Science, Technology and Medicine open access publisher.Publish, read and share novel research. Amperometric Glucose Sensors for Whole Blood Measurement Based on Dehydrogenase EnzymesMarco Cardosi1 and Zuifang Liu1[1] LifeScan Scotland Limited, a Johnson & Johnson Company,, UK1. Your use of this website constitutes acknowledgement and acceptance of our Terms & Conditions.
People with diabetes may have a new way to indicate their blood sugar level is too high or too low, by turning to our trusty canine friends, after researchers have found that dogs can help with hypoglycemia monitoring. The study, published in PLOS ONE, is the first of its kind to analyze whether trained dogs can accurately and consistently serve as an "early-warning system" to monitor blood sugar levels for their owners and notify them when the levels are too high or low.
Researchers then collected data from the owners to analyze whether the dogs were accurately able to respond to their owners' hypoglycemic levels, and also whether the owners experienced better blood sugar control and wider benefits. Additionally, the owners' data showed that the dogs notified them with "significant accuracy" during times of both low and high blood sugar. The study authors note that although dogs respond to their owners' high or low blood sugar levels, they cannot be entirely sure how they do this. They say their study confirms that trained detection dogs perform above the chance level, which is the level that would be expected if random choices were made.
Researchers recently revealed that they are creating a method for dogs to sniff out ovarian cancer. Tips on How to Stop Your Dog from BitingBeware - Not All Advertised Dog Rescues Really Are!
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Nycki Etherington was shocked when she was diagnosed with gestational diabetes mellitus (GDM). Etherington was given an oral glucose tolerance test between 24 and 28 weeks, and, like almost 10 percent of pregnant women, received a positive result. Enduring yet another procedure during pregnancy can seem like an unnecessary inconvenience—the oral glucose tolerance test takes several hours to complete—yet it’s essential to catch and treat this condition. When she was diagnosed, Etherington fretted about everything from what foods to eat to whether her baby would make it to term (premature birth is another concern with GDM). Etherington became diligent about reading food labels, counting carbs and eschewing high-sugar foods for lower-sugar ones. But despite the occasional elevated level, diet and exercise were enough to keep Etherington’s GDM under control during the pregnancy, as well as when she was pregnant with her second child. Tara Peel, a 43-year-old mother of two from Winnipeg, had a trickier time with her recent second pregnancy. Having to take insulin pushed Peel into the category of a high-risk pregnancy, which meant she had to abandon her natural at-home birth plan in favour of a hospital delivery and add monthly consultations with an endocrinologist to her schedule. A version of this article appeared in our September 2015 issue with the headline, “No sweets for two”, p.
By clicking "Create Account", I confirm that I have read and understood each of the website terms of service and privacy policy and that I agree to be bound by them. Health experts blame the increase on rising levels of obesity, which is driven by junk food diets and a lack of exercise. This lack of insulin means the body cannot break down sugar, or glucose, and use it for energy.
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Two-step mechanism commonly proposed for the oxidation of NADH at chemically modified electrodes. Example of a screen printed electrochemical cell (screen image) showing the Reference element (A), the working electrode (B) and the counter electrode (C). Calibration curve (B) for the diaphorase based ketone sensor developed and manufactured in house (unpublished results).
Examples of commercially available test strips for self-testing utilising dehydrogenases.Table 1.
IntroductionSelf-monitoring blood glucose (SMBG) is an important component of modern therapy for diabetes mellitus. A slim 29-year-old, Etherington had no risk factors for the disease, which occurs when a woman’s pancreas isn’t able to produce the extra insulin needed to keep increased blood sugar in check during pregnancy. Left unchecked, high blood sugar levels increase the risk of developing pre-eclampsia, which is life-threatening to both mom and baby. She knew she needed to adopt a healthy eating plan and incorporate more exercise into her routine to help manage her glucose levels. Through trial and error, she found that potatoes, her favourite food, spiked her glucose levels so much that she had to cut them out completely, along with her beloved pasta and baguettes. Having many risk factors for GDM (she’s over 35, had GDM in her first pregnancy and has a family history of diabetes), Peel was expecting the diagnosis, even though she had adopted a diabetes-friendly diet and boosted her exercise level early on. The disease disappeared within weeks after the birth (as it does for almost all women with GDM), but she and her children are at an increased risk of developing type 2 diabetes later in life; one in five women who had GDM will be diagnosed with type 2 diabetes within nine years of giving birth, and their children are six times more likely to develop the disease than their peers.
