An amine and polyamine detection method using ion mobiilty spectrometry

Abstract

This invention relates to an analyte detection method, an Ion Mobility Spectrometer (IMS) (10) adapted for point of care use including a novel heater assembly (600), and consumables (100; 400), including reagents (130) and a vial (410) and stopper (420) 5 assembly. The reagent is an alkali metal hydrogen carbonate solution which on heating changes pH such that it can react with a sample (120) to generate amines and polyamines which are driven off as a gas (126) and ionised (127) such that they can be detected.

Description

AN ANALYTE DETECTION METHOD

[0001] This invention relates to an analyte detection method, an Ion Mobility Spectrometer (IMS) adapted for point of care use including a novel heater assembly, and consumables, including reagents and a vial and stopper assembly.BACKGROUND

[0002] Ion mobility spectrometry (IMS) is over 100 years old. It is analytical method that separates gaseous molecules with an induced charge based on mass and charge (Verbeck et al., 2002; Creaser et al., 2004; Kanu et al., 2008; Westhoff et al., 2009; Dodds and Baker, 2019). It is a highly sensitive method of analyte detection, (ng / L to pg / L) that requires no preconcentration, or labelling, of the analyte before analysis (Westhoff et al., 2009).

[0003] It has the advantage of rapidity and extremely low limits of detection and quantitation.

[0004] IMS has been used as an in vitro diagnostic device to detect bacterial infections, viruses (Blankenstein et al., 2015; DeMuri and Wald, 2014; Slots and Slots, 2011 ; Corstjens, Paul L A M et a!., 2016) and cancers (Westhoff et a!., 2009;Gugliucci, 2004; Miller-Fleming et al., 2015).

[0005] Saliva has been used as a specimen to detect patients with HIV, HPV and Hepatitis C (Corstjens, Paul L A M et al., 2016).

[0006] However, a critical problem in the use of IMS for detecting amines and polyamines as a point of care device, is that the reagent (3.75M potassium hydroxide) for raising the pH above the maximal pKa for polyamines is extremely corrosive (The European Parliament and the Council of the European Union, 2008b, 2008a).

[0007] This problem is overcome by using a reagent which is not extremely corrosive when handled (has a pH below 11 .5) but which when subsequently heated in a modified IMS (for point of care use) can achieve a pH above the maximal pKa for polyamines, such that they become unionised and are not subject to hydrogen bonding in a polar solvent. This means they have a lower energy requirement for volatilisation, and volatalise at an appropriate temperature when above the pKa.

[0008] CN106198702 teaches detecting scheduled drugs such as heroine, ketamine, and cocaine in saliva samples utilising IMS. It does not look at polyamines and usessodium carbonate or sodium bicarbonate to adjust the pH to between 8.5 and 9.5 together with “solubilisers” - methanol, ethanol or acetonitrile to improve sensitivity.

[0009] The problem with the use of sodium carbonate or sodium bicarbonate is the pH is below the pKa of many of the amines / poyamines which Applicant seeks to detect when identifying e.g. specific viruses, bacteria or cancers in samples, where it is desired that the pH is above 9.5 and more preferably above 10 as the pKa of many amines to be detected are above pH 9.5 as illustrated in Tables 1 and 2 below which illustrate the pKa of amino acids (Table 1 ) and a range of primary, secondary, tertiary, aryl and heterocyclic amines (Table 2). Table 1Table 2List of pKb values of different amines

[0010] Table 2 is from - https : / / i.pinimg.com / 736x / d0 / 9a / 34 / d09a34ad182940bbdc878f1fc973da91.jpgBRIEF SUMMARY OF THE DISCLOSURE

[0011] In accordance with a first aspect of the invention there is provided a point of care method of analyte detection using Ion Mobility Spectrometry comprising the steps of: i) Obtaining a sample from a patient; ii) Placing the sample together with a reagent into an Ion Mobility Spectrometer; characterised in that iii) The reagent is a solution with a pH of below 11 .5 at room temperature, which on heating to up to 110°c disassociates to raise the pH above a pKa of one or more target amines and polyamines present in the sample, and iv) The reagent and sample are heated, and the one or more target amine(s) and / or polyamine(s) are gassified and ionised to form gaseous analytes which travel across a drift region of the IMS to a Faraday plate where they are detected characterised in that the reagent is one of a potassium, rubidium or caesium hydrogen carbonate solution

[0012] In one embodiment the point of care method of analyte detection further comprises the step of: v) Comparing a profile detected at the Faraday plate with a known profile indicative of a disease and making a diagnosis based on the comparison.

