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Technical Bulletin: The Advantages and Pitfalls of Reagent Strips in Dialysis, Other Medical, and Non-Medical Applications
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David A. Morris, Ph.D. and Joe D. Sweazy
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Introduction
Every day we see around us clever devices that make our lives and jobs easier. The .reagent strip or test strip is a simple device that revolutionized the way laboratory assays are done, and its user-friendly attributes allow technical and non-technical persons to conduct relatively sophisticated assays. Test strips typically have a plastic handle with a reagent area at one end.
We use test strips to provide rapid assays of aqueous or other fluid mixtures, including body fluids. In a typical assay, you dip the reagent area into a sample, remove it, and compare the color of the reagent area with a color chart. Some test strips work by presence/absence of a color change at a threshold concentration, or by measuring a color or electrochemical change with a meter.
"A Complex Mixture"
Modern test strips have a complex mixture of chemicals that can include enzymes, polymers, surfactants, buffers and indicators. The chemicals control the ability of the reagent area to pick up sample, enhance the color intensity, provide stability, reduce the tendency of the colors to run off the pad, and enhance other attributes of a test strip.
In this paper we look at the history of test strips through their impact on personal diabetes control, and on the revolution they caused in urine analysis. We also describe the wide array of environmental and industrial applications of test strips. Finally, we look at some test strips used in dialysis centers; we present some accuracy and precision data and identify the major pitfall encountered by users of these convenient easy-to-use devices.
Impact on Diabetes Control
The utility of test strips has been universally demonstrated by its role in monitoring blood sugar levels. The U.S. government conducted a landmark study from 1983 to 1993 that profoundly impacted the management of diabetes (1). That study involved 1,441 volunteers with Type 1 diabetes at 29 medical centers in the United States and Canada. The United Kingdom Prospective Diabetes Study in 1998 produced similar results for 5,102 Type 2 diabetes patients at 23 clinical centers (2-6).
The studies proved that the level of blood sugar control predicts the onset and severity of diabetes-related complications for both types of diabetes. This means that if you have diabetes, and if you can keep your blood sugar levels as close as possible to normal, you can live a normal life span with few or even no complications at all. You will lower your chances of having eye disease, kidney disease, nerve damage and heart disease.
To reach tight control, one must test his or her blood sugar levels up to several times each day. Test strips and handy pocket-sized meters made it possible to easily and conveniently monitor blood glucose and maintain tight control (7–9). The market for blood glucose meters and test strips has grown into a worldwide market that was expected to exceed 3 billion dollars by the year 2000 (10).
Some other examples of where test strips have been applied for analysis of blood are listed in Table 1. (See the bottom of the page.)
Impact on Urine Analysis
For over half a century, urine analysis using test strips has been used for health status screening. Almost all urine test screening is done in a professional setting, although there have been attempts to market urine strips as a “wellness status test” for home use. Some urine test strips measure up to ten parameters at once on a single strip. With a simple dip-and-read multi-test strip, the health professional gets the ten results listed in the urine analysis row of Table 1. The complete analysis takes a little over a minute.
The ease-of-use and speed of the multi-test strip can be contrasted with the much longer time required to conduct separate laboratory analyses for the each of the ten different urine parameters. Abnormal results of key parameters indicates a need for additional testing to confirm possible liver or kidney problems, possible incipient infection in the bladder or urinary tract, and other problems. Before test strips, a clinical lab would batch urine samples until a lab technician was free to conduct the myriad of tests on the now aging and “fragrant” samples. Since the advent of test strips, the patient presents the sample cup at the sample station at the doctor’s office and a test strip is used to get a complete urine profile of a fresh sample right away.
Non-Medical Applications
Before turning to the application of test strips in Dialysis Centers, it is instructive to point out that test strips have found their way to a variety of applications in the non-medical field. The convenience, ease of use and accuracy that make test strips attractive for blood and urine analysis have also led to their use in these areas.
Table 2 lists some test strip applications in such diverse areas as automotive fluid, aquarium, environmental and industrial testing, soil testing by garden enthusiasts, swimming pools and spas. The wide range of parameters that can be measured with test strips is also shown in the table.
Test Strip Use in Dialysis Centers
Reagent strips have been used to meet testing needs in hemodialysis operations for more than 10 years. A schematic drawing of a typical dialysis operation showing possible testing locations in the water treatment area, the dialysis machine area and the dialyzer disinfection area, is shown in Figure 1.
Figure 1 is a general example. Testing routine in hemodialysis centers may vary from the testing shown in Figure 1. Reagent strips that are commonly used in a hemodialysis center are shown in Table 3, along with typical concentration ranges.
Reagent strips that measure disinfectant levels and key parameters of dialysate solution require Section 510(k) pre-market notification to the U.S. Food and Drug Administration (FDA). The manufacturer must practice current Good Manufacturing Practices (cGMP) as set forth in the Quality System Regulation (QS) for Medical Devices: General Regulation (21 CFR Part 820).
