A SYSTEM AND DEVICE

Abstract

A cassette for use in concentrating matter in a sample suspension, the cassette comprising a housing having a support and an enclosed sample-receiving channel, the enclosed sample-receiving channel having an upper portion and a base connected by at least two walls; in which the upper portion is configured to have a width less than a width of the base and a depth greater than 400 m.

Claims

1. A cassette (1,200,300) for use in concentrating matter in a sample suspension, the cassette (1,200,300) comprising a housing (2) having a support (5) and an enclosed sample-receiving channel (6), the enclosed sample-receiving channel (6) having an upper portion (7,208) and a base (5a) connected by at least two walls (11a, 11b); in which the upper portion (7,208) is configured to have a width less than a width of the base (5a) and a depth greater than 400.

2. A cassette (1,200,300) according to claim 1, in which the walls (11a, 11b) of the enclosed sample-receiving channel (6) have a slope of between 40 and 90 with respect to the base (5a).

3. A cassette (1,200,300) according to claim 1, in which the base (5a) is formed by the housing support (5).

4. A cassette (1,200,300) according to claim 1, in which the enclosed sample-receiving channel (6) further comprises a raised platform (50) extending upwards from the base (5a).

5. A cassette (1,200,300) according to claim 4, in which the raised platform (50) is separated from the upper portion (7,208) and at least one of the walls (11a, 11b).

6. A cassette (1,200,300) according to claim 5, in which the raised platform (50) is separated from the upper portion (7,208) and both of the walls (11a,11b).

7. A cassette (1,200,300) according to claim 1, in which the enclosed sample-receiving channel (6) can have any shape selected from the group comprising straight, curved, circular, partial circle, elliptical, a connected series of two or more straight or curved lines, or a combination thereof.

8. A cassette (1,200, 300) according to claim 1, in which the enclosed sample-receiving channel (6) in cross-section is a trapezoid, an isosceles trapezoid, a truncated triangle, a rectangle, an isosceles trapezoid on a rectangle, an ellipse, an arc, is concave or convex.

9. A cassette (1,200,300) according to claim 1, in which the housing (2) further comprises an air release port (14) in fluid communication with the enclosed sample-receiving channel (6).

10. A cassette (1,200,300) according to claim 1, in which the cassette further comprises an inlet port (20) adapted to receive the sample suspension and act as a conduit to deliver the sample to the enclosed sample-receiving channel (6).

11. A cassette (1,200,300) according to claim 10, in which the air release port (14) is diametrically opposed the inlet port (20) or the air release port (14) is opposite the inlet port (20).

12. A cassette (1,200,300) according to claim 11, in which the inlet port (20) is a non-return valve.

13. A cassette (200) according to claim 1, in which the housing (2) of the cassette (1) consists of a lower section (201) configured to accommodate the enclosed sample-receiving channel (6) and an upper section (202) configured to reversibly attach to and create a seal with the lower section (201).

14. A cassette (200) according to claim 13, in which the enclosed sample-receiving channel (6) is formed when the lower section (201) and the upper section (202) are combined and form a closed, sealed housing (2).

15. A cassette (1,200,300) according to claim 1, wherein the housing further comprises a hole (30) configured to receive an actuator (101) connected to an image capturing device (100).

16. A cassette (1,200,300) according to claim 1, in which the cassette (1,200,300) is configured to move in one plane.

17. A cassette (1,200,300) according to claim 16, wherein the motion within the plane can be rectilinear-translation or rotation, or a combination thereof.

18. A cassette (1,200,300) according to claim 1, in which an internal surface of the walls (11a,11b) of the enclosed sample-receiving channel (6) is coated with a hydrophobic material or a hydrophilic material.

19. A cassette (1,200,300) according to claim 1, in which the housing (2) is transparent or translucent.

20. A cassette (1,200,300) according to claim 1, in which the housing (2) of the cassette (1) is either open or closed.

21. A cassette (1) according to claim 1, in which the cassette (1) is a single piece.

22. A cassette (1,200,300) according to claim 1, in which the cassette is disposable or suitable for re-use.

23. A cassette (1,200,300) according to claim 1, in which the matter is suspended matter or particulate matter.

24. A system (400) for concentrating particulate matter in a sample suspension, the system comprising the cassette (1,200,300) as claimed in claim 1 and an image capturing device (100) configured to accommodate the cassette (1,200,300).

