Dry Chemistry Testing: Part 1 of 3
In the past few generations, humans have taken unprecedented control of their own fertility. Much of the associated technology and scientific expertise has focused on limiting unwanted pregnancies. Hopeful parents-to-be, however, also have tools at their disposal: modern family-planning methods and a reasonable degree of diligence can allow them to manage the number of children they’d like to have, and when, with reasonable certainty.
Unfortunately, there remain a considerable number of people who want to have children, but for one reason or another experience trouble conceiving. Fortunately, the likelihood of a successful pregnancy can be increased by understanding when conception is most likely to occur.
Methods of Ovulation Detection
Women are most fertile from a period 3–5 days before ovulation to 1–2 days after ovulation. This period is referred to as the “fertility window.” At Day 5 before ovulation, the probability of conception is about 8%. That probability doubles at Day 1 before ovulation, and almost doubles again on the day of ovulation. It then falls rapidly to 7% one day after ovulation. If a woman is trying to conceive, this information can be very useful.
The challenge, therefore, is to determine when ovulation is about to happen.
Historically, several methods have been used to determine the time of ovulation. Basal body temperature (BBT) measurements and determination of urinary luteinizing hormone (uLH) are probably the most common, but both have limitations. BBT is measured with a specialized thermometer with significantly more gradations than the typical fever thermometer. It reports temperatures between 94°F and 100°F in 0.1°F increments. The method requires multiple temperature readings over several days upon awakening and before the woman gets out of bed. Ideally, data are entered into a chart each day. Ovulation causes a 0.1°F increase in temperature. However, this only occurs on the day of ovulation, which means that the period of enhanced fertility a few days before ovulation are not detected. Basal body temperature can also be affected by factors other than ovulation such as emotion, lack of sleep, and smoking. One published study found that BBT was accurate only 32% of the time[1].
Luteinizing hormone levels in blood increase dramatically 20–48 hours prior to ovulation and can be detected in urine 8–12 hours after it occurs in blood. The levels are low enough that an immunoassay typically based on a monoclonal antibody must be used to detect the hormone. While commercially available over-the-counter (OTC) tests for uLH have been available for some time, they too suffer from some limitations. The surge is only detectable 1–2 days prior to ovulation, which once again misses some of the fertility window. Interferences include the use of drugs used in the treatment of infertility and menopause. (There is also the subjective undesirability of using urine as the sample.)
Researchers have also identified changes in electrolytes in various body fluids that signal that ovulation is about to take place. Salivary conductivity changes up to six days prior to ovulation. But to increase reliability of the method, it is recommended that vaginal conductivity be measured as well[2]. Different ranges are given depending on whether the patient is taking fertility drugs. While the method is accurate, it does require the use of instrumentation which usually requires a learning curve. The cost of the instrumentation is also a barrier to adoption.
Detecting Ovulation with Dry Chemistry Tests
PortaScience, Inc., of Moorestown, New Jersey (now a division of DCN Dx) has developed a dry chemistry method for the prediction of ovulation. It is based on the observation that concentrations of calcium and magnesium ions in saliva drop significantly up to six days before ovulation. No instrumentation is necessary, as the test is formulated to be read by the naked eye. The test is simple to run. Saliva is collected in a small container, and an unmeasured amount of saliva is added to the dry chemistry test card containing the appropriate reagents. After a few minutes, there is a color change if ovulation is imminent. No instrumentation is required—the card is read with the naked eye and compared to a color chart. The method was deemed novel by the U.S. Patent Office, which resulted in the issuance of a patent[3].
This method is based on the use of chromoionophores, which are dyes known to change color in the presence of certain positively charged ions such as calcium and magnesium. These dyes are commercially available from reputable chemical manufacturers and have a long history of laboratory use. To provide a relevant threshold for the color change, ethylenediaminetetraacetic acid (EDTA) is added to the dyes. It has a higher affinity for calcium and magnesium and reacts with these ions before the ions can interact with the chromoionophore dyes. When the EDTA is saturated with these ions, then the color reaction can take place.
Developing Reliable Dry Chemistry Ovulation Tests
The chemistry described above makes an interesting science project, but much more is needed to turn that into a commercial product. Extensive experimentation was done to select the dyes, the card material, and the stabilizers and wetting agents in order for them to perform reliably. These formulations have also undergone stability testing to ensure that the card has the same performance in the hands of the user as it did in quality control testing at the manufacturer. The test was also field tested to ensure that essentially untrained users could get laboratory level results in their own homes.
This ovulation prediction assay is only one example of the kind of testing that can be done in a dry card format. The concept can be readily expanded to be applied to a wide variety of ionic species in water-based samples for which appropriate chromoionophores are known. This could include not only human clinical samples, but also veterinary samples, environmental samples, food stuff, cosmetics, and pharmaceuticals, to name just a few. The company has also developed tests for negatively charged ions (e.g., a test of iodide in table salt).
This test is read with the naked human eye, but simple instrumentation does exist to read these cards. This makes interpretation of the test results less subjective. Instrumentation brings with it the connectivity that is increasingly important in this digital age. The decision to make the test readable by eye or instrumentation is driven by several factors including cost and the frequency of testing. It makes little sense to add instrumentation cost and complexity to a test that is done only periodically. On the other hand, if many batches of animal feed need to be tested for heavy metals over periods of months to years, it is easier to justify the inclusion of instrumentation.
Conclusion
If there is a need for analytical chemistry testing for ionic species in a point-of-use setting, contacting the experts at PortaScience, Inc. is an excellent place to start. Even with a talented team of chemists and manufacturing experts in place, conversations with another group of professionals can expand the range of potential approaches.
Be sure to check out Part 2 and Part 3 of this series on dry chemistry testing.
DCN Dx is a globally recognized contract developer of point-of-care devices based in Carlsbad, California. Since its founding, DCN Dx has committed itself to furthering the rapid diagnostic and point-of-care test market through the continued evolution of technologies and applications. DCN Dx’s cross-functional team of scientists and engineers develop and integrate all aspects of the point-of-care device system, including complex binding reactions, cassettes, sample handling devices, and reader systems. The company assists clients in developing entire rapid diagnostic tests from concept to commercialization.
To learn more about DCN Dx’s services, click here.
[1] Maurizio Guida et. al. Efficacy of Methods for Determining Ovulation in a Natural Family Planning Program. Fertility and Sterility 72: 900-904 (1999)
[2] Fernando, R. S., J. Regas, and G. Betz. Prediction of ovulation with the use of oral and vaginal electrical measurements during treatment with clomiphene citrate. Fertil. Steril. 47(3): 409-415 (1987)
[3] U.S. Patent 5 914 271