In this context, the mediator reacts with NADH in a chemical step to oxidise it (a) and is itself reduced. The mediator and NADH form a charge transfer complex that dissociates to give rise to the reduced mediator and biologically active NAD+.4.
Comercially available screen printed electrodes for research purposes can be obtained from DropSens, Edificio CEEI - Parque Tecnologico de Asturias - 33428 Llanera (Asturias) Spain. The test strip incorporates gold and palladium electrodes that are orientated in a cofacial manner.
SMBG has been recommended for people with diabetes and their health care professionals in order to achieve a specific level of glycaemic control and to prevent hypoglycaemia. Since GDM has no real symptoms (common complaints of people with diabetes, such as tiredness and frequent urination, are often normal parts of pregnancy), she had no warning signs. As well, women with GDM may give birth to very large babies, which can get stuck in the birth canal and require intervention, such as a C-section.
In fact, one Ontario-based study by the Institute for Clinical Evaluative Sciences and Mount Sinai Hospital found the rate of GDM and pre-GDM (diabetes that existed prior to the pregnancy) doubled from 1996 to 2010, largely due to lifestyle changes. To ease the transition, she attended free diabetes courses at Women’s College Hospital in Toronto, where she learned about nutrition, obtained a glucose meter, and was taught how to prick her finger and interpret the results.
Because Peel’s blood sugar rose while she slept, no matter how perfect it was during the day, she had to give herself insulin injections every night before bed.
As a result, both Etherington and Peel are maintaining their healthy lifestyles, and they both schedule annual checkups to ensure they keep the diabetes at bay.
The electrochemical reduction of 1,10-phenanthroline-5,6-quinone, like other quinones, is reversible and occurs by 2e- transfer in a single step in aqueous solution and by two 1 e-transfer steps in aprotic media.7. The dimensions of the two electrodes are defined and controlled during the manufacturing process by a die-punch process. The goal of SMBG is to collect detailed information about blood glucose levels at many time points to enable maintenance of a more constant glucose level by more precise regimens.
The keys were eating well, resisting temptation (parties were tough) and exercising frequently (she favoured going for brisk walks and dancing).
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Earlier this year Harvard researchers found consuming 50g of processed red meat per day increased the risk of type 2 diabetes by 50 per cent. When the electrode is modified, in this case with o-amino phenol the oxidation of NADH is concomitant with the oxidation of the immobilised redox couple (traces b – d).
The chemical oxidation of NADH by this mechanism bypasses the problems associated with the direct oxidation at a bare electrode surface.3. The electrodes are separated from each other by a thin plastic spacer that has a nominal thickness of 95 ?m.
In fact, if women with GDM are properly monitored and work to keep their blood sugar levels in check, most go on to have healthy pregnancies. Peel even found ways to work well-loved foods into her diet: When craving a hamburger, she ate it open-faced and opted for a salad instead of fries. If you can't find this email, access your profile editor to re-send the confirmation email. The glucose-sensitive reagents, citraconate buffer salts, potassium ferricyanide mediator, and flavoprotein glucose dehydrogenase, are laid down on the “bottom” palladium electrode. This is because if untreated hypoglycaemia can result in coma for diabetic patients and is therefore a condition that diabetics try to avoid through proper action.
The user can apply blood to the test strip either from the right hand side or the left hand side.
