate way. 5.- Put the lid back on the plate (or close a lid attached). 6.- Place the plate in an incubator for 24 hours. The plates can be inverted if desired. 7.- The number of fluorescent cavities is counted after 24 hours by placing a UV light of 6 watts and 365 nm to 12.7 cm of the plate. The plate is not read before 24 hours. The results are stable at 48 hours. 8.- The number of fluorescent cavities is compared with an MPN table to determine the most probable number of bacteria present in the plate. The documents of the patent and other references mentioned herein are each incorporated by reference thereto as if each had been incorporated separately by reference in its entirety.

Dipstick Test Device for Detection and Enumeration of Microorganisms Next with reference to Figures 8-13, the concept is to use small individual absorbent materials containing a lyophilized or "dip-dried" reagent in a setting that allows a distribution of automatic sample and enumeration of target microorganisms.

A series of absorbent (hydrophilic) materials containing reagent ("reagent islets") are immobilized in a support structure made of a hydrophobic material to form individual reagent islets. The reagent islets can be embedded in the support structure, for example. However, this should not be interpreted as limiting since any way to fix the islet of reagent to the support structure can be used as long as it does not interfere with the test. The rate of liquid absorption of each islet of reagent in this mode is the same since the islets of reagent are made of the same material and have the same size. Additional configuration can be made so that there are two or more groups of reagent islets in different sizes to form an adjustment that can be used to achieve higher quantification of microorganisms without serial dilution based on the principles of the Plus Number method. Probable (MPN). In a preferred embodiment, the reagent is one developed to test the presence of target microorganisms in a sample using, but not limited to, defined substrate technology (DST) and / or Multiple Enzyme Technology (MET). An objective positive detection of microorganisms in said reagents can cause a change in the color of the reagent and / or cause the emission of a fluorescent signal from the reagent when viewed under a long-wave UV lamp. Examples of such reagents are Colilert®, Enterolert ™, Simplate TPC ™, and Simplate Cec ™. The sample inoculum is precalculated based on the maximum absorption rate (saturation point) of an islet of reagent and the total number of reagent islets in a device. When the predetermined amount of liquid sample (ie, homogeneous water, milk, juice and food) is inoculated into this device, each islet of reagent absorbs the same amount of sample. The sample can be distributed to each islet of reagent simply by moving the device from back to front or in circular motion. The area between each islet of reagent is hydrophobic and when each islet of reagent absorbs the sample to its saturation point, there will be no more sample between the reagent islets and cross-contamination between reagent islets will be avoided. The liquid sample absorbed by the reagent islets also rehydrolyzes the reagent in the reagent islets to support the growth of microorganisms in the sample. After incubation of the device containing the sample at a predetermined temperature, the reagent islets containing microorganisms targeted by the sample will have a color change and / or emit fluorescent signals (positive); Reagent islets that do not have target microorganisms in the sample will not show color change or emit fluorescent signals (a negative result). The concentration of target microorganisms in the sample being tested can be calculated based on the number of positive and negative reagent islets observed using the Most Probable Numbers (MPN) method. Another application of this concept is to have lyophilized reagent islets with different types of reagents. Each reagent is designed to test a specific aspect of a sample, such as microorganisms, chemicals, or any other detectable analyte of interest. The combination of these islets of reagent in the same device forms a test kit that provides a one-step test for multiple chemicals, analytes, or biological materials. Specifically referring to FIGS. 8 and 9, another aspect of the invention is shown comprising an immersion rod 100 containing an islet of reagent 101. The rod will generally be made of plastic, but its composition is not of fundamental importance and It can build from any hydrophobic material that does not cause leaching in the test sample or interference with the test. Figures 8 and 9 illustrate the rod with a single reagent islet. Figure 10 illustrates a preferred embodiment of the dipstick containing multiple islets of reagent 102. Reagent islet zones may be placed on the dipstick with each zone having reagent islets containing different reagents or combinations of reagents other than those other zones, thus allowing multiple tests on a single immersion rod, each zone testing a different analyte or microorganism. Figure 11 illustrates the dipstick test device of Figure 10 which has gone through the test procedure and illustrates islets of reagent showing both positive results (dark reagent islets) and negative results (white reagent islets). Figure 12 shows a preferred embodiment of this aspect of the invention. An immersion rod 100 with multiple islets of reagent 101 is shown which is bent and inserted into a test tube 103. In this embodiment, the test tube also has a cap 104 which also protects the islets of factor reagent environmental Of course, the device can be placed in other types of containers such as a plastic sleeve or any container that serves to protect the device. The reagent placed in the islet of reagent can be any reagent or combination of reagents that can be distributed on or in the reagent islet material.