[0013] Preferably the reagent is a hydrogen carbonate solution.

[0014] Preferably the hydrogen carbonate takes the form of an alkali metal salt.

[0015] Preferably the alkali metal salt is a Cesium, Potassium, Rubidium or Sodium salt, and most preferably cesium hydrogen carbonate.

[0016] Preferably the the sample and reagent are heated to between 80 and 110Gc.

[0017] In accordance with a second aspect of the invention there is provided a consumable or kit of parts comprising a consumable for use with a point of care Ion Mobility Spectrometer (IMS) analyte detection device which consumable comprises a reagent which is a hydrogen carbonate solution selected from one of potassium hydrogen carbonate solution, caesium hydrogen carbonate solution or rubidium hydrogen carbonate solution.

[0018] Preferably the reagent has a concentration greater than 2.5M.

[0019] In one embodiment the reagent is supplied in a container comprising a dropper for applying the reagent to a sample.

[0020] The container may be a squeezy bottle with a dropper or a bottle with a separate pipette dropper.

[0021] In accordance with a third aspect of the invention there is provided a consumable or kit of parts comprising a consumable comprising a vial and a stopper wherein the vial has an open end remote from a closed end of the vial and comprises at least two slots or an aperture from which gaseous analytes derived from the sample, by the action of the reagent, escape when the vial, sample and reagent are heated in an IMS with the stopper in place.

[0022] In one embodiment the slots give rise to expandable sprung walls such that the stopper locks onto the vial when inserted in the open end.

[0023] Preferably the vial further comprises a shoulder just below the at least two slots for locating the vial in a heating chamber of a heater assembly of an IMS.

[0024] Preferably the stopper comprises a shoulder to limit the degree of insertion of the stopper into the open end, such that a lower region of the slots remain open allowing gassifed analytes to enter an ionising region of the IMS on heating therein.

[0025] In one embodiment the stopper has a stick extending from a lower end with a swab located at an end of the stick remote from that attached to the stopper.

[0026] Such an arrangement is great for e.g saliva swabs.

[0027] Of course the sample may be placed in a vial absent of a swab if collected as a liquid sample e.g. urine, sputum or blood, or as a tissue sample (from e.g a biopsy).

[0028] In accordance with a fourth aspect of the invention there is provided a point of care Ion Mobility Spectrometer (IMS) analyte detection device adapted for use with the method of the first aspect or the consumable of the second or third aspects comprising: i) an ionisation region; and ii) a drift region said ionisation region comprisinga heating assembly comprising a sample receiving chamber;a gas outlet; andan ionisation sourcesaid drift region comprising d) an ion shutter; e) drift rings; f) an aperture grid; and g) a Faraday plate; characterised in that iii) the sample receiving chamber is shaped to retain a vial containing a sample and reagent, which is sealed with a stopper and contains an aperture allowing the release of a gas into the ionising region; iv) a heater is shaped to provide a chamber to heat the vial containing the sample and reagent; and v) a management system controls the heater.