Avoiding the Pitfalls
The convenience and ease of use of test strips can lead a careless user to ignore the use instructions for the specific test strip that he or she is using. This is the major pitfall, and can lead to incorrect results. There are two things to keep in mind: the activation method and the read time. These are given on the bottle label and in the product insert.
It is important to use the required activation mode for the strip you are using. Different strips require dipping the strip in the sample, or swishing the strip back and forth in the sample, or holding the reagent area in a stream of sample. Using the wrong activation method for that strip may lead to incorrect results.
Two examples of use instructions for different reagent strips are given in Table 4. While both reagent strips have directions for testing is a stream, the Water Hardness reagent strip requires 2 seconds in the stream and comparison with color chart at 15 seconds (See “Directions in Stream”), but the Total Chlorine reagent strip requires 5 seconds in the stream followed by an immediate comparison with the color chart (See “Directions for Qualitative Results”).
"Testing in a Sample Cup"
Both reagent strips also have directions for testing in a sample cup. The Water Hardness reagent strip requires dipping the reagent area in the sample for 5 seconds, and then comparing with the color chart 15 seconds after removing the reagent pad from the sample (See “Directions in Sample”). The Total Chlorine reagent strip calls for moving the strip back and forth gently in the sample for 15 seconds, then comparing the pad to the color chart immediately after removal from the sample (See “Directions for Quantitative Results”).
Test strips are designed to minimize the effect of variation of use instructions by the application of tools like Failure Mode and Effects Analysis (FMEA). However, the best results are obtained by the user following the directions exactly. The take-away message is to read and follow the directions on the bottle label and in the product insert.
The Origin of Test Strip Color
Test strips change colors because the presence of the specific parameter in the sample causes a chemical reaction that leads to a change in the color of one or more of the chemicals in the reagent area of the strip. When we say an object is colored, we are observing the selective absorption of wavelengths of incident light by molecules on the surface of the object (11).
For example, a blue object absorbs wavelengths that are complementary to blue, and reflects only the blue portion of the light. The energy is actually absorbed by electrons that are free to roam within the molecules of the blue object. When light shines on the electrons they become excited or energized. The energized electrons undergo energy level transitions and absorb selective energy at certain wavelengths depending on the nature of the molecule.
When the chemical nature of compound is changed due a chemical reaction, the possible energy transitions of the electrons can be different in the new compound. As a result, an indicator that is blue before a chemical reaction may now be green when the indicator is transformed due to a chemical reaction.
Visual Response to Colors
Perception of color begins with light entering the eye and falling on the light sensitive cells, namely rods and cones, located on the retina (12). Complex signal processing events take place in which energy is converted by the rods and cones into electrical signals. These signals are information packets that are transmitted via the optic nerve to the brain. The signals report the relative response of the rods and cones to the incident light, and the brain decodes the information to tell us what we are seeing. Three kinds of cones present in all humans sense approximately red, green and blue portions of the visible spectrum. Rods sense the lightness/darkness attributes of the reflected light.
It is estimated that there are 10 million definitely distinguishable colors (13). Color scientists have organized colors into three attributes: lightness, hue, and saturation. Persons with normal color visions can distinguish extremely slight differences in lightness, hue and saturation when comparing colors.
Modern Color Theory
Test strips utilize this sensitive color matching ability. Reacted test strip colors are compared with printed color blocks to determine concentration. Color scientists use a defined standard “color difference unit” to quantitatively describe differences between colors.
We strive to create the greatest possible color difference with concentration in terms of these units. We also use color measurement tools to select the best color match between reagent strip and the color blocks that are usually printed on the reagent strip bottle label or on other packaging materials. The match between reagent strip and color block colors is also checked in different light sources such as fluorescent, daylight and incandescent, to confirm that there is no metamerism.
Metamerism is the property of two objects matching in color when viewed under one type of lighting, but do not match in color when viewed under a different type of lighting. The design of reagent strips according to strict objectives and the good color acuity of most people combine to produce accurate results with very good precision in the wide range of testing in which reagent strips are applied.
Accuracy and Precision
Excellent accuracy and precision data for three reagent strips used in dialysis centers are given in Table 5. In this context, accuracy refers to how closely the average values agree with the target values in column 1, and precision refers to the relative value of the standard deviation. The number of replicates is also indicated. The accuracy and precision of SteriChek Sensitive Total Chlorine Reagent Strips were determined with samples to which either sodium hypochlorite or monochloramine were added to give total chlorine levels indicated. Amperometric titration was used as the reference method (15).
