CA2303384A1 - Food contamination detection system - Google Patents
Food contamination detection system Download PDFInfo
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- CA2303384A1 CA2303384A1 CA002303384A CA2303384A CA2303384A1 CA 2303384 A1 CA2303384 A1 CA 2303384A1 CA 002303384 A CA002303384 A CA 002303384A CA 2303384 A CA2303384 A CA 2303384A CA 2303384 A1 CA2303384 A1 CA 2303384A1
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- food
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- food contamination
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Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54366—Apparatus specially adapted for solid-phase testing
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/02—Food
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Chemical & Material Sciences (AREA)
- Food Science & Technology (AREA)
- General Physics & Mathematics (AREA)
- Urology & Nephrology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Physics & Mathematics (AREA)
- Medicinal Chemistry (AREA)
- Pathology (AREA)
- Biomedical Technology (AREA)
- Analytical Chemistry (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Microbiology (AREA)
- Cell Biology (AREA)
- Biotechnology (AREA)
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Investigating Or Analyzing Non-Biological Materials By The Use Of Chemical Means (AREA)
Abstract
A food contamination indicator includes a first layer impermeable support, a second capillary layer and a third impermeable cover film layer. The second layer includes an absorbent material providing for the directional flow of juices, an immunobead solution pad and a detection area. The capillary layer is supported by the impermeable support layer, and covered by the impermeable cover film layer, such that the capillary layer rests between the impermeable support layer and the impermeable cover film layer.
Description
The field of the present invention is the detection of substances or contaminants in edible consumer products.
Over the past several years there has been increasing concern over the safety of our food supply. Contamination of food can come from a variety of sources and the type of contamination possible is often dependent on the type of food involved.
Most animal derived food products, such as raw meat, are exposed to contamination before, during, or after processing. In most cases, contamination is minor and, if the food is prepared properly, may not pose a serious threat to the consumer. However, while the contamination of food is generally low, i.e. few bacteria per gram of food, if the food is not stored under satisfactory conditions, or if the food is stored for long periods of time, contaminants, such as bacteria, may grow to become a serious threat to the eventual consumer. Even if the food products reach the market in an acceptable condition, subsequent mistreatment by the consumer may lead to the development of food contamination.
Food products are often "mass produced" and sold at retail outlets in prepackaged containers. Such packages typically include a styrofoam, plastic or cardboard tray which supports the food product. The tray and food are sealed in a transparent plastic wrap material and a liner lies between the food product and the inside bottom of the tray. A bar code is often used on the products for scanning at the check-out register, to reduce errors in totaling purchases and for stock control.
The bar code comprises a series or pattern of bars, which represent a number identifying the product.
A number of incidents and factors have lead to the growing concern over the food supply. These include:
raw chicken and egg products have been found to be contaminated with Salmonella and inadequate cooking of such products has led to serious illness or death of persons who have consumed the contaminated products;
inadequately pasteurized milk products have been found to be contaminated with Listeria which has lead to serious illness or death of consumers of the products;
AMENDED SHEET
Over the past several years there has been increasing concern over the safety of our food supply. Contamination of food can come from a variety of sources and the type of contamination possible is often dependent on the type of food involved.
Most animal derived food products, such as raw meat, are exposed to contamination before, during, or after processing. In most cases, contamination is minor and, if the food is prepared properly, may not pose a serious threat to the consumer. However, while the contamination of food is generally low, i.e. few bacteria per gram of food, if the food is not stored under satisfactory conditions, or if the food is stored for long periods of time, contaminants, such as bacteria, may grow to become a serious threat to the eventual consumer. Even if the food products reach the market in an acceptable condition, subsequent mistreatment by the consumer may lead to the development of food contamination.
Food products are often "mass produced" and sold at retail outlets in prepackaged containers. Such packages typically include a styrofoam, plastic or cardboard tray which supports the food product. The tray and food are sealed in a transparent plastic wrap material and a liner lies between the food product and the inside bottom of the tray. A bar code is often used on the products for scanning at the check-out register, to reduce errors in totaling purchases and for stock control.
The bar code comprises a series or pattern of bars, which represent a number identifying the product.
A number of incidents and factors have lead to the growing concern over the food supply. These include:
raw chicken and egg products have been found to be contaminated with Salmonella and inadequate cooking of such products has led to serious illness or death of persons who have consumed the contaminated products;
inadequately pasteurized milk products have been found to be contaminated with Listeria which has lead to serious illness or death of consumers of the products;
AMENDED SHEET
a highly toxic stain of E. coli has lead to the death of several people who consumed prepared beef products which had been inadequately cooked;
a number of toxins are known, such as ciguatoxins, which contaminate fish. These toxins are not inactivated or destroyed by cooking and so their presence in fish is a threat to any consumer of the product;
shell fish, such as oysters, concentrate contaminants present in the water in which they grow and, since they are frequently eaten raw, pose a threat to the health of consumers; and fish are increasingly eaten raw which adds to the possibility of increased outbreaks of illness from water borne contaminants.
There has been a need for a reliable way to detect if a food product purchased by a consumer is fit for consumption. Any solution to this problem should be relatively inexpensive and able to detect a number of agents capable of causing illness. It should also be simple to read so that consumers, who do not have access to sophisticated testing equipment or specialized knowledge, can readily determine if the products they have purchased are free from contamination.
Co-pending United States Application Serial Nos. 08/584,984 filed January 11, 1996, 08/197,297 filed February 16, 1994 (WO-A-9427144), and 08/758,205 filed November 26, 1996 (WO-A-9814777) and United States Patent No. 5,306,466 issued April 26, 1994 (all of which are incorporated by reference as if fully set forth herein) describe inventions for the detection of contaminants in food.
EP-A-0306336 discloses assay methods and devices specifically adapted for conducting assays requiring that a sample and at least one other liquid reagent be added during the assay.
EP-A-0587222 discloses a thin film immunoassay device with an increased wash volume capacity.
It is an object of the invention to develop a more consumer friendly test device and method for detecting bad food.
According to the invention, this is achieved by the features of claim 1 or 16.
Advantageous further embodiments are described in the subclaims.
A1~'!~ENDED SHEET
a number of toxins are known, such as ciguatoxins, which contaminate fish. These toxins are not inactivated or destroyed by cooking and so their presence in fish is a threat to any consumer of the product;
shell fish, such as oysters, concentrate contaminants present in the water in which they grow and, since they are frequently eaten raw, pose a threat to the health of consumers; and fish are increasingly eaten raw which adds to the possibility of increased outbreaks of illness from water borne contaminants.
There has been a need for a reliable way to detect if a food product purchased by a consumer is fit for consumption. Any solution to this problem should be relatively inexpensive and able to detect a number of agents capable of causing illness. It should also be simple to read so that consumers, who do not have access to sophisticated testing equipment or specialized knowledge, can readily determine if the products they have purchased are free from contamination.
Co-pending United States Application Serial Nos. 08/584,984 filed January 11, 1996, 08/197,297 filed February 16, 1994 (WO-A-9427144), and 08/758,205 filed November 26, 1996 (WO-A-9814777) and United States Patent No. 5,306,466 issued April 26, 1994 (all of which are incorporated by reference as if fully set forth herein) describe inventions for the detection of contaminants in food.