[0029] The management system ensures the heater is heated to a temperature, and for a time, such that amine(s) and / or polyamine(s) in the sample are driven off as a gas, and ionised in the ionisation region forming charged gaseous analytes which travel across the drift region of the IMS to the Faraday plate where they are detected.BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Embodiments of the invention are further described hereinafter with reference to the accompanying drawing, in which:Fig 1 is a diagram showing the main components and principle of Ion Mobility Spectroscopy (IMS);Figs 2a and 2b are diagrams illustrating embodiments of a first consumable of the invention (Reagent);Figs 3a, 3b and 3c are diagrams illustrating a second consumable of the invention (Vial and stopper - optionally with a swab connected via a stick);Fig 4a is an exploded view of a heating assembly of the invention for an IMS for use as a point of care analyte detection device;Fig 4b is a cross sectional view of the heating assembly of Fig 4a together with a vial and stopper (with stick and swap attached to the stopper as per one embodiment) as per Figs 3a and 3b in place in the assembly;Fig 5 is a graph showing temperature increase v time;Fig 6 is a graph of pH with temperature for a control, potassium hydroxide solution;Fig 7 is a graph of pH with temperature for sodium carbonate (outside the scope of the invention);Fig 8 is a graph of pH with temperature for potassium carbonate (outside the scope of the invention);Fig 9 is a graph of pH with temperature for rubidium carbonate (outside the scope of the invention);Fig 10 is a graph of pH with temperature for cesium carbonate (outside the scope of the invention);Fig 11 is a graph of pH with temperature for 1 .06 M sodium hydrogen carbonate (outside the scope of the invention);Fig 12 is a graph of pH with temperature for 2 M potassium hydrogen carbonate;Fig 13 is a graph of with temperature for 2 M potassium hydrogen carbonate pH adjusted);Fig 14 is a graph of pH with temperature for cesium hydrogen carbonate;Fig 15 is a graph of pH with temperature for 3.75 M cesium hydrogen carbonate;Fig 16 is a graph of pH with temperature for 4.33 M rubidium hydrogen carbonate; andFig 17 is a bar chart showing the results of the different compounds tested.DETAILED DESCRIPTION

[0031] Referring to Fig 1 the IMS (10) comprises two regions - an ionisation region (20) and a drift region (30).

[0032] The ionisation region comprises a sample receiving chamber (not shown) an ionisation source (22) and a gas outlet (24) via which unionised molecules (26) leave.

[0033] The drift region comprises an ion shutter (32), drift rings (34) an aperture grid (36) and a Faraday plate (38).

[0034] The IMS of the invention has been adapted for use for point of care analyte detection (see Fig 4a and 4b and Figs 2a, 2b, 3a, 3b and 3c) by the inclusion of a heater assembly (600) comprising a heater (640) which is annular, the assembly being shaped to retain a vial (410) containing a sample (120) and the reagent (130) which is sealed with astopper (420) and contains one or more apertures (A), preferably formed from slots (460), allowing the release of gaseous analytes (126) generated when the sample (120) and reagent (130) are heated, the reagent causing amine(s) / polyamine(s) present in the sample to be volatilised (126) such that they enter the ionisation region (20) and are ionised (127) to charged gaseous analytes (128).

[0035] In use un-ionised molecules (26) are blocked from entering the drift region (30) by the ion shutter (32), while a drift gas (31) carries these molecules away through a gas outlet (24).

[0036] The ionised analytes (128) flow in the opposite direction to that of an electrical field (40) against the drift gas (31 ) which acts as a barrier to charged gaseous analyte (128) migration in the electrical field and reduces the length of the drift region required to achieve separation of molecules. The drift rings (34) maintain the electrical field across the column.

[0037] As the charged analytes (128) travel with the electrical field against the drift gas (31 ) flow, they will reach the aperture grid (36) which acts as a gate barrier to all ions to make the drift to the Faraday plate calculations accurate. The Faraday plate (38) detects the electrical charge of analyte as it touches its surface, and this is recorded against the time from introduction to detection.

[0038] Once the charged analytes(128) contact the Faraday plate (38), its charge dissipates, and it registers on the detector. The molecule is no longer under the attractive influence of the electric field as it can only attract ionised molecules. It remains a gas due to the temperature of the drift region.

[0039] The reagents (130) of the invention, which is used to volatilise the amines, were determined by way of the following Experiments which identified favoured hydrogen carbonates:MATERIALS AND METHODS

[0040] It was noted that heating hydrogen carbonate solutions would drive off carbon dioxide and form the carbonate anion as shown in Table 3.

[0041] Table 3Table 3 Dissociation pKa ApKa / A°C constants for carbonic acid buffer systems(Benyon and Easterby 1996)H2CO3to HCO3- 6.35 -0.0055HCO3- to CO3210.33 -0.009

[0042] The change in pKafor the hydrogen carbonate to carbonate buffer system when reaching 100 °C is calculated to be a drop of -0.72 for an 80 °C change in temperature (Benyon and Easterby, 1996). This would mean that the pH would be expected to buffer around pH 9.61 .

[0043] The alkali metal carbonate and hydrogen carbonate salts used in this study are shown in Table 4.