25. A system (400) according to claim 24, in which the system further comprises a computer, an internal storage device, a detector (122) and wireless network to send data for cloud-computing storage.

26. A kit for determining the presence of particulate matter in a liquid sample, the kit comprising a cassette (1,200,300) as claimed in claim 1 and a flotation solution.

27. A kit according to claim 26, in which the flotation solution is selected from a saturated NaCl solution with specific gravity of 1.20, a saturated sugar solution with a specific gravity of 1.280, a Sheather's sugar solution with a specific gravity of 1.20, a saturated zinc sulphate solution with a specific gravity of 1.20, a saturated sodium nitrate solution with a specific gravity of 1.20, and a saturated magnesium sulphate solution with a specific gravity of 1.280.

28. A method for determining the presence of particulate matter in a sample suspension, the method comprising the steps of: preparing the sample suspension; mixing the sample suspension thoroughly in a container; introducing the thoroughly mixed sample suspension into the closed sample-receiving channel (6) of the cassette (1,200,300) as claimed in claim 1; mounting the cassette (1,200,300) with the mixed sample suspension onto an image capturing device (100) having an objective lens (102); and moving the cassette (1,200,300) or optical image device (100) in one plane to allow the determination of the presence of particulate matter in the sample suspension, wherein the field of view of the objective lens (102) corresponds to the width of an upper portion (7,208) of the enclosed sample-receiving channel (6).

29. The method of claim 28, in which the image capturing device (100) comprises an actuator (101) configured to engage with the cassette (1,200,300) and move either the objective lens (102) or the cassette (1,200,300) in one plane only.

30. A cassette (1,200,300) for use in determining the presence of matter in a sample suspension, the cassette (1,200,300) comprising a housing (2) having a support (5); an inlet channel (20) adapted to receive the sample suspension; and a closed sample-receiving channel (6) in fluid communication with the inlet channel (20), the closed sample-receiving channel (6) having an upper portion (7,208) and a base (5a) connected by at least two walls (11a,11b) forming a long axis; in which the upper portion (7,208) is configured to have a width equal to or less than a width of the base (5a).

31. A cassette (1,200,300) for use in concentrating (particulate) matter in a sample suspension, the cassette (1,200,300) comprising a housing (2) having a support (5) and an enclosed sample-receiving channel (6), the enclosed sample-receiving channel (6) having an upper portion (7,208) and a base (5a) connected by at least two walls (11a, 11b); in which the upper portion (7,208) is configured to have a width less than a width of the base (5a).

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0106] The invention will be more clearly understood from the following description of an embodiment thereof, given by way of example only, with reference to the accompanying drawings, in which:

[0107] FIG. 1A and FIG. 1B illustrate perspective views of an example of a cassette of the invention, where the cassette is disk-shaped, with and without a cover, respectively. The cassette of FIG. 1B can be determined as being with and without a separate cover, where the overall cassette is transparent and where the cover is also transparent, thus giving the same internal view.

[0108] FIG. 2A illustrates a cross-sectional view of the cassette of FIG. 1, while FIG. 2B illustrates a cross-sectional view of the enclosed sample-receiving channel of FIG. 1 having a platform.

[0109] FIG. 3 illustrates a cross-sectional view of one embodiment of a cassette of the invention where the cassette is a single piece and the enclosed sample-receiving channel further includes a platform.

[0110] FIG. 4 illustrates an example of one embodiment of a cassette of the invention, where the cassette is a quadrilateral or linear in shape, without a platform.

[0111] FIGS. 5A and 5B illustrate a typical image capturing device for use with the cassette illustrated in FIGS. 1 to 4.

[0112] FIG. 6 illustrates a typical data collection system used in conjunction with the determination system of the invention.

[0113] FIG. 7A-C illustrate a typical photomicrograph of one FOV of an ovine faecal sample prepared as outlined below using the cassette described herein and the system as shown in FIG. 5A. FIG. 7A illustrates an oocyst (circled) from Nematodirus and the characteristics of the interior of the oocyst are clearly visible. FIG. 7B shows that air bubbles (circled) are easily identified. FIG. 7C shows that fainter oocysts (circled) represent Coccidia.