EP-A-0306336 discloses assay methods and devices specifically adapted for conducting assays requiring that a sample and at least one other liquid reagent be added during the assay.
EP-A-0587222 discloses a thin film immunoassay device with an increased wash volume capacity.
It is an object of the invention to develop a more consumer friendly test device and method for detecting bad food.
According to the invention, this is achieved by the features of claim 1 or 16.
Advantageous further embodiments are described in the subclaims.
A1~'!~ENDED SHEET
A first, separate aspect of the present invention, is directed to an improvement on the previously described methods of detecting contaminants in food by providing for the directional flow of the fluids or juices bringing these fluids or juices into contact with a detecting element. Such directional flow advantageously enhances the sensitivity and reliability of the detection system, permitting the analysis of a larger volume and more representative sample of juices.
A second, separate aspect of the present invention, is directed to an indicator dependent on the directional capillary flow of the fluids wherein contamination is to be detected. The indicator includes a first layer impermeable support, a second capillary layer having an immunobead solution pad and a detection area, this second layer providing for the directional flow of juices and being supported by the impermeable support, and a third impermeable cover film layer, having a impermeable cover film, for covering the capillary layer, such that the capillary layer rests between the impermeable support layer and the impermeable cover film layer.
A third, separate aspect of.the. present invention, is directed to a code indicia which may be modified or rendered unreadable by a contamination detector providing for the directional capillary flow of the fluids where contamination is to be detected. The code indicia may include at least one, or preferably at least two, bar codes, at least one of which is modified or rendered unreadable in the presence of the antigen or antigens the indicator is intended to detect. Alternatively, the code indicia may be characterized by utilizing, with the indicator, a first bar code for which the change in appearance of the indicator in the presence of an antigen or antigens makes the first bar code unreadable by a bar code reader, and a second bar code for which the change in appearance of the indicator in the presence of an antigen or antigens makes the second bar code readable by a bar code reader.
A fourth, separate aspect of the present invention, is directed to an indicator dependent on the directional capillary flow of the fluids wherein contamination is to be detected. The indicator includes a first layer impermeable support, a second capillary layer having an immunobead solution pad and at least two detection areas, this second layer providing for the directional flow of juices and being supported by ;~.~,:..~,1~~';, eHE~T
.".nVw.~. Jry.W r 1 the impermeable support, and a third impermeable cover film layer, having a impernzeable cover film, for covering the capillary layer, such that the capillary layer rests between the impermeable support layer and the impermeable cover film layer and each detection area is in alignment with a window in the cover film and a code indicia.
A fifth, separate aspect of the present invention, is directed to a method for the detection of food contamination for use with a food container adapted for containing a food product which produces juices. The juices are sequentially drawn through a first absorbent material into and through an immunobead solution pad, including a first set of antibodies specific to an antigen to be detected, the first set of antibodies being linked to a light absorbing material, and through a substrate, including a second set of antibodies specific to the antigen to be detected, the first set of antibodies, or to the combination thereof, said second set of antibodies being bound to a membrane in a detection area.
Figure 1 is a longitudinal cross sectional view of a contamination detector for use in food safety according to a preferred embodiment.
Figure 2 is a top plan view of the contamination detection system shown in Figure 1.
Figure 3 is a top plan view of an alternative preferred embodiment of a contamination detection system.
Figure 4 is a longitudinal cross-section schematic of an alternative arrangement for a contamination detector within the scope of the present invention.
The invention is based on the solid-phase immunobead assay and antibody sandwich principles modified to facilitate the continuous flow of product juices and contaminants. The contaminants may be microorganisms, bacteria, viruses, toxins or any other immunogenic materials or substances, and are herein referred to as "antigen" or "antigens." The assay may preferably be linked to a uniform product code bar system. The antigen or antigens are detected as they pass through the contamination detection system by binding to antigen-specific colored immunobead complex which then migrates to be captured by a second specific antibody. The AMENDED ~HEE7-S
second specific antibody is attached to a membrane forming part of the uniform product code bar system. The presence of the contaminating antigen is evident by a localized color bar formation.
The appearance-of a bar-indicating~ the presence -of antigea~~can then be determined by simple optical scanning of the bar code attached to the product or by visually observing the appearance of a symbol, a letter, a word, or the like.
The scanner can be programmed to read the type of contamination, product, and record ~ the date and location of the reading. The membranes are designated to allow entry of antigen, prevent the entrance of interfering substances, and maintain the complex formed between the immunobead and the antigen to be detected inside the system.
The contamination detection system remains closely associated with the product, thus allowing for early detection and tracking product safety from slaughter or processing/packaging plant to the consumer. The presence of a visual symbol on or in addition to the code indicia may also permit the determination of product safety in the absence of scanning devices, such as in the consumer's home.
Figure 1 is a longitudinal cross sectional view of a contamination detector for use in food safety. The contamination detection system may be used in a preformed well of a food package as described in the above referenced patent and applications or, advantageously, it may be embedded in the food package itself.
Alternatively, the contamination detection system may be embedded or closely associated with a liner, which commonly is present in the package of food products and lies between the tray and the food, such that juices collected in the liner are drawn into the contamination detection system.
In any case, the components of the contamination detection system are substantially the same. As shown in Figure 1, a cover film 2 composed of mylar acetate or some other type of transparent nonporous film contains a code indicia (not shown) such as the ubiquitous UPC bar code which codes for a particular food product that is contained in the package in which the particular contamination detection system is associated. The bars of the bar code may be printed onto the film and the light reflective spaces may be created by using adhesive white paper 4 ~1,~,~ci~~ED SKEET' or some other method such as painting or staining the back of the cover film 2, provided that light may be reflected to a bar code reading device such as a laser bar code scanner, light pen, CCD or other such device that can detect the bars and spaces of the bar code.
In the case of a bar code, the cover film 2 preferably has a window 6 which contains at least one space of the bar code. Window 6 is aligned with detection area 26 (See Figure 2), of substrate 8. Typically, the prepackaged food product will be ~ shipped to a retail outlet such as a grocery store which will attach its own bar code label (cover film 2) to and in alignment with detection area 26. When the contamination detection system detects contamination, the light reflectivity of detection area 26 will change from being light reflecting to light absorbing.
Typically, for many UPC bar codes that means a color change from white to black.
However, as known by those of ordinary skill in the art of bar code scanning, other colors, shades, or metallic lusters may be used to accomplish this effect.
Once detection area becomes light absorbing, a bar code reading device will be unable to successfully decode the code indicia of cover film 2 and contamination can be detected.
Further detection methods that may be used with the present invention are described in the above referenced patent and patent applications. For example, these may include symbols, letters, words, and the like, which become readable or unreadable (depending on the implementation), in the presence of the antigen or antigens to be detected. In particular, the indicator area 26 may be placed such that it obliterates part of the product identification code rendering it incomplete to avoid the accidental checkout of contaminated product, or it may modify it, such that it represents a different code. One or several indicator areas may be present in a substrate 8. Alternatively, the code indicia may be characterized by utilizing, with the indicator, a first bar code for which the change in appearance of the indicator in the presence of antigen makes the first bar code unreadable by a bar code reader, and a second bar code for which the change in appearance of the indicator in the presence of antigen makes the second bar code readable by a bar code reader.