To this end, regular testing can predict dangerous drop in blood glucose concentration which can lead to hypoglycaemia.
Where this is the case, the following disadvantages may outweigh the potential benefits:anxiety about one’s blood sugar control and state of healththe physical pain of finger prickingexpense to the NHS or other medical bodyNumerous trials have been carried out to determine the true impact of SMBG on glycaemic control. Among patients with type 1 diabetes, SMBG has been associated with improved health outcomes.
The test most commonly involves pricking a finger with a lancet device to obtain a small blood sample, applying a drop of blood onto a reagent strip (typically an enzyme electrode), and determining the glucose concentration by inserting the strip into an electronic meter for an automated reading.
Test results (a measure of the glucose concentration in the blood sample) are then recorded either in a logbook or stored in the glucose meter’s electronic memory. People with diabetes can be taught to use their SMBG results to correct any deviations out of a desired target range by changing their carbohydrate intake, exercising, or using more or less insulin.The frequency with which patients with diabetes should monitor their blood glucose level varies from person to person.
Most experts agree that insulin-treated patients should monitor blood glucose at least four times a day, most commonly fasting, before meals, and before bed.
In addition, patients using insulin can benefit by obtaining postprandial blood glucose readings to help them more accurately adjust their insulin regimen. A positive correlation between frequency of SMBG and glycaemic control among patients with insulin-treated type 1 or type 2 diabetes has been demonstrated. If a patient is on a stable oral regimen with HbA1c concentration within the target range, infrequent SMBG monitoring is appropriate. In such cases, patients can use SMBG data as biofeedback at times of increased stress or changes in diet or physical activity. One factor that is often overlooked is that the numbers obtained by testing are only one part of the picture, which requires additional data to be complete. The patient’s exercise regime must also be considered, as well as when and how much medication has been taken. The meter - applies potential differences in a programmed sequence to the sensor, collects current data and analyses the current time response of the sensor, records and displays results.

The enzyme electrode (or biosensor) test strip - collects the blood sample, the sample undergoes an enzymatic chemical reaction followed by an electrochemical detection step. The patient simply inserts the enzyme electrode into the meter and applies a small drop of blood to the sensor. Blood is a complex fluid and glucose levels vary widely over time in a single patient, many factors besides glucose vary in blood from healthy, patients (haematocrit, oxygen levels, and metabolic by-products) therefore great specificity is a prime requirement. In addition, patients with diabetes may have a wide range of other medical problems creating even greater variation in their blood.
Finally, biosensors can be used directly in the blood without requiring major modifications to the biological sample (increased temperature or pressure, dramatic pH changes, addition of highly reactive chemicals, etc).
The enzyme electrodes commonly used in SMBG can be defined as a combination of any electrochemical probe with a thin layer of enzyme based reagent that is selective for glucose (or other important analytes such as ?-hydroxybutyrate). In these devices, the function of the enzyme is to provide selectivity by virtue of its biological affinity for a particular substrate molecule. For example, an enzyme is capable of catalysing a particular reaction of a given substrate even though other isomers of that substrate or similar substrates may be present.
Typically, the progress of the enzyme reaction (which is related to the concentration of analyte) is monitored by the rate of formation of product or the disappearance of a reactant. If either the product or reactant is electroactive, then the progress of the reaction can be monitored directly using amperometry. The resultant current is monitored by the meter and then interpolated into an accurate measurement of glucose using on-board software algorithms giving the user a concentration value in typically less than 7 seconds.The final method of analysis used will ultimately depend on several properties of the enzyme. The main considerations are;does the enzyme contain redox active groupsare the products of the biochemical reaction electroactiveis one of the substrates or cofactors electroactivewhat is the required speed of responsewhat will be the final application of the sensorThe answer to the first three criteria will depend largely on the system under investigation. The answer to the latter three depends on the requirements and application of the sensor under consideration.
If the enzyme does not contain any redox groups, then the method of analysis will be restricted to monitoring either the release of products or the consumption of substrate by their reaction at the transducing electrode.