[0114] FIG. 8 illustrates a side, cross-sectional view of an example of a cassette of the invention.

[0115] FIG. 9 illustrates a typical photomicrograph of one FOV of eggs from ovine (FIG. 9A, 9B), bovine (FIG. 9C, 9D) and equine (FIG. 9E) faecal samples visible (circled) in the channel of the cassette of the claimed invention and which was captured using the optical system described in the invention. The types of eggs are labelled as follows; N=Nematodirus; M=Moniezia; C=coccidia; S=Strongyles. The Moniezia and Strongyles eggs were not shown in FIG. 7.

DETAILED DESCRIPTION OF THE DRAWINGS

[0116] The present invention describes a system comprising an image capturing device with a magnifying lens, a cassette designed to hold a specific volume of a liquid sample, image analysis software designed to recognise and count specific items/particles of interest, and a kit to simplify the procedure for an end user.

[0117] Referring now to the figures, where FIG. 1, FIG. 2 and FIG. 8 illustrate one embodiment of a cassette of the present invention. Specifically, FIG. 1 illustrates a perspective view of one embodiment of a cassette of the present invention and is generally referred to by reference numeral 1. FIG. 2A and FIG. 8 illustrate the cassette 1 of FIG. 1 in cross-section. The cassette 1 comprises a housing 2 having an outer wall 3, a hole 30 configured to accommodate an actuator or motor drive shaft and a support 5. The housing 2 of the cassette 1 can be with or without an outer wall 3. The support 5 forms a base 5a of an enclosed sample-receiving channel 6, which is situated between the hole 30 and outer wall 3. The enclosed sample-receiving channel 6 comprises walls 11a,11b which form a conduit 12 adapted to accommodate a liquid sample. The housing 2 further comprises an inlet port 20 in fluid communication with the enclosed sample-receiving channel 6. The inlet port 20 is configured to receive the liquid sample and deliver the sample to the conduit 12 of the enclosed sample-receiving channel 6. In some embodiments, and opposite the inlet port 20, can be positioned a (small) air release port 14, which permits air to escape when the liquid sample is being applied to the enclosed sample-receiving channel 6 via the inlet port 20 (see FIG. 2A).

[0118] As shown in FIG. 1, FIG. 2 and FIG. 8, the enclosed sample-receiving channel 6 in cross-section shows the walls 11a,11b form a trapezoid shape with an upper portion 7 parallel to the support 5. The upper portion 7 is preferably transparent or translucent. The walls 11a,11b can be coated on the side facing the conduit 12 with a hydrophobic or a hydrophilic material. The conduit 12 concentrates buoyant matter in the liquid sample at the upper portion 7. The width of the upper portion 7 is determined by the field of view (FOV) of an image capturing device 100 (see FIG. 4). Ideally, the width of the upper portion 7 corresponds to the width of the FOV, so that the entire upper portion 7 can be successively scanned simply by either rotating the cassette 1 through 360 or rotating an imaging capturing device through 360, with no additional relative motion required. As shown in the FIG. 2B, the enclosed sample-receiving channel (optionally) further comprises a raised platform 50 extending upwards from the base 5a. The raised platform 50 is distinctly separate (FIG. 2B) from one or both of the walls 11a,11b and the upper portion 7. The advantage of having a raised platform 50 is that there is a smaller, shallower depth to the channel 6 directly under the viewing area but larger volumes in the reservoirs either side of the platform 50, which results in a smaller volume of liquid under the objective lens of the microscope or image reader. This smaller volume of liquid reduces the blurriness caused by diffraction of light through a liquid. The raised platform 50 remains free of, and never comes in to contact with, the walls 11a,11b and the upper portion 7.

