GB2196117A

GB2196117A - Microbiological assay device

Info

Publication number: GB2196117A
Application number: GB8718580A
Authority: GB
Inventors: Ian Hartley Lewis; Raymond Curry
Original assignee: CURRY Ltd R; VENTURECARE Ltd
Current assignee: CURRY Ltd R; VENTURECARE Ltd
Priority date: 1986-08-07
Filing date: 1987-08-06
Publication date: 1988-04-20
Anticipated expiration: 2007-08-06
Also published as: GB8718580D0; GB8619311D0; GB2196117B

Abstract

A device for detecting the presence of and determining the extent of biological contamination of a sample is disclosed. A sample vessel 14 is brought into sealed engagement with the device 10, and a liquid reagent, liquid hydrogen peroxide, is added to the sample during operation of the device. Catalayse contained in the contaminated sample breaks down the hydrogen peroxide releasing gaseous oxygen, the increase in gaseous pressure being detected by a pressure sensor 23. <IMAGE>

Description

SPECIFICATION Microbiological assay device This invention relates to a microbiological assay device for detecting and for measuring the extent of biological contamination of a sample.

The invention has been developed primarily, though not excusively, with a view to provide an assay device which can be used to detect the presence and the extent of biological contamination of a food or drink sample, and in which the device is readily operable and is able to provide an immediate record of the presence and/or extent of biological contamination of the sample.

In the food industry, it is very important for health reasons to be able to monitor the state of hygiene of food products at regular intervals, but existing techniques are very timeconsuming. These either involve purely manual operations, in a series of separate operations, which are therefore time-consuming and an inefficient use of manpower. There are also, however, available expensive monitoring equipment which can provide automatic readings, but these are sophisticated pieces of equipment which are, necessarily expensive, and which also take a long time before a final reading is available.Thus, this equipment depends upon the variation in electrical properties of a sample, in that the electrical impedance of micro-organisms developing on a sample causes variation in the electrical properties of the sample, as a function of the extent of the micro-organisms present, but it can take from 4 to 6 hours before a final reading becomes available.

Accordingly, there is a clear need to provide a degree of automacitity in the detection and /or measurement of biological contamination, and by means of an assay device which is relatively inexpensive, as compared with the existing monitoring equipment, and which can provide relatively rapid results.

According to the present invention there is provided a microbiological assay device for detecting and for measuring the extent of biological contamination of a sample, in which the device comprises a pressure transducer which is, in use, in sealed communication with a sample vessel, and an inlet through which a liquid reagent can be brought into contact with a sample in the sample vessel, the reagent being of the type which reacts with a contaminated sample to liberate gas and the pressure transducer being responsive to the action of the gaseous presure generated by said liberated gas, and an electronic monitoring device connected to the pressure transducer and operable to indicate the presence and/or the extent of contamination of the sample as a function of the said gaseous pressure.

The device may include a portable housing which defines a sealable reaction chamber, the housing having a first port therein in which a sample vessel may be detachably mounted in a substantially sealed manner, and a second port in which a reagent vessel can be detachably mounted in substantially sealed manner, the vessel being intended to contain the liquid reagent.

It is preferred that the housing is pivotally mounted on a siutable support stand for movement about a substantially horizontal pivot axis, and the second port is arranged at a higher level than the first port. Thus, upon loading of the device with the sample vessel and the reagent vessel (when the housing will be in a loading position about its pivot axis), the housing can then be pivoted to an operative (mixing) position in which the movement of the second port from the loading position is such that the liquid reagent in the reagent vessel can drain from the vessel and into the sample vessel. Usually, the sample vessel will be located substantially at the lowest part of the reaction chamber when the housing is in the operative position.