Of particular relevance to this article, are the devices that incorporate nicotinamide adenine dinucleotide (NAD), flavin adenine dinucleotide (FAD) or pyrroloquinoline quinone (PQQ) dependent dehydrogenase enzymes.2.
Electrochemical oxidation of NAD(P)HGiven that the nicotinamide coenzymes are electron carriers, and therefore by definition electroactive, it would appear at first sight (points 2 and 3 above) that these systems would be ideal candidates for commercial enzyme electrode devices.
The electrochemical oxidation of NADH to NAD+ is however both chemically and kinetically complicated at common electrode surfaces such as gold, platinum or glassy carbon.The electrochemical detection of NADH has generated great interest because the pyridine nucleotides NAD+ and NADP+ are ubiquitous in all living systems and are required for the reactions of more than 450 oxidoreductases.
SCE [8, 9] significant over-potential is often required for the direct oxidation of NADH at bare electrodes. The large amount of energy required to produce these intermediates is the origin of the large overpotential (typically 1 Volt) required at bare electrodes. As a result, the direct electrochemical oxidation of NADH has been shown to produce a mixture of products including biologically active NAD+, (NAD)2 dimers and products from the side reactions of the electrogenerated NAD. In addition, the direct oxidation of NADH is often accompanied by electrode fouling due to the polymerisation oxidation products on the electrode surface. Both of these processes are irreversible and result in the gradual blocking of the electrode during continued oxidation.
It is this electrode fouling that results in the irreproducibility of the analytical signal from bare electrodes. In addition, if this method were to be used in commercial glucose sensors for SMBG, the high over potentials required would result in the response being a combination of signals from the oxidation of both NADH and common interferents, e.g. These molecules can transfer charge from the reduced coenzyme directly to the electrode thereby bypassing the direct oxidation of NADH. The use of this approach has three potential advantages when compared to the direct oxidation of the reduced nicotinamide.
First, by judicious choice of the mediator problems associated with electrode fouling or competing reactions can be avoided (especially true if he chosen mediator has a site for hydride transfer). Second, the rate of electron transfer between the NADH molecule and the mediator can be enhanced resulting in a more stable increased signal. The rate at which NADH is chemically oxidised will governed to a large extent by the potential difference between the formal redox potential of the two species.
Finally, a mediator can be selected with a redox potential that will limit the effects of interference. The electrocatalytic oxidation of NADH at a modified electrode is shown schematically in Figure 1. These show the direct oxidation of NADH (e) at the glassy carbon electrode and the cyclic voltammogram (a) of the immobilised species. The chemical oxidation of NADH by this mechanism bypasses the problems associated with the direct oxidation at a bare electrode surface.To design an NADH sensor, the mediator is normally immobilized on the electrode surface or within the electrode material. The method that is ultimately chosen to produce the modified electrode depends upon the method of mass production (for commercial sensors) and the materials used in the device. For example, in sensors that utilise screen printed carbon based electrodes (screen-printing technology is a kind of low-cost thick film technology which allows to deposit thick films, a few to hundreds micrometers and is well suited for mass production and portable devices. Such a micro fabrication route offers high-volume production of extremely inexpensive and yet highly reproducible disposable enzyme electrodes – this will be discussed further in the text) it is convenient to incorporate the mediator directly into the carbon ink particularly if the mediator contains delocalised aromatic rings, as found in quinine and phenoxazine dye based mediators, which form strong chemisorbed bonds with the carbon and graphite plates. The mediator loading, activity stability etc can all be investigated using conventional electrochemical techniques such as DC cyclic voltammetry. Kinetic modelling of NADH oxidation at chemically modified electrodes.In the scientific literature, NADH oxidation at chemically modified electrodes is most commonly suggested to occur via a two-step reaction mechanism. In the first step, NADH forms a charge transfer complex with the oxidised form of the mediator bound to the electrode surface. In the second step, electron exchange takes lace and the complex breaks down producing NAD+ and a reduced mediator site. Because the electrode is polarised, the reduced mediator site is reoxidised in a non-rate limiting electron exchange to the bulk electrode material. Figure 3.Two-step mechanism commonly proposed for the oxidation of NADH at chemically modified electrodes. This type of model assumes that the rate of electron transfer between the mediating species in the film and the NADH is sufficiently fast so as not to be rate limiting, the NADH freely diffuses into the film whereupon it adsorbs to the catalytic site and it undergoes oxidation to NAD+.