[0119] Referring now to FIG. 3, there is illustrated a cross-section view of one embodiment of the cassette of the invention, and is given reference numeral 200, in which parts or steps described with reference to the previous embodiment are assigned the same numerals. The cassette 200 is a two-piece object consisting of a lower section 201 and an upper section 202. The lower section 201 comprises an outer wall 3; a hole 30 configured to accommodate an actuator or motor drive shaft; a support 5 with an enclosed sample-receiving channel 6 positioned thereon and between the outer wall 3 and the hole 30. The lower section 201 of the cassette 200 can be used with or without an outer wall 3. The enclosed sample-receiving channel 6 in cross-section shows the walls 11a,11b angled such that they form a trapezoid shape. The portion of the support 5 closed off by the walls 11a,11b forms the base 5a of the enclosed sample-receiving channel 6. The walls 11a,11b can be coated on the side facing the conduit 12 with a hydrophobic or hydrophilic material.

[0120] The upper section 202 has a top surface 206 and a bottom surface 207 and is preferably transparent or translucent. The upper section 202 further comprises a small air release port 14 in fluid communication with the closed sample-receiving channel 6. The small air release port 14 is configured to permit air to escape when the liquid sample is being applied to the enclosed sample-receiving channel 6 via the inlet port 20. The inlet port 20 is configured to receive the liquid sample and deliver the sample to the conduit 12 of the enclosed sample-receiving channel 6. The upper section 202 is configured to reversibly engage with the walls 11a,11b of the enclosed sample-receiving channel 6. The bottom surface 207 is configured to sit parallel to the support 5 and to rest on top of the walls 11a,11b of the enclosed sample-receiving channel 6, forming an upper portion 208 through which a lens of the image capturing device 100 can detect buoyant matter in the conduit 12. The engagement of the upper section 202 with the walls 11a,11b form an air-tight seal. The conduit 12 concentrates buoyant matter in the liquid sample where the upper section 202 forms the seal with the enclosed sample-receiving channel 6, that is, at the area of the upper portion 208.

[0121] Referring now to FIG. 4, there is illustrated another embodiment of a cassette, which is provided with reference numeral 300, and in which parts or steps described with reference to the previous embodiment are assigned the same numerals. The cassette 300 is linear or quadrilateral in shape. The cassette 300 comprises a housing 301 having an (optional) outer wall 3, a support 5 with an enclosed sample-receiving channel 6 positioned thereon. The enclosed sample-receiving channel 6 comprises walls 11a,11b which from form a conduit 12 adapted to accommodate a liquid sample. The enclosed sample-receiving channel 6 shown here has walls 11a,11b angled to form a trapezoid shape with an upper portion 7 parallel to the support 5. The portion of the support 5 closed off by the walls 11a,11b forms the base 5a of the enclosed sample-receiving channel 6. The upper portion 7 is preferably transparent or translucent. The walls 11a,11b can be coated on the side facing the conduit 12 with a hydrophobic or a hydrophilic material.

[0122] The conduit 12 concentrates buoyant matter in the liquid sample at the upper portion 7. The width of the upper portion 7 is determined by the FOV of an image capturing device 100 (see FIG. 3). Ideally, the width of the upper portion 7 corresponds to the width of the FOV, so that the entire upper portion 7 can be successively scanned simply by either moving the cassette 300 or the image capturing device 100 in one plane.

[0123] Referring now to FIG. 5, there is illustrated an image capturing device 100 suitable for use with the cassette 1,200,300 described above. In this figure, a typical compound microscope is illustrated as the image capturing device 100, with a stage 120, an objective lens 102 (4/NA0.1 plan), corrected for 160 mm focal distance, and a detector 122 (Raspberry Pi camera with 8 Mpi). The image capturing device 100 comprises an actuator 101 that rotates or moves the cassette 1,200,300 along a certain axis, the objective lens 102, a sensor (illustrated as the detector 122), one or more illumination source(s), a computer, a transmitting device, a suitable housing 124, and a power source.

[0124] The actuator 101 is an electric motor that rotates the cassette 1,200, or moves the cassette 300 along a plane, causing the enclosed sample-receiving channel 6 to pass under the objective lens 102 of the image capturing device 100. The hole 30 of cassette 1,200,300 is configured to accommodate the actuator 101, locking the actuator 101 and cassette 1,200,300 together so that they rotate/move together. The locking of the actuator 101 and the cassette 1,200,300 can be achieved by the typical systems known in the art for mounting a cassette or slide on a microscope stage. For example, the locking can be achieved by a magnetic force generated between a magnet 110 that is mounted on the actuator 101 and a magnetic disc 112 that is placed inside the cassette 1,200,300. Alternatively, the locking can be achieved by a receptacle moulded into the base 5 of the cassette 1,200,300 that is a fit for a shaft coupled to the actuator 101 locking them together. The cassette 1,200,300 remains square with respect to the central axis of the device 100, thereby ensuring a constant distance from the objective lens 102 of the device 100 to the upper portion 7,208 of the cassette 1,200,300.