[0044] Table 4

[0045] Table 4 Formula weight, CAS number, supplier, catalogue number and lot number of alkali metal hydrogen carbonates and carbonates.

[0046] Materials were weighed using a three decimal place balance Bonvoisin # 5002 S / N HZ5003B. pH was measure using a Hanna Instruments Edge HI 2020 S / N C1021024 software v1.18. The electrode was a Hanna Instruments H1 11310 S / N 533505, firmware 1 .06. This has an integral temperature sensor. pH was measured at30 second intervals using the logging function on the Edge.

[0047] Solutions were weighed into a glass Universal, containing a magnetic stirrer bar. The sample was then place on a preheated hot plate stirrer SH-2 with stirring at the lowest setting, and heating at the highest setting. The pH was measured as the temperature was increased as a measure of the carbon dioxide driven off.

[0048] The data were downloaded from the Edge to a USB drive as a .CSV file and this was opened in MS Excel, where graphs were generated.RESULTS

[0049] Fig 5 shows the rate of change in temperature of a 10 g volume of water on the SH-2 hotplate stirrer set at maximum heating rate. The initial lag phase represents the time taken for the hotplate stirrer to heat up. a) 3.75 M (15 %) POTASSIUM HYDROXIDE

[0050] Fig 6 (control) shows that 3.75 M potassium hydroxide is above the threshold of pH 11 .5 for corrosive alkali solutions (The European Parliament and the Council of the European Union, 2008a). Heating reduces the pH measured just below the corrosive limit. b) 3.5 M SODIUM CARBONATE

[0051] Fig 7 shows a 3.5 M solution.. The pH of the 3.5 M sodium carbonate is above the corrosion limit of pH 11 .5 (The European Parliament and the Council of the European Union, 2008a). Like 3.75 M potassium hydroxide, the pH declines with increasing temperature, and does not fall below pH 11 .5 until the temperature is around 50 °C. c) POTASSIUM CARBONATE

[0052] Fig 8 shows the pH of 3.75 M potassium carbonate is also above the corrosion limit of pH 11.5 (The European Parliament and the Council of the European Union, 2008a). Like 3.75 M potassium hydroxide, the pH declines with increasing temperature, and does not fall below pH 11.5 until the temperature is 60 °C.d) 2 M RUBIDIUM CARNONATE

[0053] Figure 9 shows the effect of heat on the pH of 2 M rubidium carbonate solution

[0054] Rubidium carbonate was prepared as a 2 M solution due to the limited amount of material available. The pH of 2 M rubidium carbonate is above the corrosion limit of pH 11 .5 (The European Parliament and the Council of the European Union, 2008a). Like 3.75 M potassium hydroxide, the pH declines with increasing temperature, and does not fall below pH 11 .5 until the temperature is 65 °C. e) 4M CESIUM CARBONATE

[0055] Fig 10 shows the pH of 4 M cesium carbonate is above the corrosion limit of pH 11 .5 (The European Parliament and the Council of the European Union, 2008a). The pH remains above 11 .5 even when heated above 100 °C.

[0056] Cesium has the largest ionic radius of the stable metals and is the most alkaline. The solution did not boil at 105 °C due to the high salt concentration in the solution which causes boiling point elevation. f) 1.06 M SODIUM HYRDOGEN CARBONATE

[0057] The concentration of the sodium hydrogen carbonate solution was at near the limit of solubility. This is why the concentration is lower than the potassium hydroxide and potassium carbonate solutions.

[0058] Fig 11 shows the pH of 1 .06 M sodium hydrogen carbonate starts around 8.0 and on heating increases by around 1.2 pH units at around 100 °C. The change in pH is limited by the solubility of sodium hydrogen carbonate. g) 2 M POTASSIUM HYDROGEN CARBONATE

[0059] The concentration of the potassium hydrogen carbonate solution was at near the limit of solubility. This is why the concentration is lower than the potassium hydroxide and potassium carbonate solutions

[0060] Fig 12 shows the pH of the 2 M potassium hydrogen carbonate starts around pH 8 like the 1 .06 M sodium hydrogen carbonate solution. Due to the higher concentration the pH is around 0.5 pH units higher than the 1 .06 M sodium hydrogen carbonate solution. h) 2 M POTASSIUM HYDROGEN CARBONATE - PH ADJUSTED