Also, the expression for ik is valid only for thin films where the concentration of NADH is insufficient to saturate the binding sites.[12]Due to the formation of the charge-transfer complex, this reaction scheme is commonly analysed using Michaelis-Menten kinetics.
From figure 3 it is possible to construct the following kinetic argument.This is now a straight line plot. Values for kObs can be obtained from Koutecky-Levitch plots under steady state oxidation conditions as described by Compton and Hancock.[12] Typically, values of kObs tend to be in the range 10-3 to 10-1 cm s-1. Exogenous NAD+ is incorporated into the reagent ink which is deposited onto the individual electrode by screen printing. The mediator molecule that is used to recycle the reduced form of the coenzyme is phenanthroline quinone (Manufacturers own vial insert data sheet). Advantages often cited with this type of chemistry include good selectivity and no reaction with oxygen.5.
Screen printing as a means of mass-manufacturing enzyme sensorsScreen printing is arguably the most versatile of all printing processes.
It can be used to print on a wide variety of substrates, including paper, paperboard, plastics, glass, metals, fabrics, and many other materials including paper, plastics, glass, metals, nylon and cotton. Some common products from the screen printing industry include posters, labels, decals, signs, and all types of textiles and electronic circuit boards. The advantage of screen printing over other print processes is that the press can print on substrates of any shape, thickness and size.A significant characteristic of screen printing is that a greater thickness of the ink can be applied to the substrate than is possible with other printing techniques. This allows for some very interesting effects that are not possible using other printing methods. Because of the simplicity of the application process, a wider range of inks and dyes are available for use in screen printing than for use in any other printing process. The major chemicals used include screen emulsions, inks, and solvents, surfactants, caustics and oxidizers used in screen reclamation. The screen printing process uses a porous mesh stretched tightly over a frame made of wood or metal.
The stencil defines the image to be printed in other printing technologies this would be referred to as the image plate.Screen printing ink is applied to the substrate by placing the screen over the material. Ink is then forced through the fine mesh openings using a squeegee that is drawn across the screen, applying pressure thereby forcing the ink through the open areas of the screen. Ink will pass through only in areas where no stencil is applied, thus forming an image on the printing substrate. The diameter of the threads and the thread count of the mesh will determine how much ink is deposited onto the substrates. Figure 4 shows an example of an image for an electrochemical cell consisting of a working, a counter and a reference element. At one time most blades were made from rubber which, however, is prone to wear and edge nicks and has a tendency to warp and distort. While blades continue to be made from rubbers such as neoprene, most are now made from polyurethane which can produce as many as 25,000 impressions without significant degradation of the image.If the item was printed on a manual or automatic screen press the printed product will be placed on a conveyor belt which carries the item into the drying oven or through the UV curing system. Air drying of certain inks, though rare in the industry, is still sometimes utilized.The rate of screen printing production was once dictated by the drying rate of the screen print inks. Some specific innovations which affected the production rate and have also increased screen press popularity include:Development of automatic presses versus hand operated presses which have comparatively slow production times.
Flat-bed and cylinder presses are similar in that both use a flat screen and a three step reciprocating process to perform the printing operation. The screen is first moved into position over the substrate, the squeegee is then pressed against the mesh and drawn over the image area, and then the screen is lifted away from the substrate to complete the process. With a flat-bed press the substrate to be printed is positioned on a horizontal print bed that is parallel to the screen. The open-ended cylinders are capped at both ends and fitted into blocks at the side of the press. During printing, ink is pumped into one end of the cylinder so that a fresh supply is constantly maintained.