[0125] The cassette 1,200,300 also maintains its square position with the central axis of the capturing device 100 to the upper portion 7,208 of the cassette 1,200,300 to maintain the correct distance from camera to the cassette. If necessary any deviation of this can be negated by, for example, minor adjustments of the vertical distance of the cassette 1,200,300 relative to the lens 102 using actuators.

[0126] To use the cassette 1,200,300 with the image capturing device 100, a liquid sample must be prepared. The sample is prepared by a method as described in the prior art which is typical of preparing a sample for analysis and is as follows. For example, 45 ml of a flotation solution (FS) is measured into a graduated cylinder. A sample is firstly homogenised thoroughly and 5 g of the homogenised sample is added to the flotation solution to provide a sample suspension. The dilution ratio of sample:flotation solution is measured by volume (typically a dilution ratio of 1:10). The sample suspension is further homogenised thoroughly in the graduated cylinder using a spatula and passed through a filter. The pore size of the filter will be determined to be at least 5% greater than the largest dimension in the range of specific matter being identified in the test. For example, if the specific matter being identified in the test has a diameter of 100 m, the pore size of the filter will be 105 m. The sample is then further filtered through a wire mesh having a pore size of, for example, 212 m, so as to remove any remaining large debris.

[0127] A portion of the filtered sample is obtained using a syringe. The filtered sample is mixed thoroughly and constantly while the sample is being taken.

[0128] Once the sample is prepared by one of the methods set out above (or any suitable method for preparing such samples), the sample is introduced with minimum delay after preparation to the inlet channel 20, with maximum retention of the buoyant particles of interest, and with minimum (ideally zero) entrapped air or gas, especially when the latter is in the form of microscopic bubbles. The prepared sample is passed into the conduit 12 of the enclosed sample-receiving channel 6 via the inlet port 20 using a syringe or similar pumping mechanism. A small air release port 14, opposite the inlet port 20, allows the air in the conduit 12 to escape as the liquid sample enters the enclosed sample-receiving channel 6 (see FIG. 1B and FIG. 2A). The cassette 1,200,300 can be held, during sample loading, so that the central axis of the cassette 1,200,300 is horizontal and parallel to the ground, with the plane of the cassette 1,200,300 therefore vertical and perpendicular to the ground. The inlet port 20 should be directly below the central axis of the cassette 1,200,300, and the air release port 14 at the opposite point, directly above the central axis, so that bubbles will naturally tend towards the air release port 14 due to their buoyancy in the sample. The filling of the conduit 12 is complete when the sample reaches this small air release port 14. The effect of the inlet port 20 will be to hold the sample inside, with no tendency to leak out at the air release port 14, which can therefore be left unsealed and open.

[0129] Now looking at FIG. 5A, the lens 102 of the image capturing device 100 is configured to achieve the maximum FOV with the minimum requirement of resolution needed for the automated analysis to optimise processing time and performance. This is achieved by matching the optical resolution power of the lens 102 to the pixel resolution of the camera, as denoted by the following formula:

R=1.22/(NAobj+NAcond)0.61/NA/2

[0130] For =520 nm (green light)

[0131] R=3 m, the minimum distance between resolvable points, in the same units as lambda specified.

[0132] NA=0.1 (Numerical aperture of the lens used)

[0133] This is the level of detail recognisable by the objective lens 102 used. The detector 122 should perform similar levels to get the best of the system. Therefore, one must decrease the working distance to reduce magnification and gain with respect to the field of view.