Conveniently, the first and second ports may be angularly spaced from each other by about 90 about the pivot axis, and the second port will be below the pivot axis when in the loading position and above the pivot axis when in the operative position. By this arrangement of the ports, it is ensured that the sample always remains in the sample vessel, and therefore this device does not involve any appreciable risk of contamination of the walls of the reaction chamber. This is a significant advantage as compared with known techniques, in which the sample comes into contact with working surfaces of any apparatus which is used, and therefore lengthy sterilisation of these working surfaces is required e.g.

by autoclaving between each sample testing.

The sample vessel and the reagent vessel may take the form of open ended tubes, and the arrangement of the respective ports is such that the tubes can be readily detachably mounted therein in sealed manner. Bayonettype or other suitable releasable engagements may be provided.

One disadvantage of this particular embodiment is that the pivoting of the device causes turbulance which can lead to erratic results.

This is alleviated by an alternative embodiment in which the reagent is added directly to the sample vessel from a position vertically above the sample vessel, via a valve.

The device also preferably incorporates a temperature sensor for monitoring the temperature during the reaction, so that the pressure may be indicated as a function of temperature as well as time.

According to another aspect of the invention there is provided a method of detecting and measuring the extent of biological contamination of a sample using a microbiological assay device, in which a sample is introduced into the device, liquid reagent is brought into contact with the sample so as to liberate gas upon contact with any contaminant which may be present on the sample, the increase in ga seous pressure within the device is detected by a pressure transducer, and the presence and the extent of contamination of the sample is indicated by electronically monitoring electri cal signals derived from the pressure trans ducer.

The invention is primarily based upon a re cognition of the fact that food and drink which becomes biologically contaminated de velops and enzyme catalayse, and that the ap plication of liquid hydrogen peroxide to catala yse causes the liberation of gaseous oxygen in an amount which is dependent upon the extent of catalayse (and hence contamination) which is present in a sample. Accordingly, in use of the device, the preferred liquid reagent is liquid hydrogen peroxide.

Two embodiments of microbiological assay device according to the invention will now be described in detail, by way of example only, with reference to the accompanying drawings in which: Figure 1 is a schematic drawing of one em bodiment of the present invention, Figure 2 is a front elevation of an alternative embodiment of the invention, Figure 2A is a plan view of the device shown in Fig. 2, Figure 2B is a view on line A-A of Fig. 2, Figure 3 is a side elevation of the device shown in Fig. 2, Figure 3A is a cross section through line B-B of Fig. 2, Figure 3B is a close up of the lever arm shown in Fig. 3A.

Referring to Fig. 1, the microbiological assay device is designated generally by reference 10, and is intended to detect and to measure the extent of biological contamination of a food or drink, or other sample. The device 10 comprises a portable housing 11 which de fines a sealable reaction chamber 12. A first port 13 is formed in the housing 11 and in which a sample vessel in the form of a sam ple tube 14 can be detachably mounted in substantially sealed manner by any convenient means, such as a bayonet-type coupling. A second port 15 is formed in the wall of the housing 11 and, as will be seen from the drawing, at a higher level than the first port 13.The second port 15 serves to detachably mount a reagent vessel in the form of a reag ent tube 16, this tube being also capable of being mounted in substantially sealed manner and being intended to contain a liquid reagent of the type which reacts with a contaminated sample in order to liberate gas.

A pressure transducer 17 is arranged to be exposed to the action of the gaseous pressure generated in the reaction chamber 12 when a sample in the tube 14 is brought into contact with the liquid reagent from the reagent tube 16. An electronic monitoring device is connected to the transducer 17 and is operable to indicate the presence and/or the extent of contamination of a sample as a function of the gaseous pressure generated in the reaction chamber. The electronic monitoring device comprises an amplifier 18 which is connected to the transducer 17 in order to amplify electrically the signal from the transducer, a computer 19, a printer 20 and a voltage regulator 21 connected to a 13 amp mains supply.