The squeegee is a free floating steel bar inside the cylinder and squeegee pressure is maintained and adjusted by magnets mounted under the press bed.
Screen printing inks are moderately viscous inks which exhibit different properties when compared to other printing inks such as offset, gravure and flexographic inks though they have similar basic compositions (pigments, solvent carrier, toners, and emulsifiers). There are five different types of screen ink to include solvent, water, and solvent plastisol, water plastisol, and UV curable. UV curable inksUV curable inks consist of liquid pre-polymers, monomers, and initiators which upon being exposed to large doses of U.V. They also require less energy, overall, to dry or "cure" compared to gas or electric driers.

The down side of UV inks is they can cost as much as three times that of regular inks and must be handled differently than conventional inks due to safety issues. Plastisol InksPlastisol inks (both solvent and water based) are used in textile screen printing. Plastisol ink is a PVC (polyvinyl chloride) based system that essentially contains no solvent at all. Along with UV ink used in graphic screen printing, it is referred to as a 100% solid ink system. Plastisol is a thermoplastic ink in that it is necessary to heat the printed ink film to a temperature high enough to cause the molecules of PVC resin and plasticizer to cross-link and thereby solidify, or cure.
Solvent inks & water inksSolvent and water based screen printing inks are formulated with primarily solvent or water.
Water based inks, though they contain significantly less, may still emit VOC's from small amounts of solvent and other additives blended into the ink.
Water-based inks also have the advantage of being an excellent ink system for high speed roll-to-roll yardage printing.
Such printing is done on large sophisticated equipment that has very large drying (curing) capacity. Finally, because of the fragility of the components used in the manufacture of enzyme electrodes, e.g.
First the customer provides the screen printer with objects, photographs, text, ideas, or concepts of what they wish to have printed. The printer must then transfer a "picture" of the artwork (also called "copy") to be printed into an "image" (a picture on film) which can then be processed and eventually used to prepare the screen stencil. Once the artwork is transferred to a positive image that will be chemically processed onto the screen fabric (applying the emulsion or stencil) and eventually mounted onto a screen frame that is then attached to the printing press and production begins.
Each colour represents a different layer requiring different screens and artworks.Enzyme electrodes are normally built up in layers using different art works and different inks. The blood monitors (the systems that the user receives) are made up of three main parts: the Optium meter, the Optium Plus blood glucose electrodes, and the Optium blood ?-Ketone electrodes. When the blood sample is applied to the electrode, the glucose or ketone (?-hydroxybutyrate) in the blood reacts with the chemicals on the electrode.
This reaction produces a small electrical current that is measured and the result is displayed by the sensor. Optium Xceed monitors are designed for testing blood obtained from a finger prick, but you can also use it to test blood from other, less painful, sites such as the base of the thumb, forearm or upper arm.
A recent study published in Clinical Chemistry and Laboratory Medicine [13] concluded that the Optium Blood glucose test strip had a within-run imprecision coefficient of variation (CV) of 4.2%. The authors concluded that meter had good precision and accuracy when compared to the laboratory method and met the quality recommendations of the National Committee of Clinical Laboratory Standards (NCCLS, currently the Clinical Laboratory Standards Institute), the National Academy of Clinical Biochemistry (NACB) and the International Organization for Standardization (ISO).6.
Ketone sensors which incorporate a NAD-linked dehydrogenaseIn addition to glucose, another important analyte which is of particular relevance to diabetic patients is the ketone body, ?-hydroxybutyrate. Diabetic ketoacidosis (DKA), a condition were the level of ketone bodies in the blood are elevated, is a problem that occurs in people with diabetes. It occurs when the body cannot use sugar (glucose) as a fuel source because there is no insulin or not enough insulin. People with type 1 diabetes do not have enough insulin, a hormone the body uses to break down sugar (glucose) in the blood for energy. When glucose is not available, fat is broken down instead.As fats are broken down, acids called ketones build up in the blood and urine. Diabetic ketoacidosis is often the first sign of type 1 diabetes in people who do not yet have other symptoms.