[0134] The size of the field of view now matches the width of the enclosed sample channel 6 in the cassette 1,200,300. The internal surfaces of the holder of the lens 102 can have a sandpaper or felt lining (or similar surface which reduces or inhibits reflection of light) to reduce any internal reflections, which could distort or reduce the clarity of the image on the sensor/detector 122 of the image capturing device 100. The column between the lens 102 and the sensor/detector 122 in the image capturing device 100 has a minimum diameter to alleviate internal reflections that may cause aberration of the image being taken. The image capturing device 100 has been selected to achieve a low-cost solution while achieving suitable resolution and quality of image.

[0135] The width of the upper portion 7,208 is determined by the FOV of the objective lens 102 of the image capturing device 100 at the chosen magnification. By making the width of the upper portion 7,208 correspond to the width of the FOV, the entire surface of the enclosed channel 6 can be scanned by motion of the cassette 1,200,300 in one direction or in one plane only or by motion of the objective lens 102 in one direction or one plane only.

[0136] FIG. 6 illustrates, by way of a block-diagram, a system 400 for obtaining images of a sample loaded in the closed sample-receiving channel 6 of the cassette 1,200,300. Images obtained from the closed sample-receiving channel 6 are captured in sequence as the actuator 101 moves the cassette 1,200,300 in a one plane only until the entire surface area of the enclosed sample-receiving channel 6 is covered or a sufficient representative series of images has been captured. Alternatively, the actuator 101 moves the objective lens 102 until the entire surface area of the enclosed sample-receiving channel 6 is covered or a sufficient representative series of images has been captured. These images are processed by image analysis software to establish the presence or absence of specific microorganisms or other items of interest. An example of this would be a farmer carrying out a count of eggs of one or more specific parasites present in a faecal sample taken from animals on a farm. It is also possible for the user to send the results of any test to a cloud-based database for storage and subsequent analysis. In this way, or by other methods, by comparing results from different locations, or from the same location over time, the growth, spread or containment of parasitic infection can be monitored. This information could be very valuable at local, regional, national and international levels, for example to monitor patterns of infestation of livestock. National entities, such as a government department of agriculture, or regional authorities, for example at a European level, could use this information for setting policies on animal treatment, animal movements, disease control, veterinary practice, etc.

[0137] Turning now to FIG. 7, the ability to differentiate between species using an image recognition algorithm will be directly proportional to the quality of the image captured and the contrast achieved. The contrast in FIG. 7A is sufficient for the human eye to differentiate shapes based on aspect ratio, contrast and characteristic of the interior of the oocyst. The dark ringed spherical shapes marked in FIG. 7B clearly identifies air bubbles. The fainter oocysts marked in FIG. 7C represent Coccidia. The concentration of the items of interest (for example, oocysts) in the cassette described herein increases the capturing efficiency over the prior art devices by decreasing the number of images required for an automated capture. Thus, the cassette and the system of the claimed invention provides a distinct advantage of being capable of analysing higher volumes while acquiring less images, in an automated process, where these images can be captured at a suitable resolution to be able to see items of interest clearly enough to identify them.

[0138] Turning now to FIG. 9, there is illustrated a typical photomicrograph of one FOV of eggs from ovine (FIG. 9A, 9B), bovine (FIG. 9C, 9D) and equine (FIG. 9E) faecal samples visible (circled) in the channel of the cassette of the claimed invention and which was captured using the automated system of the invention. The types of eggs are labelled as follows; N=Nematodirus; M=Moniezia; C=coccidia; S=Strongyles. The cassette concentrates the particles for identification which enables the process to be automated. As explained above, the concentration of the items of interest (for example, oocysts) in the cassette described herein increases the capturing efficiency over the prior art devices by decreasing the number of images required for an automated capture. The image quality captured and the contrast achieved by concentrating the items of interest using the automated system permits the user to see the items of interest clearly enough to identify them user fewer images, which provides a distinct advantage over the prior art devices, which cannot be automated. This also demonstrates that the cassette and system of the claimed invention can be used to determine the presence of matter from samples across different species.

