COOL SCIENCE THINGS
Pregnancy Tests + COVID-19 Testing: An Explanation of Lateral Flow Assays
“Lateral flow immunochromatographic assays” is a big phrase and seems intimidating. I’ll try to break it down for you.
When I’ve asked the older women in my life who have been pregnant, they told me that they found out through “peeing on a stick.” I was very confused. I thought they meant a tree stick.
What sort of tree stick would you need to pee on in order to tell you that there’s a baby in you? Isn’t that dirty? Why would people ever want to pee on a stick? Isn’t that dirty?
Just imagine: picking up a random stick from the side of the tree trunk. You pull it off, wrestling with the tree, and victorious when you finally do. You’re like “yes, this is gonna tell me that there is a clump of cells in my body that will cause a bunch of symptoms and develops into this child that needs to be forced out.” And then you pee on it. I don’t think tree stick is that smart, guys.
Anyway, as I grew older I finally figured out that it probably didn’t mean a tree stick — it was some medical thing that was way beyond my head to comprehend. It is a stick — but the main identification part was on this weird sheet of inexplicable paper that somehow told you you were pregnant? 🤯
And then I went down a rabbit hole (again). If this piece of paper is beyond my head, then I can do better. And I can (try to) communicate information about it (and, the science!).
How and Why These Strips Work
Remember the pH strips that you would use in high school chemistry during those few points of time that you got to do a lab?
👆🏽 These strips would turn to a certain color, and based on that, we could identify the alkalinity (basic-ness) or acidity of that weird green mixture you made when you clearly didn’t listen to your chem teacher.
If you want to know the specifics of how they work: 👇🏽
On a high level way (as an indicative measure, not necessarily chemically) lateral flow assays work in a similar way.
Lateral flow assays are diagnostic tests that implement a system of point-of-care testing: testing in real-time. They identify the biomarkers that indicate the existence of the target (i.e. a bacteria, virus, particular hormone, etc). They use stuff like blood, pee, food — you get the gist.
There are two different types:
- 🍞 Sandwich types — basically, colored line shows up means that test is positive.
- 🏆 Competitive types — means that absence of colored line indicates a positive test.
Lateral flow Immunoassays contain a few different parts to it
First they have to detect the specific biomarker indicative of the test. The biomarker is deposited on a specific section of the assay.
Without the assistance of external forces, this moves along a polymeric strip which has different zones characteristic of different antibodies and molecules.
Since the liquid is first deposited on the conjugate pad, this acts as the first line where the flow is very controlled and goes along a very specific pathway. This uses capillary action → meaning that the liquid goes on a very controlled path and does not have external forces like gravity acting upon it.
The conjugate pad contains the nanoparticles and antibodies which stick to the specific biomarker if it’s there.
- Antibodies are Y-shaped blood proteins that are produced typically in the body to identify the existence of foreign particles, and nanoparticles are microscopic molecules with high chemical affinity that can typically label the existence of such products.
- For coloration of the lines → they use different types of nanoparticles, which produce the colored line that shows up whenever the test is taken.
- Specific types = gold, carbon, or colored latex nanoparticles.
🏆Gold is commonly used because of its intense colors, as well has high binding affinity, surface area and stability.
⚫️ Carbon nanoparticles are also used because they have a high sensitivity, are hydrophobic and can bind well to different types of biomarkers. It is also accessible at low cost and has a good range for capillary action.
🔴 Colored latex nanoparticles are used because they utilize covalent binding and physical adsorption (adhesion of molecules to the surface) and are especially used because of their red color, which augments the visualization pathway in these tests.
These form a 3-way bond with the antibody and the biomarker.
- These continue moving through the nitrocellulose membrane, which is a common device used for protein-blotting because they have a high protein-binding affinity, so it is really easy to detect + stop glycoproteins, proteins and nucleic acids.
There are two different reaction lines: a test line and a control line.
- At the test line, we have the biomarker which was already bound to the conjugated antibody, now bound to the test antibody.
- At the control line, we verify if the test was valid or not. Excess antibodies which are not bound to the biomarker are allowed to flow to this line and bind to secondary molecules. This makes sure that the flow was 👌🏽.
- There are then specific lines with reagents (chemicals used to test if a reaction occurs) — in some cases, if binding happens, that’s a positive results. In other cases, if binding does not happen, that’s a negative result.
- The results are based on whether or not a line shows up. Sometimes, you can do a quantitative analysis of the biomarker levels based on how dense the line is.
- In order to do this, we use readers to test this.
In the case of pregnancy tests
For a pregnancy test, if one line shows up → you’re not pregnant.
If two lines show up → you are pregnant.
You can typically get these types over the counter. The other one, which doctors use, check your urine or your blood via a test in the lab. They are typically much faster and get results in a few minutes. However, the over-the-counter tests are not bad → they do not show false negatives, and they can show results within 10–15 minutes!
Additionally, they both operate on the same principle of HCG analysis.
For these, the main indicator is identifying a hormone called HCG (Human chorionic gonadotropin). This is formed by the trophoblast cells that surround a growing embryo → eventually, this stuff makes up the placenta (yes, that’s the protective barrier around the embryo. Yes, it is filled with bacteria and is released after the baby is out. Yes, cats eat their own placentas so that the baby doesn’t get infected. It’s pretty cool).
HCG is usually bound to gold nano-spheres in the test, which are sandwiched with the conjugated antibody and the test antibody. As more and more show up → the test shows a positive result.
In your urine, this is the typical accumulation of hCG → therefore, the levels of it vary depending on the time that one is taking the test. For the test, the detection limit is typically between 20–100 mIU/mL.
What about COVID-19 Testing?
Using lateral flow assays for COVID-19 testing would lead to a more rapid diagnosis than the more common method, which RT-PCR. I talked about this in a previous article 👇🏽
These typically analyze swabs from the throat and nose, and based on the presence of viral proteins in the analyte, the test will prove to be positive. The specific viral proteins used are antigens, which bind to specific antibodies.
In considering the sensitivity vs. specificity of lateral flow assays, there are commonly high sensitivities, meaning that they will provide a high accuracy in classification because true positive identification is almost always spot-on. However, this does reduce the specificity of the test, which identifies the true negatives from the false positives. Thus, a false positive rate is typically very common.
However, in the case where the test has a high specificity, sensitivity is also lowered.
- According to Gavi, the WHO recommends that the sensitivity of a test is >70 percent, while the specificity is 97 percent.
- In comparison to PCR tests, however, this would lead to a 25 percent less sensitive test for rapid diagnostic immunoassays. However, it is great with it’s high specificity → it can prove whether or not someone suspected of having COVID-19 really has it.
However, because of its high portability and easy administration, it is one of the ideal methods being utilized for COVID-19 Testing. Typically, it would be used at health camps in developing countries, where clinical infrastructure may not be completely well developed. In addition, countries like Slovakia and the UK would use it in order to screen the whole population for COVID-19.
Essentially — this thing is of high impact.
Hey there! I’m Orna, a 15-year-old innovator at TKS interested in biotech, AI, and in particular longevity. Thanks for reading my article, and I hope you’ll connect.
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