A11~ENDED SH~EE7-Additionally, the indicator area 26 could include a number which appears upon contamination to complete a numeric code corresponding to the data code by the contamination detecting bar code. With these additional elements, a checkout clerk could not inadvertently price contaminated product when bar code is not recognized or read by the scanner.
Above substrate 8 is migration film 10 which is composed of mylar acetate or some other transparent nonporous film. Below substrate 8 is a support 12 which can be a plastic strip, a film such as mylar acetate or some other nonporous film or substance. These two components ensure that liquids pass through the contamination detection system by the inflow orifice 14 described herein.
Figure 2 is a top plan view of the contamination detection system shown in Figure 1. As shown in Figures 1 and 2 the contamination detection system functions as follows. Liquids associated with a food product such as blood and juices from meat and fish, or liquids packaged with the food or the food itself, such as milk and juices, enter an inflow orifice 14 of the contamination detection system. The liquid passes through a protective filter 16 which filters out larger materials such as proteins and clumped dead cells. The liquid then passes through a selective filter 18 that filters out other materials and molecules larger than the toxin of interest such as ciguatoxin, brevetoxin or other toxins associated with food poisoning, or a bacteria of interest such as E. coli, Salmonella, Listeria, Campylobacter or other bacteria associated with food poisoning. The filters are not considered essential.
Moreover, other means of filtering, well known in the art, may be used.
Absorbent pad 20 includes a primary absorbent material 22. As used herein, "absorbent pad" or "absorbent material" means any material providing for the directional capillary flow of juices or fluids. These materials include, for example, blotting paper, Whatman paper, structures having thin capillary conduits made of any of the several well known in the art materials that do not detrimentally affect the sample travelling therein, and the like. After the filtered liquid passes through the primary absorbent material 22, it passes through the immunobead solution pad 24.
Immunobead solution pad 24 contains monoclonal antibodies bound to colored latex AMENDED SHEET
microspheres, and is created by standard techniques as later described herein.
As the filtered liquid passes through the immunobead solution pad 24, antigens bind to monoclonal antibodies that recognize the specific antigen. Advantageously, the immunobead solution pad may have one type of monoclonal antibody or multiple types of monoclonal antibodies to ensure detection of a single antigen.
Alternatively, the immunobead solution pad may have multiple types of monoclonal antibodies to detect multiple antigens. Alternatively, the immunobead solution pad may use polyclonal antibodies instead of and/or in addition to monoclonal antibodies.
The preparation of such antigen-specific antibodies is well known in the art.
In some cases it may be necessary to conjugate a toxin antigen to a protein to "mask"
the toxicity of the antigen. Otherwise injection of the toxic antigen may result in the death of the animal in which the antibodies are to be prepared. Methods of conjugating compounds are well known in the art and one such method is described by Hokama et al., Mycotoxins and Phycotoxins 188, A Collection of Invited Papers at the Seventh International ILJPAC Symposium of Mycotoxins and Phycotoxins, Tokyo, Japan 1988, pp. 303-310 (Elsevier Science Publishers, Amsterdam), which is incorporated herein by reference.
The filtered liquid now laden with monoclonal antibodies bound to both a colored latex microsphere and an antigen passes from the immunobead solution pad 24 to substrate 8. Substrate 8 is a membrane, such as Immobilon-P, to which another monoclonal antibody is bound to a detection area 26. Alternatively, a polyclonal antibody may be used on substrate 8 to form detection area 26. The detection area 26 may encompass the entire area of the substrate 8, or only a portion of it. Antibodies on the substrate 8 can be specific to the antibodies contained in the immunobead solution pad, the antigen or antigens, the antigen to be detected bound to the antibodies contained in the immunobead solution pad, or mixtures thereof.
Although the first and second antibodies could be the same, they are preferably different. The second antibody preferably recognizes different antigenic determinants on the antigen than the first antibody. Consequently, as the filtered liquid passes across substrate 8, the antigens bound to antibodies bound to the colored latex A1~ENDED ShtEET
microsphere are captured in detection area 26. As the number of antibodies bound to the colored latex microsphere and a toxin or bacteria increases, so does the color of detection area 26 so that eventually the light reflectivity of detection area 26 changes from light reflecting to light absorbing at which point contamination is considered to have occurred. The filtered liquid is then withdrawn by a secondary absorbent material 28.
Figure 3 is a top plan view of an alternative embodiment of the contamination detection system. This embodiment functions essentially the same as the embodiments shown in Figures 1 and 2 except that secondary absorbent material also functions as a filter which prevents antibodies bound to a colored latex microsphere and antigen to pass beyond the substrate 8 area to the outflow orifice.
The membrane forming substrate 8 is channeled to allow the antibodies bound to a colored latex microsphere and a antigen to recirculate over the detection area to further enhance the sensitivity of the contamination detection system.
Figure 4 is a longitudinal cross-section schematic of an alternative arrangement for a contamination detector within the scope of the present invention.
As shown in the figure, a first layer includes an impermeable support 12, serving as a support for a second layer having an immunobead solution pad 24, described previously. Two or more substrates 8 and 9 (two shown in the figure), having detection areas 26 and 27, described previously, abut and are in contact with the immunobead solution pad 24 in the second layer. A third layer has a cover film composed of a transparent nonporous film that separates code indicia 40, such as the UPC bar codes shown 42 and 44, from the second layer. The detection areas 26 and 27 are aligned with the window 6 or windows in the cover film 2 and the code indicia 40, 42, 44.
In use this alternative embodiment draws product juices through inflow orifice 14 into or onto the immunobead solution pad 24. As the juices pass through the immunobead solution pad 24, antigens, if present, bind a first set of antibodies in the immunobead solution pad 24. The fluid - now laden with the first set of antibodies bound to both a colored latex microsphere and an antigen - passes from the ANxENDED SHEET
immunobead solution pad 24 onto the substrates 8 and 9. As the fluid passes across substrates 8 and 9, the first set of antibodies bound to both a colored latex microsphere and an antigen are captured in detection areas 26 and 27 by a second set of antibodies specific to the antibodies contained in the immunobead solution pad, 5 the antigen or antigens, the antigen bound to the antibodies contained in the immunobead solution pad, or mixtures thereof. As the number of antibodies bound to the colored latex microspheres and antigen increases, so does the color of ~ detection areas 26 and 27, so that eventually the light reflectivity of detection areas 26 and 27 changes from light reflecting to light absorbing. At that point 10 contamination is considered to have occurred.
The code indicia may have a first bar code 44 for which the change in appearance of the first indicator 27 in the presence of antigen makes the first bar code 44 unreadable by a bar code reader, and a second bar code 42 for which the change in appearance of the second indicator 26 in the presence of antigen makes the second bar code 42 readable by a bar code reader. Other alternatives, explained in detail throughout this application, may also be used with this embodiment.
EXAMPLES
As an illustrative example, a contamination detection system for the detection of E. coli may be produced as described below.
Materials Anti-E.coli 0157:H7 polyclonal antibody, 1 mg/mL, commercially available from Kirkegaard & Perry Laboratories, Inc., Gaithersburg, MD; catalog number 95-90. Sodium phosphate buffer (SPB), 0.1 M, pH 6Ø Phosphate buffered saline (PBS), pH 7.4. Polystyrene (latex) black microspheres, 0.388~xn diameter, 10%
solids, commercially available from Bangs Laboratories, Inc., Carmel, IN;
catalog No. D000388PK. Bovine serum albumin (BSA). Blocking solution consisting of 1 % BSA in PBS. Glycine, CZH3N0z, FW = 75.07. Second blocking solution consisting of 1 M glycine in PBS. Immobilon-P (PVDF) membrane commercially available from Millipore Corporation, Bedford, MA; catalog No. IPVH 101 00.