The accuracy and precision of SteriChek Residual Peroxide Reagent Strips were determined with solutions made by diluting hydrogen peroxide or peroxide/peracetic acid solutions with reverse osmosis/distilled water. The concentrations are indicated in Table 5. We measured the peroxide concentrations with a reference method (16). The accuracy and precision of SteriChek Sensitive Hardness Reagent Strips were demonstrated with solutions containing 0, 5, 10, and 20 ppm as calcium carbonate using total hardness standard solutions from Hach Company. An accredited reference laboratory using the ICP method following approved EPA methodologies measured the calcium levels in the standard solutions.
The average and standard deviation results for total chlorine in Table 5 show that the reagent strips readily distinguish total chlorine concentration levels of 0.0, 0.1, 0.5, 1.0, 3 and 10 ppm. The strip therefore provides a reliable means of measuring total chlorine at all the levels, including the levels important for feed water (0.1 ppm) and rinse water (0.5 ppm).
The results for residual peroxide in Table 5 show that the reagent strip readily distinguishes peroxide concentration levels of 0.0, 1.0, 3, 5 and 10 ppm. The strip therefore provides a reliable means of measuring the residual peroxide in rinse water at the concentrations shown, and is a reliable indicator of residual peroxide when the concentration is below 1 ppm.
The average and standard deviation results for water hardness in Table 5 show that the total hardness reagent strip readily distinguishes water hardness levels of 0, 5, 10 and 20 ppm. These concentrations are equivalent to 0.0, 0.3, 0.6 and 1.2 grains per gallon, respectively. The strip therefore provides a convenient and accurate means of measuring and monitoring total hardness at very low levels in water.
Conclusion
Reagent strips are used in medical and non-medical applications to measure properties of aqueous solutions and other fluids. In medical applications, reagent strips are now used all over the world by laypersons to measure their own blood sugar. They are widely used in urine analysis screening, usually by health professionals because of their reliability and laborsaving attributes. They are also increasingly used for water analysis and to test disinfectant residuals and potency in dialysis centers. To get the best accuracy and precision of these devices, users are advised to follow use directions closely. The accuracy and precision results shown in this report are typical of reagent strip performance and underscore the reasons why these convenient, easy-to-use devices have found such wide application.
Dan Morris, PhD, is director of Technology, Environmental Test Systems (ETS, manufacturer of SteriChek reagent test strips), Elkhart, IN. Joe Sweazy is Technical Service associate at ETS.
References
- The Effect of Intensive Treatment of Diabetes on the Development and Progression of Long-Term Complications in Insulin-Dependent Diabetes Mellitus. New England Journal of Medicine (September 30, 1993), 329(14)
- Cost effectiveness analysis of improved blood pressure control in hypertensive patients with type 2 diabetes. UKPDS Study Group, British Medical Journal 317: 720-726, 1998
- Efficacy of atenolol and captopril in reducing risk of macrovascular and microvascular complications in type 2 diabetes. UKPDS Study Group, British Medical Journal 317: 713-720, 1998
- Tight blood pressure control and risk of macrovascular and microvascular complications in type 2 diabetes. UKPDS Study Group, British Medical Journal 317: 703-713, 1998
- Effect of intensive blood-glucose control with metformin on complications in overweight patients with type 2 diabetes. UKPDS Study Group, Lancet 352: 854-865, 1998
- Intensive blood glucose control with sulphonylureas or insulin compared with conventional treatment and risk of complications in patients with type 2 diabetes. UKPDS Study Group, Lancet 352: 837-853, 1998
- Skyler JS et al: Home blood glucose monitoring as an aid in diabetes management. Diabetes Care 1:150-157, 1978.
- Sonksen PH, Judd Sl, and Lowy C: Home monitoring of blood glucose- method for improving diabetic control. Lancet 1: 729-732, 1978.
- Walford S et al: Self-monitoring of blood glucose -- improvement of diabetic control: Lancet 1: 7320735, 1978.
- R. W. Waynant, Ph.D.* and V. M. Chenault, Ph.D., MT(ASCP)**: Overview of Non-Invasive Fluid Glucose Measurement Using Optical Techniques to Maintain Glucose Control in Diabetes Mellitus, Leos Newsletter 12 (2), 1998. *Food and Drug Administration; Center for Devices and Radiological Health; Office of Science and Technology* and Office of Device Evaluation**
- R. G. Kuehni, Color – An Introduction to Practices and Principles, p. 14-15, John Wiley and Sons, 1997.
- ibid. p. 28-36.
- D. B. Judd and G. Wyszecki, Color in Business, Science and Industry, pp. 314, John Wiley and Sons, 1975.
- ibid. pp. 317-321.
- Amperometric Titration Method, Standard Methods for the Examination of Water and Wastewater, 18th Edition, pp. 4-41 to 4-43, American Public Health Association, Washington, D.C., 1991
- A. Claiborne et al. Spectrophotometric Method – Journal of Biological Chemistry, 254, pp. 4245-4252, 1979.
- From blind studies conducted with multiple persons.
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