[0061] The pH of the 2 M potassium hydrogen carbonate solution was raised by adding 2 M equivalents of potassium hydroxide. The behaviour was identical to the potassium carbonate solution in c)

[0062] Fig 13 shows the heating of the pH adjusted solution makes a buffered system that sits on the pKa. This drifts down on heating to close to pH 9.61 which is the pKa for bicarbonate-carbonate buffer at 100 °C. i) CESIUM HYDROGEN CARBONATE

[0063] Fig 14 shows Cesium hydrogen carbonate is freely soluble in water and easily dissolved in 10 ml of water. This was not volumetric, so the concentration is unknown as the dissolved salt increased the total volume in the vial. j) 3.75 M CESIUM HYDROGEN CARBONATE

[0064] A volumetric solution of cesium hydrogen carbonate was prepared at 3.75 M concentration.

[0065] Fig 15 shows the higher concentration of cesium hydrogen carbonate in the volumetric solution gave a higher final pH than the 72 % w / v solution. k) 4.33 M RUBIDIUM HYDROGEN CARBONATE

[0066] A volumetric solution of rubidium hydrogen carbonate was prepared at 4.33 M concentration.

[0067] Fig 16 shows that the 4.33 M rubidium hydrogen carbonate solution increases pH similarly to the cesium hydrogen carbonate solution.I) START AND END PH FOR SOLUTIONS

[0068] Table 5 below shows the start and end pH for the tested reagents.

[0069] Table 5

[0070] The data from Table 5 is shown graphically in Fig 17.CONCLUSION

[0071] A critical problem in the use of IMS for detecting polyamines as a point of care device is that the reagent for raising the pH above the maximal acid dissociation constant, pKa, for polyamines is extremely corrosive (The European Parliament and the Council of the European Union, 2008b, 2008a).

[0072] The relatively low solubility of sodium and to a lesser extent the potassium salt of the hydrogen carbonate anion limits the concentration in the reagent and make these salts less preferred as a replacement for 3.75 M potassium hydroxide (The European Parliament and the Council of the European Union, 2008b) than the cesium (and rubidium) salts which can raise the pH of a specimen above the highest pKa for polyamines and amines.

[0073] The limit of solubility for both rubidium and cesium salts of the hydrogen carbonate are substantially greater than those of sodium and potassium making them preferred reagents.

[0074] The data show that cesium hydrogen carbonate can reach pH > 10.5 when heated to 100 °C. This is close the pH of potassium and sodium carbonates and potassium hydroxide after heating to 100 °C. Cesium is the most alkaline ion of any stable element in the periodic table (Sharpe, 1986).

[0075] Cesium chloride has similar toxicity to the chloride salts of sodium and potassium (Melnikov and Zanoni, 2010; Johnson et al., 1975). The low toxicity, the high solubility of cesium and rubidium salts of the hydrogen carbonate anion, and the capacity to drive off carbon dioxide when heated to 100 °C makes these ideal compounds for use in point of care testing for amines and polyamines by IMS.

[0076] Having established that certain reagents can be used to overcome the issues surrounding using an extremely corrosive reagent to volatilise amines and polyamines for use in point of care testing using IMS the Applicant modified an IMS device and developed novel presentations of consumables for use with the method of the invention.

[0077] Referring to Figs 2a and 2b there is illustrated a first consumable (100) of the invention (Reagent (130)) which is one of a solution of: sodium hydrogen carbonate (Fig 11 ), potassium hydrogen carbonate (Figs 12 and 13 ), and cesium hydrogen carbonate (Figs 14 and 15).

[0078] The reagent (130) is provided in a bottle (300) provided with a dropper (310) and lid (320) (Fig 2a) or an independent dropper (310) comprising a pipette (312), lid (320) and teat (314).

[0079] The dropper simplifies the methodology of the invention since the reagent can be simply and safely added to the sample (120) in the vial (410).

[0080] Referring to Figs 3a, 3b and 3c there is illustrated a second consumable (400) of the invention (Vial (410) and stopper (420).)