Methanol CH30H HPLC grade, commercially available from Chempure, Houston, :~~~~WED Sh~EET
TX; catalog No. 831-296. Immunobead pad, made from dried alcohol swab material from Baxter Scientific Products, McGaw Park, IL; catalog B3062. Sucrose 50%
solution in PBS. Deionized distilled (dd) water. Plastic support S-PIA stick, commercially available from Millipore Corp. Acetate film, as used in overhead transparencies. Absorbent paper Whatman # 42. Non-toxic glue stick, available commercially from Conros Corp., Detroit, MI Ross 5tik # 06-074.
PBS ( 1 OmM) standard solution may be prepared according to Harlow, E. and D. Lane, Antibodies: a laboratory manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY at 726 ( 1988). Antibody stock solution ( 1 mg/mL) may be prepared by rehydration with 50% glycerol solution, according to antibody manufacturer's instructions. SPB may be prepared as indicated in "Protocols for Adsorbing Proteins one Polystyrene-Based Microspheres," by Bangs Laboratories, Inc., 9025 Technology Drive, Fishers, IN 46038-2886, telephone (317) 570-7020, Fax (317) 570-7034. Surfactants may be removed from microspheres as indicated in "How to Work with Microspheres," and "Protocols for Adsorbing Proteins onto Polystryrene-Based Microspheres," by Bangs Laboratories, Inc. The immunobead solution may be prepared following the Sodium Phosphate/PBS/BSA method indicated in "Protocols for Adsorbing Proteins onto Polystyrene-Based Micrpspheres,"
by Bangs Laboratories, Inc. For example, a 6 mL final volume of immunobead solution may be prepared with 120 ~,L antibody solution, 300 p,L black latex microspheres, and 5,580 ~L SPB.
Preparation of Immunobead Pad An immunobead pad for use in the present invention may be prepared as follows. Alcohol is evaporated from the Baxter alcohol swab (pad). The pad is then rinsed by repeated immersion in dd water. The step may then be repeated in a new amount of dd water, and the pad dried in an oven at 37 °C. The dried pad is placed on a glass surface and wetted with 1 % BSA using a pipettor, and then dried by suspending it from a corner inside an oven at 37 °C. The dried pad is placed on a glass surface and wetted with 50% sucrose using a pipettor, and then dried in an oven at 37 °C. The dried pad is placed on a glass surface and wetted with ANI~h~DED SHEET
immunobead solution using a pipettor. About 15 p,L of immunobead solution is applied to a 0.9 cm x 0.9 cm section of blocked pad. The pad is again dried in an oven at 37 °C for about 10 minutes. The drying process is completed at 4 °C in a refrigerator. The immunobead pads may be stored between sheets of non-adhesive paper tissue in plastic bags at 4 °C, and may be cut into appropriately sized pieces before contamination detection system assembly.
Preparation of Detection Area in the Substrate Immobilon-P membrane (Millipore Corp.), is pre-wetted as provided with the manufacturer's instructions. While the Immobilon-P membrane remains wet (opaque), primary antibody is applied with a 10-~L pipettor (about 3 ~.L of anti-E.
coli polyclonal antibody per membrane) in the shape desired for the detection area.
For example, in the case of a bar code a vertical bar (0.7 cm x 0.2 cm) is applied.
The antibody solution preferably has a concentration of 20 p,g/mL, PBS being used as the diluting solution. If the membrane partially dries (i.e. the membrane shows traces of white coloration), the pre-wetting procedure should be repeated. The applied antibody-is-dried-by-incubatingthe-membrane-overa-KimwipeTM tissue in an oven at 37 °C for IO-15 minutes, or in a refrigerator at 4 °C
for a longer period.
The membrane is then blocked by immersing the membrane in 1 % BSA
blocking solution for 30 minutes at room temperature. It is desirable that bubbles be prevented from forming on the surface of the membrane during the immersion procedure. The membrane is then three times rinsed by immersions in PBS, each immersion lasting about five minutes. Excess PB5 is eliminated by vertically placing the membrane on KimwipeTM tissue for about two minutes. The membrane is then dried on KimwipeTM tissue in an oven at 37 °C for 15 minutes or in the refrigerator at 4 °C for a longer period. Once dry, the blocked membrane is ready for assembly into the contamination detection system.
Assembly of the Contamination Detection System According to Preferred Embodiments A contamination detection system may be assembled by cutting an 5-PIA
plastic strip into two 4.5 cm x 1 cm pieces, one of them with the attached membrane ~~C7ED SHEET
with antibody in detection area 26. Using an additional piece of S-PIA plastic strip (3 cm x 1 cm) as a basis, the two 4.5 cm x 1 cm pieces are glued end to end so that the strip returns to its original dimensions (9 cm x 1 cm), but with the membrane on the central part of the plastic strip. A thin line of non-toxic glue is applied along both edges of the strip, except on the membrane. A 0.9 cm x 0.9 cm piece of immunobead pad is attached to the strip so that one edge is flush against the membrane. A 1.5 cm x 1 cm piece of absorbent paper is attached to the plastic strip ~ so that it is flush against the free end of the immunobead pad, while a second 1.5 cm x 1 cm piece of absorbent paper is attached to the plastic strip so that it is flush against the free end of the membrane. At this point the plastic strip should have four components glued side by side in the following sequence: paper-membrane-pad-paper.
Glue may then be applied in a thin line against both edges of a 2 cm x 0.9 cm piece of acetate. The acetate is glued so as to cover both the membrane and the pad. The acetate may slightly overlap on the piece of absorbent paper next to the membrane or pad.
A bar code may be incorporated into the assembled contamination detection system by attaching a second piece of acetate (7 cm x 3 cm) with a printed Uniform Product Code bar code to one end of the plastic strip such that the bars to be modified, or made unreadable, match the area where the antibody was applied (detection area 26) on the underlying membrane (substrate 8). The contamination detection system may be stored under refrigeration.
Thus, a food contamination detector providing for the directional capillary flow of juices has been disclosed. While embodiments and applications of this invention have been shown and described, it would be apparent to those skilled in the art that many more modifications are possible without departing from the inventive concepts herein. For example, while the example describes the use antibodies against E. coli, those of skill in the art will recognize that different antibodies, and antibodies to different antigen or antigens may be used.
Moreover, mixtures of antibodies, against a number of different antigens, or combinations -~~I~GEp ~i-~~t ~
thereof may be used. With the use of different antibodies, multiple different antigens which could be present in the sample can be detected. Those of skill in the art will also recognize other manners of assembling food contamination detectors within the teachings of the present disclosure. For example, the absorbent pad can be made of any material providing for the directional capillary flow of juices or fluids, the immunobead pad may be prepared using other materials and membranes well known in the art, as can the detection area. The invention, therefore is not to be restricted except in the spirit of the appended claims.
