SENSORS FOR MEASURING ANALYTE
CONCENTRATIONS AND METHODS OF
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates generally to electrochemical sensors and, more particularly, to enzyme catalyzed electrochemical sensors including lactate sensors.
2. Field of the Invention
Under normal conditions, glucose is metabolized to pyruvate which, in turn, is metabolized to carbon dioxide and water with little or no lactate production. Therefore, in a healthy human, the concentration of lactate in blood should be below some critical level. However, if there is a metabolic problem, the level of lactate in the blood may increase above mis critical level. In certain systems, it may be desirable to measure the lactate concentration in other biological fluids such as plasma, sweat, tears or urine.
A high level of lactate in blood can indicate a lack of oxygen in the blood resulting from a variety of problems such as circulation shock, a fatal disease with a high mortality rate. Circulation shock occurs when the heart cannot distribute enough blood through the body, and typical causes for this condition are hemorrhaging, dehydration, heart attack, sepsis and certain other infections.
One traditional method of measuring lactate concentrations in blood include reacting lactate with NAD+ to form pyruvate and NADH and subsequently making a spectrcphotometric determination of the NADH concentration, which serves as a measure of the lactate concentration. This technique, however, is relatively slow, inefficient and labor intensive.
In a clinical setting, accurate and relatively fast determinations of lactate levels can be determined from blood samples utilizing electrochemical sensors. Conventional sensors are fabricated to be large, comprising many serviceable parts, or small, planar-type sensors which may be more convenient in many circumstances. The term "planar" as used herein refers to the well-known procedure of fabricating a substantially planar structure comprising layers of relatively thin materials, for example, using the well-known thick or thin-film techniques. See, for example, Liu et al., U.S. Pat No. 4,571,292, and Papadakis et at, U.S. Pat. No. 4,536,274, both of which are incorporated herein by reference.
In the clinical setting, it is a goal to maximize the data obtainable from relatively small test sample volumes (microliters) during chemical blood analysis. Fabrication of a sensor sample chamber for holding a blood sample in contact with a sensor is desirable in this regard so that many determinations may be simultaneously performed on a test sample, for example, using a series of interconnected sensors, each constructed to detect a different analyte, from a small test sample volume. However, as a sample chamber is made smaller, the concentration of contaminants in a sample, as those released from sensor components themselves, especially components denning the sample chamber, and/or certain reaction products of the sensor itself is increased. Such contamination may result in premature sensor failure.
Lactate electrode sensors include an enzyme-oontaining layer which converts lactate to reaction products including hydrogen peroxide according to the following reactions:
[lactate oxidase] H2O2 ^02+2H++2e
In these reactions, lactate reacts with oxygen to form hydrogen peroxide. A suitable electrode can then measure the formation of hydrogen peroxide, as an electrical signal.
10 Current is generated as a result of peroxide oxidation, and under suitable conditions the current is proportional to the lactate concentration.
Numerous devices for determination of lactate have been described, however most of them have some limitation with
15 respect to reproducibility, speed of response, test same volume, number of effective uses, and the range of detection. Some existing commercial methods rely on utilization of hydrogen peroxide measurement as outlined above. Some known enzyme electrodes have a two membrane
20 system. In these electrodes, the blood, including lactate and certain interferants, diffuses through a primary membrane of the sensor. Certain blood components then reach a second membrane and interact with an enzyme, such as lactate oxidase, that catalyzes the conversion of lactate to hydrogen
25 peroxide. The hydrogen peroxide may diffuse back through the primary membrane, or it may further diffuse through the second membrane to an electrode where it can be reacted to form oxygen and a proton, resulting in a current proportional to the lactate concentration.
30 The electrode's membrane assembly serves several functions, including selectively allowing the passage of lactate therethrough, providing a location between the primary and secondary membranes for an enzyme to catalyze the reaction of lactate and oxygen passing through the
35 primary membrane, and allowing only hydrogen peroxide through the secondary membrane to the electrode.
A single-layered electrode membrane was described by lones in EP Patent No. 207 370 Bl. This reference is directed to an electrochemical sensor including three pri
40 mary components: a metal electrode, a reactive layer of immobilized enzyme directly on an anode, and a singlelayered membrane. The membrane is formed from a dispersion of a polymerizable silicon-containing compound applied in an incompletely cured form, having a liquid
45 carrier which is essentially insoluble in the dispersed phase and removable from the dispersion during curing.
It has been found, however, that the single membrane layer disclosed in EP 207,370 Bl minimizes only anionic interfering substances, such as ascorbic acid and uric acid,
50 from passing therethrough. Neutral species, such as acetaminophen, can diffuse through the membrane and influence the sensor's sensitivity and accuracy.
As noted above, enzyme electrodes convert lactate or glucose into hydrogen peroxide, which can be reacted to
55 produce a current proportional to the lactate or glucose concentration. Enzyme electrodes adapted to measure other analytes have also been described in the art. An enzyme electrode having an electrically conductive support member which consists of, or comprises, a parous layer of resin
60 bonded carbon or graphite particles is disclosed by Bennetto et al., in U.S. Pat. No. 4,970,145. The carbon or graphite particles have a finely divided platinum group metal intimately mixed therewith, to form a porous, substantially homogeneous, substrate layer into which the enzyme is
65 adsorbed or immobilized. The preferred substrate materials are resin bonded, platinized carbon paper electrodes, comprising platinized carbon powder particles bonded onto a