[0081] The vial (410) has been specially adapted for use in the IMS (10) of another aspect of the invention and comprises an opening (440) for receiving a sample (120) and the reagent (130) remote from a closed end (450). Extending from the opening (440) are at least a pair of slots (460), which extend to a shoulder (470) creating slightly expandable, sprung walls (430). When the stopper (420) is inserted into the opening (440) apertures (A) are formed, where the lower region (462) of the slots (460) meet the shoulder (470), which apertures align with apertures (A’) in a mid-casing (650) component of the heating assembly (600) of the IMS (10).

[0082] In a preferred embodiment of the second consumable (400) the sample is collected on a swab (510) which forms an integral part of the consumable being connected, by a suitably sized stick (500) which is connected to a lower end (422) of the stopper (420) with the swab being attached to the end (502) of the stick remote from that connected to the stopper. Of course, as illustrated in Fig 3c), which shows the stopper (420) inserted in the vial (410), the stick and swab are not essential.

[0083] The stopper (420) also comprises a shoulder (424) above a stepped or sloping region (426) which facilitates substantial closing of the vial (stopper retention in the vial opening (440) ) and most importantly ensures a path via which the gas (126) generated on heating leaves the vial via apertures (A) and is also able to leave the heating assembly.

[0084] Fig 4a and 4b (in conjunction with Fig 1) show a novel heating assembly (600) for a point of care IMS.

[0085] Fig 4a is an exploded view of a heating module assembly (600) for a second consumable (400) as described above.

[0086] The heating module assembly (600) comprises an annular chamber (642) in which a vial (410) containing a sample (120) and reagent (130) is evenly heated, controlled by a management system - not shown.

[0087] The heater (640) is located in a base casing member (610) and held firmly in place by a washer (620) and screws (630). A mid casing member (650) comprises apertures (A’) located such that they are aligned with the openings (A) in the vial (410).

[0088] A lid (660) and sprung plunger (670), with compression spring (672), are held in place by a plunger retainer (680) and screws (682).

[0089] In use, the second consumable (400) is located within an aperture (690) running through the lower (612) assembly components and is retained in place by the upper (614) assembly components which serve as a lid.

[0090] Fig 4b shows the assembly (600) with the heater (640) and gas exit (E) with the vial (410), stopper (420), stick (500) and swap (510) in place in the device.REFERENCES

[0091] Benyon, R.J. and Easterby, J.S. (1996), A1 Properties of Common Buffers - carbonic acid: Buffer Solutions - the basics, Oxford University Press, Oxford

[0092] . Blankenstein, T., Lytton, S.D., Leidl, B., Atweh, E., Friese, K. and Mylonas, I. (2015), “Point-of-care (POC) diagnosis of bacterial vaginosis (BV) using VGTest™ ion mobility spectrometry (IMS) in a routine ambulatory care gynecology clinic”, Archives of gynecology and obstetrics, Vol. 292 No. 2, pp. 355-362.

[0093] Corstjens, Paul L A M, Abrams, W.R. and Malamud, D. (2016), “Saliva and viral infections”, Periodontology 2000, Vol. 70 No. 1 , pp. 93-110.

[0094] Greaser, C.S., Griffiths, J.R., Bramwell, C.J., Noreen, S., Hill, C.A. and Thomas, C.P. (2004), “Ion mobility spectrometry: a review. Part 1. Structural analysis by mobility measurement”, Analyst, Vol. 129 No. 11 , pp. 984-994.

[0095] DeMuri, G.P. and Wald, E.R. (2014), “The Group A Streptococcal Carrier State Reviewed: Still an Enigma”, Journal of the Pediatric Infectious Diseases Society, Vol. 3 No. 4, pp. 336-342.

[0096] Dodds, J.N. and Baker, E.S. (2019), “Ion mobility spectrometry: Fundamental concepts, instrumentation, applications, and the road ahead”, Journal of the American Society for Mass Spectrometry, Vol. 30 No. 11 , pp. 2185-2195.

[0097] Firpo, M.R. and Mounce, B.C. (2020), “Diverse functions of polyamines in virus infection”, Biomolecules, Vol. 10 No. 4, p. 628. NB 2022-01 133 Gugliucci, A. (2004), “Polyamines as clinical laboratory tools”, Clinica chimica acta, Vol. 344 No. 1-2, pp. 23-35.