Gtd~~~, ~~.~~6__~
A second, separate aspect of the present invention, is directed to an indicator dependent on the directional capillary flow of the fluids wherein contamination is to be detected. The indicator includes a first layer impermeable support, a second capillary layer having an immunobead solution pad and a detection area, this second layer providing for the directional flow of juices and being supported by the impermeable support, and a third impermeable cover film layer, having a impermeable cover film, for covering the capillary layer, such that the capillary layer rests between the impermeable support layer and the impermeable cover film layer.
A third, separate aspect of.the. present invention, is directed to a code indicia which may be modified or rendered unreadable by a contamination detector providing for the directional capillary flow of the fluids where contamination is to be detected. The code indicia may include at least one, or preferably at least two, bar codes, at least one of which is modified or rendered unreadable in the presence of the antigen or antigens the indicator is intended to detect. Alternatively, the code indicia may be characterized by utilizing, with the indicator, a first bar code for which the change in appearance of the indicator in the presence of an antigen or antigens makes the first bar code unreadable by a bar code reader, and a second bar code for which the change in appearance of the indicator in the presence of an antigen or antigens makes the second bar code readable by a bar code reader.
A fourth, separate aspect of the present invention, is directed to an indicator dependent on the directional capillary flow of the fluids wherein contamination is to be detected. The indicator includes a first layer impermeable support, a second capillary layer having an immunobead solution pad and at least two detection areas, this second layer providing for the directional flow of juices and being supported by ;~.~,:..~,1~~';, eHE~T
.".nVw.~. Jry.W r 1 the impermeable support, and a third impermeable cover film layer, having a impernzeable cover film, for covering the capillary layer, such that the capillary layer rests between the impermeable support layer and the impermeable cover film layer and each detection area is in alignment with a window in the cover film and a code indicia.
A fifth, separate aspect of the present invention, is directed to a method for the detection of food contamination for use with a food container adapted for containing a food product which produces juices. The juices are sequentially drawn through a first absorbent material into and through an immunobead solution pad, including a first set of antibodies specific to an antigen to be detected, the first set of antibodies being linked to a light absorbing material, and through a substrate, including a second set of antibodies specific to the antigen to be detected, the first set of antibodies, or to the combination thereof, said second set of antibodies being bound to a membrane in a detection area.
Figure 1 is a longitudinal cross sectional view of a contamination detector for use in food safety according to a preferred embodiment.
Figure 2 is a top plan view of the contamination detection system shown in Figure 1.
Figure 3 is a top plan view of an alternative preferred embodiment of a contamination detection system.
Figure 4 is a longitudinal cross-section schematic of an alternative arrangement for a contamination detector within the scope of the present invention.
The invention is based on the solid-phase immunobead assay and antibody sandwich principles modified to facilitate the continuous flow of product juices and contaminants. The contaminants may be microorganisms, bacteria, viruses, toxins or any other immunogenic materials or substances, and are herein referred to as "antigen" or "antigens." The assay may preferably be linked to a uniform product code bar system. The antigen or antigens are detected as they pass through the contamination detection system by binding to antigen-specific colored immunobead complex which then migrates to be captured by a second specific antibody. The AMENDED ~HEE7-S
second specific antibody is attached to a membrane forming part of the uniform product code bar system. The presence of the contaminating antigen is evident by a localized color bar formation.
The appearance-of a bar-indicating~ the presence -of antigea~~can then be determined by simple optical scanning of the bar code attached to the product or by visually observing the appearance of a symbol, a letter, a word, or the like.
The scanner can be programmed to read the type of contamination, product, and record ~ the date and location of the reading. The membranes are designated to allow entry of antigen, prevent the entrance of interfering substances, and maintain the complex formed between the immunobead and the antigen to be detected inside the system.
The contamination detection system remains closely associated with the product, thus allowing for early detection and tracking product safety from slaughter or processing/packaging plant to the consumer. The presence of a visual symbol on or in addition to the code indicia may also permit the determination of product safety in the absence of scanning devices, such as in the consumer's home.
Figure 1 is a longitudinal cross sectional view of a contamination detector for use in food safety. The contamination detection system may be used in a preformed well of a food package as described in the above referenced patent and applications or, advantageously, it may be embedded in the food package itself.
Alternatively, the contamination detection system may be embedded or closely associated with a liner, which commonly is present in the package of food products and lies between the tray and the food, such that juices collected in the liner are drawn into the contamination detection system.
In any case, the components of the contamination detection system are substantially the same. As shown in Figure 1, a cover film 2 composed of mylar acetate or some other type of transparent nonporous film contains a code indicia (not shown) such as the ubiquitous UPC bar code which codes for a particular food product that is contained in the package in which the particular contamination detection system is associated. The bars of the bar code may be printed onto the film and the light reflective spaces may be created by using adhesive white paper 4 ~1,~,~ci~~ED SKEET' or some other method such as painting or staining the back of the cover film 2, provided that light may be reflected to a bar code reading device such as a laser bar code scanner, light pen, CCD or other such device that can detect the bars and spaces of the bar code.
In the case of a bar code, the cover film 2 preferably has a window 6 which contains at least one space of the bar code. Window 6 is aligned with detection area 26 (See Figure 2), of substrate 8. Typically, the prepackaged food product will be ~ shipped to a retail outlet such as a grocery store which will attach its own bar code label (cover film 2) to and in alignment with detection area 26. When the contamination detection system detects contamination, the light reflectivity of detection area 26 will change from being light reflecting to light absorbing.
Typically, for many UPC bar codes that means a color change from white to black.
However, as known by those of ordinary skill in the art of bar code scanning, other colors, shades, or metallic lusters may be used to accomplish this effect.
Once detection area becomes light absorbing, a bar code reading device will be unable to successfully decode the code indicia of cover film 2 and contamination can be detected.
Further detection methods that may be used with the present invention are described in the above referenced patent and patent applications. For example, these may include symbols, letters, words, and the like, which become readable or unreadable (depending on the implementation), in the presence of the antigen or antigens to be detected. In particular, the indicator area 26 may be placed such that it obliterates part of the product identification code rendering it incomplete to avoid the accidental checkout of contaminated product, or it may modify it, such that it represents a different code. One or several indicator areas may be present in a substrate 8. Alternatively, the code indicia may be characterized by utilizing, with the indicator, a first bar code for which the change in appearance of the indicator in the presence of antigen makes the first bar code unreadable by a bar code reader, and a second bar code for which the change in appearance of the indicator in the presence of antigen makes the second bar code readable by a bar code reader.
A11~ENDED SH~EE7-Additionally, the indicator area 26 could include a number which appears upon contamination to complete a numeric code corresponding to the data code by the contamination detecting bar code. With these additional elements, a checkout clerk could not inadvertently price contaminated product when bar code is not recognized or read by the scanner.
Above substrate 8 is migration film 10 which is composed of mylar acetate or some other transparent nonporous film. Below substrate 8 is a support 12 which can be a plastic strip, a film such as mylar acetate or some other nonporous film or substance. These two components ensure that liquids pass through the contamination detection system by the inflow orifice 14 described herein.
Figure 2 is a top plan view of the contamination detection system shown in Figure 1. As shown in Figures 1 and 2 the contamination detection system functions as follows. Liquids associated with a food product such as blood and juices from meat and fish, or liquids packaged with the food or the food itself, such as milk and juices, enter an inflow orifice 14 of the contamination detection system. The liquid passes through a protective filter 16 which filters out larger materials such as proteins and clumped dead cells. The liquid then passes through a selective filter 18 that filters out other materials and molecules larger than the toxin of interest such as ciguatoxin, brevetoxin or other toxins associated with food poisoning, or a bacteria of interest such as E. coli, Salmonella, Listeria, Campylobacter or other bacteria associated with food poisoning. The filters are not considered essential.