[0139] Materials and Methods

[0140] Fresh faecal samples were collected from naturally infected livestock. These were then fixed using a 10% formalin solution (4% formaldehyde) and stored at 4 C. until testing. Composite samples for groups of animals were combined in groups of 10 individuals and mixed well (Morgan et al., 2005 (Morgan, E. R., Cavill, L., Curry, G. E., Wood, R. M., Mitchell, E. S. (2005) Effects of aggregation and sample size on composite faecal egg counts in sheep. Veterinary Parasitology. 131:79-87), Rinaldi et al., 2014 (Rinaldi, L., Levecke, B., Bosco, A., Ianniello, D., Pepe, P., Charlier, J., Cringoli, G., Vercruysse, J., 2014. Comparison of individual and pooled faecal samples in sheep for the assessment of gastrointestinal strongyle infection intensity and anthelmintic drug efficacy using McMaster and Mini-FLOTAC. Vet. Parasitol. 205, 216-223)).

[0141] Sample Preparation

[0142] The saturated sodium chloride flotation solution is prepared in the laboratory by adding NaCl to 11 of heated H.sub.2O (40-50 C.) until no more salt goes into solution (400-500 g). NaCl is dissolved using a stirrer and the solution left to fully saturate overnight. The specific gravity (S.G.) is then checked using a hydrometer for an S.G of 1.2 (Cringoli et al., 2010 (Cringoli G. Rinaldi L. Maurelli M. P. & Utzinger J (2010) FLOTAC: new multivalent techniques for qualitative and quantitative copromicroscopic diagnosis of parasites in animals and humans, Nature Protocols: 5 (3): 503-515)).

[0143] 45 ml of the flotation solution (FS) is measured into a graduated cylinder. The preserved sample was well homogenised and 5 g was added to the flotation solution, measured by volume (dilution ratio 1:10). The sample suspension is homogenised thoroughly in the cylinder using a spatula and passed through a (tea strainer) filter having a pore size 5% larger than the particles of interest in the sample. The sample is then further filtered through a wire mesh (pore size of 212 m) to remove any remaining large debris.

[0144] Filling the Device

[0145] 4 ml of the filtered sample suspension is drawn up into a syringe. Care must be taken at this point to work quickly and to mix the sample in the syringe to avoid egg flotation within the syringe. This can be achieved by inverting the syringe 2-3 times before filling. Fit the syringe into the inlet port (the filling hole) of the cassette, with the cassette held vertically and the inlet port positioned below the horizontal axis. Depress the plunger and fill the enclosed sample-receiving channel, allowing the air to escape through the air-release port at the top. Once the enclosed sample-receiving channel is full, the syringe is removed.

[0146] Egg Identification and Count

[0147] The number of eggs per gram (EPG) of faeces is calculated by multiplying the observed count with the dilution factor and then divided by the volume of the solution examinedthe default unit for reporting FECs is EPG.

[0148] Eggs per gram (EPG) calculation for the cassette and system described herein.

[0149] 1 g of faeces in 9 ml flotation solution; (45 ml of solvent to 5 ml of solute) 2.2 ml in cassette; and multiplication factor of 4.55 to calculate EPG value. (1 g faeces in 9 ml flotation solution (giving a total volume of 10 ml); 2.2 ml examined, therefore, 10 ml/2.2 ml=4.55)

[0150] EPG Calculation for McMaster System

[0151] 1 g of faeces in 9 ml flotation solution; 0.3 ml in slide (given the standard 0.15 ml volume in chamber 1, 0.15 m in chamber 2); and multiplication factor of 33.33 to calculate EPG value. (1 g faeces in 9 ml (giving a total volume of 10 ml); 0.3 ml flotation solution examined, therefore, 10 ml/.3 ml=33.33)

[0152] EPG Calculation for Mini-Flotac

[0153] 1 g of faeces in 9 ml flotation solution; 2 ml in device (given the standard 1 ml in chamber 1, 1 ml in chamber 2); and multiplication factor of 5 to calculate EPG value. (1 g faeces in 9 ml (giving a total volume of 10 ml), 2 ml flotation solution examined, therefore, 10 ml/2 ml=5).

[0154] Results

TABLE-US-00001 TABLE 1 Comparison chart for volume examined versus area to be examined, and sensitivity of eggs per gram (EPG): Suitable for Area required Centrifuge automated Method Dilution Total Volume to be viewed Sensitivity needed recognition McMaster 1:15 2 0.15 ml 2 1 cm.sup.2 50 EPG No No Flotac 1:10 5 ml 3.24 cm.sup.2 2 EPG Yes No Mini- 1:10 2 1 ml 2 3.24 cm.sup.2 5 EPG No No Flotac Invention* 1:10 2.2 ml 1 2.82 cm.sup.2 4 EPG No Yes *with a 2.2 ml volume and a closed sample-receiving channel profile of 60 angled sidewalls, a channel depth of 3.5 mm, a base of channel 5.54 mm, and a platform size 1.5 mm 2.5 mm.