The housing 11 is pivotally mounted on a suitable support stand (not shown) for movement about a substantially horizontal pivot axis, which coincides with the general longitudinal extent of the transducer 17. The housing 11 is shown in the drawing in a loading position in which the sample tube 14 and the reagent tube 16 can be mounted in the respective ports, in sealed manner, and after charging of each tube respectively with a sample and with a suitable liquid reagent, which is preferably liquid hydrogen peroxide. Following loading of the device 10, the housing 11 is then pivotable to an operative (mixing) position in which the sample tube 14 is brought to a lower most part of the reaction chamber 12, in that it is brought substantially in line with the vertical axis 22.During this pivotal movement, the second port 15 moves from its loading position which is below the pivot axis, to a position above the pivot axis, whereby the liquid reagent can drain from the tube 16, along the adjacent lower wall of the reaction chamber, and into the sample tube 14.

It will be noted that the angular spacing apart of the tubes 14 and 16, about the pivot axis, is approximately 90 , and the tubes are open ended which readily permit contact to be achieved between the liquid reagent and the sample after the reaction chamber 12 has been sealed and loaded with the sample and the liquid reagent. However, other means may be provided (not shown) to permit communication to be achieved between the liquid reagent and the sample only after loading and sealing of the reaction chamber, By the arrangement of the ports 13 and 15, it is ensured that the sample always remains in the sample tube 14, and therefore the device 10 does not involve any appreciable risk of contamination of the walls of the reaction chamber 12. Furthermore, liquid hydrogen peroxide is known for its sterilizing properties, so that it maintains the walls of the reaction chamber 12 in a sterile condition when it drains downwardly into the sample tube 14.

In addition to the pressure transducer 17 for monitoring the instantaneous gaseous pressure generated in the chamber 12, a coarse pressure check device is provided in the form of a psi meter 23 which can also be brought into communication with the pressure prevailing with the reaction chamber 12. In the communication line between the meter 23 and the reaction chamber 12, there is also arranged a vernier pump 24. This coarse pressure check arrangement can also be used in testing final results and consistency of the results provided by the pressure transducer 17.

The sample tube 14 conveniently is supplied as a sterile disposable tube, in which the sample to be tested is housed, and it will usually be necessary to hydrate solid samples in the tube, using distilled water, and a preferred range of hydration would be 1 to 14 grammes of sample, dependent on the contamination level, together with 10 to 30 ml of distilled water. The reagent tube 16 is charged with liquid hydrogen peroxide (100 vol), and is mixed with a little tartrazine so that complete or incomplete mixing can be noted.As indicated above, the transducer 17 is located on the pivot axis of the housing, and has a transducer range - 1.0 to + 1.0 psi, and its arrangement is such as to minimise gravitational interference when the housing 11 is rotated from the loading position, as shown, to the operative or mixing position, in which the tube 14 is in line with the vertical axis 22.

The programming of the computer 19 is such that there is continuous monitoring of pressure signals derived from the transducer 17, this sampling being carried out over 24 separate 10 second intervals, and during each 10 second interval there will be averaging of approximately 1,000 pulses. A final reading will normally be achievable therefore within about 2 minutes, which is very substantially quicker than existing more sophisticated, or more expensive equipment.

If it should be desired to be able to carry out multiple testing, a plurality of reaction chambers 12 may be arranged in a bank, each being intended to carry out monitoring of a respective sample.

An alternative embodiment of the invention which does not require to be pivoted during its operation is shown in Figs. 2 and 3. Instead of the reaction chamber 11 there is provided a two-valve head 26, and the sample vessel 28 is brought into sealing engagement with the base 27 of the valve head by means of a vessel holder 30. By pivoting of the lever arm 32 as shown in Figs. 3A and 3B the sample vessel 28 is raised and pressed against sealing ring 31. The vessel is then in fluid communication with a pressure sensor 4, a reagent inlet valve 2 and vent valve 1.

Once the sample vessel has been pressed against the valve head 26, the system is vented by opening valve 1. The valves 1 and 2 are then closed by computer-controlled solenoids. A measured amount of hydrogen peroxide is then admitted by opening valve 2 and at the same time vent valve 1 is opened to vent the system and facilitate clean entry of the hydrogen peroxide into the sample vessel.