Moreover, other means of filtering, well known in the art, may be used.
Absorbent pad 20 includes a primary absorbent material 22. As used herein, "absorbent pad" or "absorbent material" means any material providing for the directional capillary flow of juices or fluids. These materials include, for example, blotting paper, Whatman paper, structures having thin capillary conduits made of any of the several well known in the art materials that do not detrimentally affect the sample travelling therein, and the like. After the filtered liquid passes through the primary absorbent material 22, it passes through the immunobead solution pad 24.
Immunobead solution pad 24 contains monoclonal antibodies bound to colored latex AMENDED SHEET
microspheres, and is created by standard techniques as later described herein.
As the filtered liquid passes through the immunobead solution pad 24, antigens bind to monoclonal antibodies that recognize the specific antigen. Advantageously, the immunobead solution pad may have one type of monoclonal antibody or multiple types of monoclonal antibodies to ensure detection of a single antigen.
Alternatively, the immunobead solution pad may have multiple types of monoclonal antibodies to detect multiple antigens. Alternatively, the immunobead solution pad may use polyclonal antibodies instead of and/or in addition to monoclonal antibodies.
The preparation of such antigen-specific antibodies is well known in the art.
In some cases it may be necessary to conjugate a toxin antigen to a protein to "mask"
the toxicity of the antigen. Otherwise injection of the toxic antigen may result in the death of the animal in which the antibodies are to be prepared. Methods of conjugating compounds are well known in the art and one such method is described by Hokama et al., Mycotoxins and Phycotoxins 188, A Collection of Invited Papers at the Seventh International ILJPAC Symposium of Mycotoxins and Phycotoxins, Tokyo, Japan 1988, pp. 303-310 (Elsevier Science Publishers, Amsterdam), which is incorporated herein by reference.
The filtered liquid now laden with monoclonal antibodies bound to both a colored latex microsphere and an antigen passes from the immunobead solution pad 24 to substrate 8. Substrate 8 is a membrane, such as Immobilon-P, to which another monoclonal antibody is bound to a detection area 26. Alternatively, a polyclonal antibody may be used on substrate 8 to form detection area 26. The detection area 26 may encompass the entire area of the substrate 8, or only a portion of it. Antibodies on the substrate 8 can be specific to the antibodies contained in the immunobead solution pad, the antigen or antigens, the antigen to be detected bound to the antibodies contained in the immunobead solution pad, or mixtures thereof.
Although the first and second antibodies could be the same, they are preferably different. The second antibody preferably recognizes different antigenic determinants on the antigen than the first antibody. Consequently, as the filtered liquid passes across substrate 8, the antigens bound to antibodies bound to the colored latex A1~ENDED ShtEET
microsphere are captured in detection area 26. As the number of antibodies bound to the colored latex microsphere and a toxin or bacteria increases, so does the color of detection area 26 so that eventually the light reflectivity of detection area 26 changes from light reflecting to light absorbing at which point contamination is considered to have occurred. The filtered liquid is then withdrawn by a secondary absorbent material 28.
Figure 3 is a top plan view of an alternative embodiment of the contamination detection system. This embodiment functions essentially the same as the embodiments shown in Figures 1 and 2 except that secondary absorbent material also functions as a filter which prevents antibodies bound to a colored latex microsphere and antigen to pass beyond the substrate 8 area to the outflow orifice.
The membrane forming substrate 8 is channeled to allow the antibodies bound to a colored latex microsphere and a antigen to recirculate over the detection area to further enhance the sensitivity of the contamination detection system.
Figure 4 is a longitudinal cross-section schematic of an alternative arrangement for a contamination detector within the scope of the present invention.
As shown in the figure, a first layer includes an impermeable support 12, serving as a support for a second layer having an immunobead solution pad 24, described previously. Two or more substrates 8 and 9 (two shown in the figure), having detection areas 26 and 27, described previously, abut and are in contact with the immunobead solution pad 24 in the second layer. A third layer has a cover film composed of a transparent nonporous film that separates code indicia 40, such as the UPC bar codes shown 42 and 44, from the second layer. The detection areas 26 and 27 are aligned with the window 6 or windows in the cover film 2 and the code indicia 40, 42, 44.
In use this alternative embodiment draws product juices through inflow orifice 14 into or onto the immunobead solution pad 24. As the juices pass through the immunobead solution pad 24, antigens, if present, bind a first set of antibodies in the immunobead solution pad 24. The fluid - now laden with the first set of antibodies bound to both a colored latex microsphere and an antigen - passes from the ANxENDED SHEET
immunobead solution pad 24 onto the substrates 8 and 9. As the fluid passes across substrates 8 and 9, the first set of antibodies bound to both a colored latex microsphere and an antigen are captured in detection areas 26 and 27 by a second set of antibodies specific to the antibodies contained in the immunobead solution pad, 5 the antigen or antigens, the antigen bound to the antibodies contained in the immunobead solution pad, or mixtures thereof. As the number of antibodies bound to the colored latex microspheres and antigen increases, so does the color of ~ detection areas 26 and 27, so that eventually the light reflectivity of detection areas 26 and 27 changes from light reflecting to light absorbing. At that point 10 contamination is considered to have occurred.
The code indicia may have a first bar code 44 for which the change in appearance of the first indicator 27 in the presence of antigen makes the first bar code 44 unreadable by a bar code reader, and a second bar code 42 for which the change in appearance of the second indicator 26 in the presence of antigen makes the second bar code 42 readable by a bar code reader. Other alternatives, explained in detail throughout this application, may also be used with this embodiment.
EXAMPLES
As an illustrative example, a contamination detection system for the detection of E. coli may be produced as described below.
Materials Anti-E.coli 0157:H7 polyclonal antibody, 1 mg/mL, commercially available from Kirkegaard & Perry Laboratories, Inc., Gaithersburg, MD; catalog number 95-90. Sodium phosphate buffer (SPB), 0.1 M, pH 6Ø Phosphate buffered saline (PBS), pH 7.4. Polystyrene (latex) black microspheres, 0.388~xn diameter, 10%
solids, commercially available from Bangs Laboratories, Inc., Carmel, IN;
catalog No. D000388PK. Bovine serum albumin (BSA). Blocking solution consisting of 1 % BSA in PBS. Glycine, CZH3N0z, FW = 75.07. Second blocking solution consisting of 1 M glycine in PBS. Immobilon-P (PVDF) membrane commercially available from Millipore Corporation, Bedford, MA; catalog No. IPVH 101 00.
Methanol CH30H HPLC grade, commercially available from Chempure, Houston, :~~~~WED Sh~EET
TX; catalog No. 831-296. Immunobead pad, made from dried alcohol swab material from Baxter Scientific Products, McGaw Park, IL; catalog B3062. Sucrose 50%
solution in PBS. Deionized distilled (dd) water. Plastic support S-PIA stick, commercially available from Millipore Corp. Acetate film, as used in overhead transparencies. Absorbent paper Whatman # 42. Non-toxic glue stick, available commercially from Conros Corp., Detroit, MI Ross 5tik # 06-074.