Application of test sample to the dipstick device The methods for applying samples to the device are varied.

The immersion rod can be immersed in the test sample, and left in contact long enough for the islets of reagent to absorb a predetermined amount of fluid. This will generally be less than 3 seconds, but may be more depending on what material the reagent islets are constructed from. In a preferred embodiment, the reagent islets are constructed of cellulose. However, they can also be constructed of any material that is capable of absorbing and containing a volume of fluid. The test sample can also be pipetted from the test solution to the reagent islets with a transfer pipette. The support structure may also comprise a box made of plastic or other suitable hydrophobic material. The reagent islets can be placed inside the box and the box can be opened and the test solution is applied using a pipette or by pouring it, or by any appropriate means. The box may have a lid to further protect the device fixed on it or as a separate piece. Referring now to Figure 13, the support structure may also comprise a central support 104 to which are attached "sheets" 105 that hold the islets of reagent 106. The sheets may be arranged in a three-dimensional structure as illustrated in FIG. Figure 13, thus maximizing the number of sheets that can be accommodated in the device. The device can be placed in a circular container 108 and a lid 107 is placed on it to provide greater protection for the device before, during, and after developing the test.

Pipetting Device Referring now to Figure 14, in another aspect, the invention provides a device comprising a support structure 109 which is contained within a laboratory pipette 111. Multiple islets of reagent 110 are contained on or within the support structure 109. The reagent islets comprise a solid absorbent material, such as cellulose, which is first cut into the desired shape and fixed to the support structure 109. Preferably, the support structure 109 has more than one side with reagent islets 110. The support structure 109 is most preferably formed of a synthetic polymer, such as plastic, and inserted into a laboratory pipette 111 which preferably comprises glass or plastic and has a tapered end for absorption of the sample. The sample can be simply introduced into the pipette by means of a suction apparatus known in the art, left for a sufficient time for the islets of reagent to absorb a predetermined volume of test sample, and is removed from the device. Then the device is incubated for a sufficient time to develop the test, and the results are determined. The pipette itself comprises both a simple means for applying the sample to the test device, and a protective function.

EXAMPLE 1 Bacterial detection system for coliforms and E. coli in milk using the dipstick device The following is an example of how the present invention provides a method of detecting coliforms in milk that is easy, does not involve many steps, and provides results that are easy to interpret. The test was conducted using a test device with multiple islets of reagent, each of which absorbed .033 ml of milk. Milk was added to the test device and with it aliquots were self-prepared for all reagent islets. The sample was incubated for 24 hours and provided the number of coliforms present by examining how many of the reagent islets changed color. The number of coliforms present is determined based on the MPN table. The data obtained from the test is shown in Table 1. The flexibility of the absorption ability of the test device is accentuated since the volume absorption can easily be adjusted by adjusting the number and / or sizes of the reagent islets. Also, the device also has the ability to have multiple tests on the same device, impregnating islets of reagent with different media.

TABLE 1 Milk samples Immersion wand device (cfu / ml) # 7197A 10.3 # 7197B 11.8 # 7197C 16.4 # 710 56.3 # 713 18 # 3B1 0 # 3B2 37 # 2B2 0 # 2B1 9.3 # 618 69.1 # 618A A11 + # 622A 12 # 622B 36 # 622C 3 # 7697A 56 # 7697B 22 # 7697C 16 # 7697D 31 # 811 3 # 815 16 # 813 6.2 Average: 23.41 Standard deviation 20.75 The invention has been described in detail with particular reference to certain preferred embodiments thereof, but it will be understood that variations and modifications may be made within the spirit and scope of the invention.