Infection, injury, a serious illness, or surgery can lead to diabetic ketoacidosis in people with type 1 diabetes.
In ketoacidosis, the body fails to adequately regulate ketone production causing such a severe accumulation of keto acids that the pH of the blood is substantially decreased.
14 Despite considerable advances in diabetes therapy, key epidemiological figures related to DKA remained nearly unchanged during the last decades at a global level. Prevention of DKA – especially in sick day management – relies on intensive self-monitoring of blood glucose and subsequent, appropriate therapy adjustments.
Self-monitoring of ketone bodies during hyperglycemia can provide important, complementary information on the metabolic state.
Both methods for self-monitoring of ketone bodies at home are clinically reliable and there is no published evidence favouring one method with respect to DKA prevention.Ketone sensitive test strips can be manufactured using NAD-linked enzyme, ?-hydroxybutyrate dehydrogenase and chemically modified electrodes like the ones described above.
It is interesting to note however that in the case of this particular enzyme representatives of the common classes of quinoid NADH redox mediator, including Meldola Blue, 4-methyl-1,2-benzoquinone, 1-methoxy phenazine methosulphate and 2,6-dichloroindophenol, were shown to inhibit the NAD-dependent enzyme ?-hydroxybutyrate dehydrogenase, severely limiting their utility in the construction of a stable biosensor electrode for the ketone body ?-hydroxybutyrate.
The electrochemical reduction of 1,10-phenanthroline-5,6-quinone, like other quinones, is reversible and occurs by 2e- transfer in a single step in aqueous solution and by two 1 e-transfer steps in aprotic media.This technology resulted in the launch of the MediSense® Optium™ ?-Ketone electrode.
The strip was stable, (?10% loss in response at 30 C versus 4 C) with a long shelf life of 18 months.
Typical acceptor molecules include dyes such as 2, 6- dichlorophenolindophenol and tetrazolium dyes and redox couples such as ferricyanide anions and ferricinium cations.
The test time was 7 seconds.In this configuration, NAD+, Diaphorase, acceptor molecule and ?-hydroxybutyrate dehydrogenase are all formulated together in the enzyme ink and laid down on the test strip using an appropriate manufacturing method.
Data generated in-house (unpublished results) using this kind of prototype test strip (with ferricyanide as the acceptor) is shown below for the clinically relevant concentration range of ?-hydroxybutyrate.
Ferricyanide, buffer salts, NAD+, binders, diaphorase and ?-hydroxybutyrate dehydrogenase were mixed into a suitable enzyme ink and printed onto carbon electrodes. Glucose test strips using PQQ linked and FAD-linked glucose dehydrogenasePQQ-GDH (pyrrolo quinoline quinone glucose dehydrogenase) belongs to a class of enzymes called quinoproteins which require ortho-quinone cofactors to oxidize a wide variety of alcohols and amines to their corresponding aldehydes and ketones.
One monomer (50 kDa, 454 residues) has been reported to bind one PQQ molecule and two Ca(II) ions.
One of the Ca(II) ions is required for activation of the cofactor; the other is needed for functional dimerisation of the protein.
The natural electron acceptor of PQQ-GDH is ubiquinone although the enzyme will react with a variety of artificial acceptors such as the ferricyanide anion ion and the ferricinum cation. The oxidised form of PQQ can be converted into the reduced form PQQH2 by the transfer of 2 electrons and two protons from the substrate molecule. One is intracellular and soluble (sPQQ-GDH) whereas the other molecule is insoluble and firmly bound to the outer surface of the cytoplasmic membrane (mPQQ-GDH).