[0098] Johnson, G.T., Lewis, T.R. and Wagner, W.D. (1975), “Acute toxicity of cesium and rubidium compounds”, Toxicology and applied pharmacology, Vol. 32 No. 2, pp. 239-245.

[0099] Kanu, A.B., Dwivedi, P., Tam, M., Matz, L. and Hill Jr, H.H. (2008), “Ion mobilitymass spectrometry”, Journal of mass spectrometry, Vol. 43 No. 1 , pp. 1-22.

[0100] Melnikov, P. and Zanoni, L.Z. (2010), “Clinical effects of cesium intake”, Biological trace element research, Vol. 135, pp. 1-9.

[0101] Michael, A.J. (2016), “Polyamines in eukaryotes, bacteria, and archaea”, Journal of Biological Chemistry, Vol. 291 No. 29, pp. 14896-14903.

[0102] Miller-Fleming, L., Olin-Sandoval, V., Campbell, K. and Raiser, M. (2015), “Remaining Mysteries of Molecular Biology: The Role of Polyamines in the Cell”, Journal of Molecular Biology, Vol. 427 No. 21 , pp. 3389-3406.

[0103] Moinard, C., Cynober, L. and Bandt, J.-P. de (2005), “Polyamines: metabolism and implications in human diseases”, Clinical Nutrition, Vol. 24 No. 2, pp. 184-197.

[0104] Mounce, B.C., Olsen, M.E., Vignuzzi, M. and Connor, J.H. (2017), “Polyamines and their role in virus infection”, Microbiology and Molecular Biology Reviews, Vol. 81 No. 4, e00029-17.

[0105] Sharpe, A.G. (1986), 10 The Alkali Metals. Inorganic Chemistry, 2nd, Longman Scientific and Technical, Harlow, Essex.

[0106] Slots, J. and Slots, H. (2011), “Bacterial and viral pathogens in saliva: disease relationship and infectious risk”, Periodontology 2000, Vol. 55 No. 1 , pp. 48-69.

[0107] The European Parliament and the Council of the European Union (2008a), 3.2.3.1.2 Classification Criteria for Mixtures. PART 3: HEALTH HAZARDS. Regulation (EC) No 1272 / 2008 of the European Parliament and of the Council of 16 December 2008 on classification, labelling and packaging of substances and mixtures, amending and repealing Directives 67 / 548 / EEC and 1999 / 45 / EC, and amending Regulation (EC) No 1907 / 2006: CLP, Vol. 53.

[0108] The European Parliament and the Council of the European Union (2008b), Potassium hydroxide [1310-58-3]. Annex VI - Table 3.1 List of harmonised classification and labelling of hazardous substances. Regulation (EC) No 1272 / 2008 of the European Parliament and of the Council of 16 December 2008 on classification, labelling and packaging of substances and mixtures, amending and repealing Directives 67 / 548 / EEC and 1999 / 45 / EC, and amending Regulation (EC) No 1907 / 2006: CLP, Vol. 53.

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Claims

CLAIMS1 . A point of care method of analyte detection using Ion Mobility Spectrometry comprising the steps of: i) Obtaining a sample (120) from a patient; ii) Placing the sample together with a reagent (130) into an Ion Mobility Spectrometer (10); characterised in that iii) The reagent is a solution with a pH of below 11 .5 at room temperature, which on heating to up to 110sC dissociates to raise the pH above a pKa of one or more target amines and polyamines present in the sample, iv) The reagent and sample are heated and the one or more target amine(s) and / or polyamine(s) are gassified (126) and ionised (127) to form gaseous analytes (128) which travel across a drift region (30) of the IMS to a Faraday plate (38) where they are detected characterised in that the reagent is one of a potassium, rubidium or caesium hydrogen carbonate solution2. A point of care method of analyte detection as claimed in claim 1 further comprising the step of: v) Comparing a profile detected at the Faraday plate with a known profile indicative of a disease and making a diagnosis based on the comparison.

3. A point of care method of analyte detection as claimed in claim 1 or 2 in which the reagent is a caesium hydrogen carbonate solution.

4. A point of care method of analyte detection as claimed in claim 1 or 2 in which the reagent is a potassium hydrogen carbonate solution.