The valves 1 and 2 are then closed and the pressure sensor 4 and associated monitoring device monitors the pressure increase and computes the rate of change of pressure with respect to both time and temperature. For this purpose a temperature sensor 3 is built into the device.

There will now be discussed the chemical reactions upon which the apparatus and the method are based.

In the presence of the enzyme catalayse, liquid hydrogen peroxide readily and rapidly releases oxygen, the catalyse acting as a specific and powerful catalyst.

2H202-12H20 + 02 Most bacteria, molds and yeasts contain the enzyme catalayse. Thus, the catalayse content of bacteria, for example, is reported to be of the order of from 1 to 2% of the total protein content, which would be a quantity equivalent to about 103 to 104 catalayse molecules per organism. Given that one molecule of catalayse is capable (under optimum conditions) of braking down 106 molecules of hydrogen peroxide, a high degree of sensitivity, even to low levels of contamination should be possible.

Possible uses of the device include the following: 1. Food manufacturers for testing their products.

2. Milk producers testing output samples, since payment is now based upon contamination levels.

3. The Milk Marketing Board, who will now be able to test quickly on receipt of deliveries.

4. Medical-testing of urine samples-as a first time rapid test, to establish contamination levels either from bacteria, or bacteria and cell debris-filtration of samples could isolate the causes of contamination.

5. Soil sterility testing for use in horticulture.

6. Air contamination levels, in which case a vacuum pump drawing measured volumes of air through filter paper would be required, and the test conducted on the resultant filter paper.

7. Detection of bateriological warfare.

Claims

1. A microbiological assay device for detecting and for measuring the extent of biological contamination of a sample, in which the device comprises a pressure trasducer which is, in use, in sealed communication with a sample vessel, and an inlet through which a liquid reagent can be brought into contact with a sample in the sample vessel, the reagent being of the type which reacts with a contaminated sample to liberate gas and the pressure transducer being responsive to the action of the gaseous pressure generated by said liberated gas, and an electronic monitoring device connected to the pressure transducer and operable to indicate the presence and/or the extent of contamination of the sample as a function of the said gaseous pressure.

2. A device according to Claim 1 and including a portable housing which defines a sealable reaction chamber, the housing having a first port therein in which a sample vessel may be detachably mounted in a substantially sealed manner, and a second port in which a reagent vessel can be detachably mounted in substantially sealed manner, the vessel being intended to contain the liquid reagent.

3. A device according to Claim 2 in which the housing is pivotally mounted on a suitable support stand for movement about a substantially horizontal pivot axis, and the second port is arranged at a higher level than the first port.

4. A device according to Claim 2 or Claim 3 in which the first and second ports are angularly spaced from each other by about 90" about the pivot axis, and the second port is below the pivot axis when in the loading position and above the pivot axis when in the operative position.

5. A device according to any of Claims 2 to 4 in which the sample vessel and the reagent vessel take the form of open ended tubes, and the arrangement of the respective ports is such that the tubes can be readily detachably mounted therein in sealed manner.

6. A device according to Claim 1 in which the reagent is admitted through a valve located directly above the sample vessel.

7. A method of detecting and measuring the extent of biological contamination of a sample using a microbiological assay device, in which a sample is introduced into the device, liquid reagent is brought into contact with the sample so as to liberate gas upon contact with any contaminant which may be present on the sample, the increase in gaseous pressure within the device is detected by a pressure transducer, and the presence and the extent of contamination of the sample is indicated by electronically monitoring electrical signals derived from the pressure transducer.

8. A method according to Claim 7 in which the reagent is liquid hydrogen peroxide.

9. A microbiological assay device substantially as herein described with reference to the accompanying drawings.

10. A device according to Claim 1 and substantially as herein described.

11. A method of detecting and measuring the extent of biological contamination of a sample, the method being substantially as herein described with reference to the accompanying drawigs.

12. A method according to Claim 7 and substantially as herein described.