For these reasons, mPQQ-GDH has not been successfully commercialised for biosensor application.
It can oxidise a number of monosaccharides, in addition to glucose, such as maltose, mannose and lactose.
Consequently, patients that have high levels of, for example maltose in the blood (which could result as a side effect of peritoneal dialysis) or have an inbred genetic disorder resulting in impaired carbohydrate metabolism, would obtain an inaccurate high reading when testing with glucose electrodes incorporating this enzyme. The increasing demand for dialysis and slower growth in capacity for haemodialysis has reinforced the need for an integrated approach to providing dialysis. Peritoneal dialysis is the preferred option for a proportion of patients with end stage renal failure. Peritoneal dialysis fluid usually contains glucose as an osmotic agent to enable water to pass across the peritoneum.
Some patients lose the osmotic effect of glucose quickly, but large icodextrin molecules, which are not easily transported across the peritoneal membrane, maintain an osmotic gradient. Icodextrin is not metabolised in the peritoneal cavity, but the polymer can move into the blood stream via the lymphatic system.
During systemic circulation, icodextrin is mainly metabolised into maltose which accumulates due to a lack of circulating maltase. It is the accumulation in the systemic circulation of these metabolites of icodextrin that may lead to the disparity between finger stick and formal blood glucose measurement. Maltose interferes with glucose assays that use glucose dehydrogenase with cofactor pyrroloquinolinequinone (PQQ-GDH) leading to falsely increased readings.Because of the oxygen insensitivity of PQQ-GDH however, there is much commercial interest in producing a mutant form of the enzyme that retains its non-reactivity to oxygen but improving its specificity with respect to D-glucose. Recently, Roche have reported the successful production of a mutant strain of PQQ-GDH which shows no cross-reactivity with maltose. Whilst flavoproteins such as glucose oxidase exchange electrons with an electrode surface via small molecular weight mediators such as ferrocene, ferricyanide etc, PQQ containing enzymes can exchange electrons via cytochrome b562. This cytochrome will exchange electrons directly with the electrode surface without the need for any mediator molecule. As its name suggests, the enzyme catalyses the oxidation of glucose but does not utilise dioxygen as a co-reactant.
It can react with a number of artificial electron acceptors such as the ferricyanide anion and the ferricinium cation. The enzyme, which is isolated from Aspergillus terreus shows good thermal stability and high selectivity for glucose. Its absorption spectrum is typical of flavoproteins showing two distinctive peaks corresponding to the oxidised flavin cofactor at 465nm and 385 nm. In the presence of glucose the enzyme is bleached and these characteristic absorption bands disappear.
The test strip incorporates gold and palladium electrodes which are orientated in a co-facial manner.
The electrodes are separated from each other by a thin plastic spacer that has a nominal thickness of 95 µm.
The glucose-sensitive reagents, citraconate buffer salts, potassium ferricyanide mediator and flavo-protein glucose dehydrogenase (FAD-GDH), are laid down on the ‘bottom’ palladium electrode. FAD-GDH enzyme was selected for its high substrate-specificity and non-reactivity towards oxygen. The strip may be defined as a ‘side-fill strip’ because blood may be applied to the 400 nL sample chamber from either the left or the right side of the test strip (Figure 7). The glucose level is reported within 5 seconds of the start of the test procedure.Manufacturing process controls and built-in signal processing compensation mechanisms eliminate the need for user calibration coding, thus reducing the potential for user error. The meter uses a novel multi-pulse signal and has an improved glucose-Hct-temperature-antioxidant compensation algorithm for higher accuracy and precision over a wide range of blood samples.
The OneTouch Verio BG test strip is designed to work with meters that are technically equivalent but have different user interfaces.

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  1. 16.01.2014 at 10:44:29

    Tools to find out how your.

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  2. 16.01.2014 at 17:57:11

    Measured six to eight hours after your pancreas.

    Author: PRESIDENT