5. A point of care method of analyte detection as claimed in claim 1 or 2 in which the reagent is a rubidium hydrogen carbonate solution.

6. A point of care method of analyte detection as claimed in any of the preceding claims wherein the sample and reagent are heated to between 80 and 1 10SC.

7. A consumable (100), or kit of parts comprising a consumable, for use with a point of care Ion Mobility Spectrometer (IMS) analyte detection device (10) which consumable comprises a reagent which is a hydrogen carbonate solution selected from one of potassium hydrogen carbonate solution, caesium hydrogen carbonate solution or rubidium hydrogen carbonate solution.

8. A consumable (100), or kit of parts comprising a consumable, comprising a reagent as claimed in claim 7 wherein the reagent is caesium hydrogen carbonate solution9. A consumable (100), or kit of parts comprising a consumable, comprising a reagent as claimed in claim 8 which has a concentration of at least 2.5M.

10. A consumable (100) or kit of parts comprising comprising a consumable, comprising a reagent as claimed in claim 8 or 9 which is supplied in a container (300) comprising a dropper (310) for applying the reagent (130) to a sample (120).11 . A consumable (400), or kit of parts comprising a consumable, comprising a vial (410) and a stopper (420) wherein the vial has an open end (440) remote from a closed end (450) of the vial, and comprises at least two slots or an aperture (460) from which gaseous analytes (126) derived from a sample (120) by the action of a reagent (130) escape when the vial, sample and reagent are heated in an IMS (10) with the stopper in place12. A consumable (400), or kit of parts comprising a consumable, as claimed in claim 11 wherein the slots give rise to expandable sprung walls (430) such that the stopper locks onto the vial when inserted into the open end (440).

13. A consumable (400), or kit of parts comprising a consumable, as claimed in claim 11 or 12 wherein the vial further comprising a shoulder (470), just below the at least two slots (460), for locating the vial in a heating chamber (642) of an IMS.

14. A consumable (400), or kit of parts comprising a consumable, as claimed in any of claims 11 to 13 wherein the stopper (420) comprises a shoulder (424) to limit the degree of insertion of the stopper into the open end (440) such that a lower region (462) of the slots (460) remain open allowing gassifed analytes (126) to enter an ionising region (20) of the IMS (10) on heating therein.

15. A consumable (400) or kit of parts comprising a consumable, as claimed in any of claims 11 to 14 wherein the stopper has a stick (500) extending from a lower end (422) with a swab (510) located at an end (502) of the stick remote from that attached to the stopper. (420).

16. A point of care Ion Mobility Spectrometer (IMS) analyte detection device (10) adapted for use with the method of any of claims 1 to 6 or with a consumable (100; 400) as claimed in any of claims 7 to 15 comprising: i) an ionisation region (20); and ii) a drift region (30); said ionisation region comprising a) a heating assembly (600) comprising a sample receiving chamber (690); b) a gas outlet (24); and c) an ionisation source (22) said drift region comprising d) an ion shutter (32); e) drift rings (34); f) an aperture grid (36); and g) a Faraday plate (38); characterised in that iii) the sample receiving chamber (690) is shaped to retain a vial (410) containing a sample (120) and reagent (130) which is sealed with a stopper (420) and contains an aperture (A) allowing the release of a gas (126) into the ionising region (20); iv) a heater (640) is shaped to provide a chamber (642) to heat the vial (410) containing the sample (120) and reagent (130); and a management system that controls the heater (640).

17. A point of care Ion Mobility Spectrometer (IMS) analyte detection device (10) as claimed in claim 16 wherein the heater (640) forms an annular chamber (642).

18. A point of care Ion Mobility Spectrometer (IMS) analyte detection device (10) as claimed in claim 16 or 17 comprising a lower (612) heating assembly which retains the vial (410) and an upper (614) heating assembly which serves as a lid.

19. A point of care Ion Mobility Spectrometer (IMS) analyte detection device as claimed in claim 16 wherein the management system manages the temperature of the heater (640) such that amine(s) and / or polyamine(s) in the sample (120) are driven off as a gas (126), and are ionised (127) in the ionisation region (20), forming charged gaseous analytes (128) which travel across the drift region (30) of the IMS (10) to the Faraday plate (38) where they are detected.

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