PBS ( 1 OmM) standard solution may be prepared according to Harlow, E. and D. Lane, Antibodies: a laboratory manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY at 726 ( 1988). Antibody stock solution ( 1 mg/mL) may be prepared by rehydration with 50% glycerol solution, according to antibody manufacturer's instructions. SPB may be prepared as indicated in "Protocols for Adsorbing Proteins one Polystyrene-Based Microspheres," by Bangs Laboratories, Inc., 9025 Technology Drive, Fishers, IN 46038-2886, telephone (317) 570-7020, Fax (317) 570-7034. Surfactants may be removed from microspheres as indicated in "How to Work with Microspheres," and "Protocols for Adsorbing Proteins onto Polystryrene-Based Microspheres," by Bangs Laboratories, Inc. The immunobead solution may be prepared following the Sodium Phosphate/PBS/BSA method indicated in "Protocols for Adsorbing Proteins onto Polystyrene-Based Micrpspheres,"
by Bangs Laboratories, Inc. For example, a 6 mL final volume of immunobead solution may be prepared with 120 ~,L antibody solution, 300 p,L black latex microspheres, and 5,580 ~L SPB.
Preparation of Immunobead Pad An immunobead pad for use in the present invention may be prepared as follows. Alcohol is evaporated from the Baxter alcohol swab (pad). The pad is then rinsed by repeated immersion in dd water. The step may then be repeated in a new amount of dd water, and the pad dried in an oven at 37 °C. The dried pad is placed on a glass surface and wetted with 1 % BSA using a pipettor, and then dried by suspending it from a corner inside an oven at 37 °C. The dried pad is placed on a glass surface and wetted with 50% sucrose using a pipettor, and then dried in an oven at 37 °C. The dried pad is placed on a glass surface and wetted with ANI~h~DED SHEET
immunobead solution using a pipettor. About 15 p,L of immunobead solution is applied to a 0.9 cm x 0.9 cm section of blocked pad. The pad is again dried in an oven at 37 °C for about 10 minutes. The drying process is completed at 4 °C in a refrigerator. The immunobead pads may be stored between sheets of non-adhesive paper tissue in plastic bags at 4 °C, and may be cut into appropriately sized pieces before contamination detection system assembly.
Preparation of Detection Area in the Substrate Immobilon-P membrane (Millipore Corp.), is pre-wetted as provided with the manufacturer's instructions. While the Immobilon-P membrane remains wet (opaque), primary antibody is applied with a 10-~L pipettor (about 3 ~.L of anti-E.
coli polyclonal antibody per membrane) in the shape desired for the detection area.
For example, in the case of a bar code a vertical bar (0.7 cm x 0.2 cm) is applied.
The antibody solution preferably has a concentration of 20 p,g/mL, PBS being used as the diluting solution. If the membrane partially dries (i.e. the membrane shows traces of white coloration), the pre-wetting procedure should be repeated. The applied antibody-is-dried-by-incubatingthe-membrane-overa-KimwipeTM tissue in an oven at 37 °C for IO-15 minutes, or in a refrigerator at 4 °C
for a longer period.
The membrane is then blocked by immersing the membrane in 1 % BSA
blocking solution for 30 minutes at room temperature. It is desirable that bubbles be prevented from forming on the surface of the membrane during the immersion procedure. The membrane is then three times rinsed by immersions in PBS, each immersion lasting about five minutes. Excess PB5 is eliminated by vertically placing the membrane on KimwipeTM tissue for about two minutes. The membrane is then dried on KimwipeTM tissue in an oven at 37 °C for 15 minutes or in the refrigerator at 4 °C for a longer period. Once dry, the blocked membrane is ready for assembly into the contamination detection system.
Assembly of the Contamination Detection System According to Preferred Embodiments A contamination detection system may be assembled by cutting an 5-PIA
plastic strip into two 4.5 cm x 1 cm pieces, one of them with the attached membrane ~~C7ED SHEET
with antibody in detection area 26. Using an additional piece of S-PIA plastic strip (3 cm x 1 cm) as a basis, the two 4.5 cm x 1 cm pieces are glued end to end so that the strip returns to its original dimensions (9 cm x 1 cm), but with the membrane on the central part of the plastic strip. A thin line of non-toxic glue is applied along both edges of the strip, except on the membrane. A 0.9 cm x 0.9 cm piece of immunobead pad is attached to the strip so that one edge is flush against the membrane. A 1.5 cm x 1 cm piece of absorbent paper is attached to the plastic strip ~ so that it is flush against the free end of the immunobead pad, while a second 1.5 cm x 1 cm piece of absorbent paper is attached to the plastic strip so that it is flush against the free end of the membrane. At this point the plastic strip should have four components glued side by side in the following sequence: paper-membrane-pad-paper.
Glue may then be applied in a thin line against both edges of a 2 cm x 0.9 cm piece of acetate. The acetate is glued so as to cover both the membrane and the pad. The acetate may slightly overlap on the piece of absorbent paper next to the membrane or pad.
A bar code may be incorporated into the assembled contamination detection system by attaching a second piece of acetate (7 cm x 3 cm) with a printed Uniform Product Code bar code to one end of the plastic strip such that the bars to be modified, or made unreadable, match the area where the antibody was applied (detection area 26) on the underlying membrane (substrate 8). The contamination detection system may be stored under refrigeration.
Thus, a food contamination detector providing for the directional capillary flow of juices has been disclosed. While embodiments and applications of this invention have been shown and described, it would be apparent to those skilled in the art that many more modifications are possible without departing from the inventive concepts herein. For example, while the example describes the use antibodies against E. coli, those of skill in the art will recognize that different antibodies, and antibodies to different antigen or antigens may be used.
Moreover, mixtures of antibodies, against a number of different antigens, or combinations -~~I~GEp ~i-~~t ~
thereof may be used. With the use of different antibodies, multiple different antigens which could be present in the sample can be detected. Those of skill in the art will also recognize other manners of assembling food contamination detectors within the teachings of the present disclosure. For example, the absorbent pad can be made of any material providing for the directional capillary flow of juices or fluids, the immunobead pad may be prepared using other materials and membranes well known in the art, as can the detection area. The invention, therefore is not to be restricted except in the spirit of the appended claims.
Gtd~~~, ~~.~~6__~
Claims (16)
1. A food contamination detector for use with a food container adapted for containing a food product which produces juices comprising a first layer comprising an impermeable support (12);
a second layer supported by the impermeable support (12) and comprising a capillary material (20) having in sequence a) a first absorbent material (22), wherein a sample may be introduced into the detector;
b) an immunobead solution pad (24) comprising a first set of antibodies linked to a light absorbing material and specific to an antigen to be detected;
c) a membrane (8) having a detection area (26), the detection area (26) comprising a second set of antibodies specific to at least one composition selected from the group consisting of the antigen to be detected, the first set of antibodies and the antigen to be detected bound to the first set of antibodies, said second set of antibodies being bound to the membrane (8) in the detection area (26); and a third layer comprising an impermeable cover film (2) covering the second layer (20), such that the second layer rests between the first layer (12) and the third layer (2);
wherein the juices flow directionally from the first absorbent material (22) through the immunobead solution pad (24) and into the membrane (8).
a second layer supported by the impermeable support (12) and comprising a capillary material (20) having in sequence a) a first absorbent material (22), wherein a sample may be introduced into the detector;
b) an immunobead solution pad (24) comprising a first set of antibodies linked to a light absorbing material and specific to an antigen to be detected;
c) a membrane (8) having a detection area (26), the detection area (26) comprising a second set of antibodies specific to at least one composition selected from the group consisting of the antigen to be detected, the first set of antibodies and the antigen to be detected bound to the first set of antibodies, said second set of antibodies being bound to the membrane (8) in the detection area (26); and a third layer comprising an impermeable cover film (2) covering the second layer (20), such that the second layer rests between the first layer (12) and the third layer (2);
wherein the juices flow directionally from the first absorbent material (22) through the immunobead solution pad (24) and into the membrane (8).