[0155] The McMaster method as described in Table 1 would theoretically require a minimum contamination level of 50 EPG of faeces to identify 1 egg in 0.3 ml, as opposed to the device and system described herein which would only require a contamination level of 4 EPG to identify 1 egg in 2.2 ml. Therefore, the device and system described herein is more accurate and would be less likely to produce a false negative result.

[0156] The device and system described herein can easily increase the volume to be examined, which is in line with the 5 ml examined in the Flotac method described in Table 1. However, the automated process time will be increased proportionally and as the Flotac system is rarely used due to its long processing time, centrifuging requirements and labour-intensive nature of the method. It is suggested that the volume examined in the device and system described herein should represent an increase above the highest level of the most frequently used method, while maintaining minimal process time.

[0157] The sensitivity of the test is directly proportional to the volume examined and this volume determines the required number of images to be analysed. The proposed concentration of the suspended buoyant matter at the viewing surface area relative to the volume examined allows the number of images to be transmitted to be kept to a minimum while still maintaining sensitivity. As internet connectivity increases and improves, the device described herein can be optimised to increase the number of image acquisitions and therefore further increasing the sensitivity of the device.

[0158] A comparison of coprological examination techniques between the mini-FLOTAC, McMaster and the system described herein are provided in Table 2 below. Three ovine faecal samples were analysed for the presence of GIT parasites coccidian oocysts, Nematodirus, Strongyles and Strongoloides eggs.

TABLE-US-00002 TABLE 2 Trial Report Results No. of Eggs No. of Eggs No. of Eggs No. of Eggs detected in detected in detected in detected in No. of 1.sup.st 2.sup.nd 1.sup.st Mini- 2.sup.nd Mini- Eggs Sample McMaster McMaster Flotac Flotac detected in Ref. No Chamber Chamber EPG Chamber Chamber EPG Invention EPG 68 1 1 66.66 4 3 35.00 16 72.8 68 0 1 33.33 1 2 15.00 10 45.5 68 1 0 33.33 1 1 10.00 12 54.6 Mean 44.44 20.00 57.63 68 0 1 33.33 2 3 25.00 9 40.95 68 1 0 33.33 6 2 40.00 9 40.95 68 1 0 33.33 0 4 20.00 8 36.4 Mean 33.33 28.33 39.43 68 2 1 99.99 3 2 25.00 5 22.75 68 2 0 66.66 2 4 30.00 12 54.6 68 0 1 33.33 3 3 30.00 10 45.5 68 3 2 166.65 4 1 25.00 12 54.6 Mean 91.66 27.50 44.36 Overall 59.99 25.50 46.87 Mean FEC

[0159] These results demonstrate that the invention provides an accurate estimation of the true faecal egg count when compared to either the McMaster and Mini-FLOTAC systems. The importance of the higher volume being analysed is demonstrated by the McMaster results which provide a skewed mean value due to the high extrapolation factor. In this case, the high multiplication factor has resulted in a higher reading but the major downside is that any false negative result has a far greater consequence on the overall analysis. By this we mean that when a very small sample volume is found to be negative for the presence of a parasite, that result is extrapolated to give a negative result for the test as a whole. The distribution of parasite eggs within a sample is not homogenous and therefore, the greater the sample size, the less the variability in the results and the higher the sensitivity.

[0160] The availability of testing the sample immediately on site with the suggested automated process will also increase the accuracy of the results as samples, once excreted, deteriorate over time and due to exposure to the environment.

[0161] In the specification, the terms comprise, comprises, comprised and comprising or any variation thereof and the terms include, includes, included and including or any variation thereof are considered to be totally interchangeable and they should all be afforded the widest possible interpretation and vice versa.

[0162] The invention is not limited to the embodiments hereinbefore described but may be varied in both construction and detail.