Claims (24)

NOVELTY OF THE INVENTION CLAIMS

1. - A test device for determining the presence or quantity of an analyte or a microorganism in a sample, said device comprising: a pipette having two open ends, one of which is tapered for the absorption of liquid; an elongate support structure contained within said pipette and comprising at least one surface; and a plurality of islands of absorption of discrete sample, each of said absorption islands is in said elongated support structure, each of said islets is adapted to contain an aliquot of liquid and is sized, configured, and shaped as a material appropriate to contain said aliquot within said absorption islet, said absorption islets contain at least one reagent for the detection of an analyte or a microorganism, further characterized in that said device will not provide any positive response for said analyte or microorganism in the absence of said analyte or microorganism present in a sample applied to said device.

2. The device according to claim 1, further comprising a plurality of different reagents.

3. The device according to claim 2, further characterized in that different reagents are provided to different islets of sample absorption between the plurality of discrete sample absorption islets.

4. The device according to claim 3, further characterized in that the different reagents provided to the different islets among the plurality of discrete sample absorption islets comprise different tests.

5. The device according to claim 1, further characterized in that said reagent (at least one) comprises a specific medium for a target organism.

6. The device according to claim 5, further characterized in that the target organisms are E. coli and / or coliform bacteria.

7. The device according to claim 5, further characterized in that the target organisms are total viable bacteria.

8. The device according to claim 5, further characterized in that the target organisms are Enterococci.

9. The device according to claim 1, further characterized in that said elongated support structure has more than one surface and said islets are placed in said more than one surface.

10. The device according to claim 1, further comprising means for sucking liquid into and out of said pipette.

11. - The device according to claim 1, further characterized in that said pipette has marks to check the suction of a predetermined amount of liquid inside said pipette.

12. A test device for determining the amount of an analyte or a microorganism in a sample, comprising: a pipette having two open ends, one of which is tapered for the absorption of liquid; an elongate support structure contained within said pipette and comprising two or more surfaces; and a plurality of islands of absorption of discrete sample, each of said islets is in said two or more surfaces of said elongated support structure, each of said islets is adapted to contain an aliquot of liquid and is dimensioned, configured, and In the form of a material suitable for containing said aliquot inside said absorption islet, said absorption islets contain at least one reagent for the detection of an analyte or a microorganism, further characterized in that said device will not provide any positive response for said analyte or microorganism in the absence of said analyte or microorganism present in a sample applied to said device.

13. The device according to claim 12, further comprising a plurality of different reagents.

14. The device according to claim 13, further characterized in that different reagents are provided to different islets between the plurality of said islets.

15. - The device according to claim 14, further characterized in that at least some of said different reagents provide different tests.

16. The device according to claim 12, further characterized in that said reagent (at least one) comprises a specific medium for cells or at least one bacterial species.

17. The device according to claim 12, further comprising means for sucking liquid into and out of said pipette.

18. The device according to claim 12, further characterized in that said pipette has marks to check the suction of a predetermined amount of liquid inside said pipette.

19. A test device for determining the amount of an analyte in a sample, said device comprising: a pipette having two open ends, one of which is tapered, said pipette having markings to check the suction of a predetermined amount of liquid inside said pipette; an elongate support structure contained within said pipette and comprising at least one surface; and a plurality of discrete sample absorption islands, each of said absorption islands is in said support structure, each of said islets is adapted to contain an aliquot of liquid and is sized, configured, and shaped as an appropriate material to contain said aliquot within said absorption islet, said absorption islets contain at least one reagent for the detection of an analyte present in a sample applied to said device; and means for sucking liquid in and out of said pipette.

20. The device according to claim 19, further comprising a plurality of different reagents.

21. The device according to claim 20, further characterized in that different reagents are provided to different islets between the plurality of said islands.

22. The device according to claim 21, further characterized in that at least some of said different reagents provide different tests.

23. The device according to claim 19, further characterized in that said reagent (at least one) comprises a specific medium for a target organism.

24. The device according to claim 19, further characterized in that said elongated support structure has more than one surface and said islets are placed in said more than one surface.