2. The food contamination detector of claim 1, wherein the impermeable cover film (2) has a window (6) aligned with the detection area (26).
3. The food contamination detector of claim 2, the cover film (2) having a code indicia.
4. The food contamination detector of claim 3, the code indicia being at least one bar code.
5. The food contamination detector of claim 3, the code indicia being at least two bar codes.
6. The food contamination detector of claim 4, the presence of antigen obliterating at least one bar code such that it is not readable by a scanner.
7. The food contamination detector of claim 4, the presence of antigen creating at least one bar in a bar code, such that it represents a different code to the scanner.
8. The food contamination detector of claim 1, the second layer (20) comprising an inflow orifice (14) for transporting juices and other fluids from the food container to a protective filter (16), the protective filter being set between the inflow orifice (14) and a selective filter (18), the selective filter being set between the protective filter (16) and the first absorbent material (22).
9. The food contamination detector of claim 1, the second layer further comprising d) a second absorbent material (28), abutting and in contact with the membrane (8).
10. The food contamination detector of claim 9, the second absorbent material (28) serving as a filter which prevents microspheres from leaving the membrane (8) area.
11. The food contamination detector of claim 1, the membrane (8) having a channeled surface.
12. The food contamination detector of claim 1, further comprising an inflow orifice (14) for transporting juices and other fluids from a food container to the first absorbent material (22).
13. The food contamination detector of claim 12, further comprising a filter (16) set between the inflow orifice (14) and the first absorbent material (22).
14. The food contamination detector of claim 1, the light absorbing material linked to the first set of antibodies comprising latex microspheres.
15. The food contamination detector of claim 1, the food contamination detector being embedded in the food package.
16. A method for the detection of food contamination for use with a food container adapted for containing a food product which produces juices comprising drawing the juices sequentially through (a) a first absorbent material (22);
(b) an immunobead solution pad (24), comprising a first set of antibodies specific to an antigen to be detected, the first set of antibodies being linked to a light absorbing material; and (c) a membrane (8);
wherein a second set of antibodies specific to at least one compound selected from the group consisting of the antigen to be detected, the first set of antibodies, and the antigen to be detected bound to the first set of antibodies is bound to the membrane in a detection area (26).
(b) an immunobead solution pad (24), comprising a first set of antibodies specific to an antigen to be detected, the first set of antibodies being linked to a light absorbing material; and (c) a membrane (8);
wherein a second set of antibodies specific to at least one compound selected from the group consisting of the antigen to be detected, the first set of antibodies, and the antigen to be detected bound to the first set of antibodies is bound to the membrane in a detection area (26).
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US5887397P | 1997-09-15 | 1997-09-15 | |
| US60/058,873 | 1997-09-15 | ||
| PCT/US1998/019167 WO1999014598A1 (en) | 1997-09-15 | 1998-09-15 | Food sentinel system ?tm¿ |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA2303384A1 true CA2303384A1 (en) | 1999-03-25 |
Family
ID=22019438
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA002303384A Abandoned CA2303384A1 (en) | 1997-09-15 | 1998-09-15 | Food contamination detection system |
Country Status (7)
| Country | Link |
|---|---|
| EP (1) | EP1018013A1 (en) |
| JP (1) | JP2001516883A (en) |
| CN (1) | CN1278920A (en) |
| AU (1) | AU9483998A (en) |
| CA (1) | CA2303384A1 (en) |
| NO (1) | NO20001297L (en) |
| WO (1) | WO1999014598A1 (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP4109101A1 (en) * | 2021-06-24 | 2022-12-28 | Technische Universität München | Portable testing device for testing susceptibility of a biological probe to one or more bioactive substances |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1119771B1 (en) * | 1998-10-06 | 2003-07-02 | Verseau Inc | Toxin detector |
| DE102004052729A1 (en) * | 2004-10-30 | 2006-05-04 | Roche Diagnostics Gmbh | Immunocomplex-specific antibody for zero-reduction in the detection of antigen-specifically bound antibodies of a specific immunoglobulin class in array test formats |
| GB2427271A (en) * | 2005-06-16 | 2006-12-20 | Porvair Filtration Group Ltd | Diagnostic device |
| WO2013115478A1 (en) * | 2012-02-03 | 2013-08-08 | 동국대학교 산학협력단 | Food wrapping kit for detecting bacteria |
Family Cites Families (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4981786A (en) * | 1987-09-04 | 1991-01-01 | Syntex (U.S.A.) Inc. | Multiple port assay device |
| US5958339A (en) * | 1992-08-31 | 1999-09-28 | Clinical Diagnostic Systems, Inc. | Format for immunoassay in thin film |
| US5306466A (en) * | 1993-05-19 | 1994-04-26 | California South Pacific Investors | Detection of contaminants in food |
| AU686166B2 (en) * | 1993-05-19 | 1998-02-05 | Sira Technologies, Inc. | Detection of contaminants in food |
| NZ505583A (en) * | 1996-09-30 | 2002-10-25 | California South Pacific Investors | Indicator colour change alters appearance of product and contamination labels of food package |
-
1998
- 1998-09-15 WO PCT/US1998/019167 patent/WO1999014598A1/en not_active Ceased
- 1998-09-15 CA CA002303384A patent/CA2303384A1/en not_active Abandoned
- 1998-09-15 JP JP2000512083A patent/JP2001516883A/en active Pending
- 1998-09-15 EP EP98948221A patent/EP1018013A1/en not_active Withdrawn
- 1998-09-15 CN CN98811173A patent/CN1278920A/en active Pending
- 1998-09-15 AU AU94839/98A patent/AU9483998A/en not_active Abandoned
-
2000
- 2000-03-13 NO NO20001297A patent/NO20001297L/en not_active Application Discontinuation
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP4109101A1 (en) * | 2021-06-24 | 2022-12-28 | Technische Universität München | Portable testing device for testing susceptibility of a biological probe to one or more bioactive substances |
| WO2022268827A1 (en) | 2021-06-24 | 2022-12-29 | Technische Universität München | Portable testing device for testing susceptibility of a biological probe to one or more bioactive substances |
Also Published As
| Publication number | Publication date |
|---|---|
| NO20001297D0 (en) | 2000-03-13 |
| JP2001516883A (en) | 2001-10-02 |
| EP1018013A1 (en) | 2000-07-12 |
| AU9483998A (en) | 1999-04-05 |
| NO20001297L (en) | 2000-05-15 |
| WO1999014598A1 (en) | 1999-03-25 |
| CN1278920A (en) | 2001-01-03 |
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