Infographic on how antibody tests work. Rapid diagnostic tests are similar to pregnancy tests. The patient sample flows down a strip, and if the sample contains antibodies they bind to the test strip. Gold nanoparticles carried with the antibodies turn the test line red to indicate a positive test. These tests only tell us if antibodies are present. Neutralisation assay tests involve mixing serial dilutions of the patient sample with a suspension of the virus. If antibodies are present they reduce the plaques that form when the samples are incubated, which can tell us about antibody effectiveness. Enzyme-linked immunoabsorbent assay tests (ELISA) involve adding a patient sample to a microplate coated with deactivated antigen. If the sample contains antibodies, they bind to the antigen. Enzyme-labelled antibodies bind to the patient antibodies, then an enzyme substrate is added which changes colour when it binds to the enzyme. This gives information on the level of antibodies in the sample. A number of other tests work on a similar basis to ELISA but with slight differences.
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Last week, we looked at what coronavirus antibody tests can and can’t tell us. In part 2 of this series of graphics on antibody testing, we look at how the tests work to arrive at their results. As we’ll see, there’s not only one type of antibody test and they can vary in accuracy and the time they take to produce results.

At the time of writing, the US has approved 13 different antibody tests for detecting coronavirus antibodies. Two different tests are currently used in the UK. These approved tests work in differing ways and can sometimes tell us different things. We’ll examine the main categories and how they work below.

Rapid diagnostic tests
When people talk about home testing, this is usually the type of test they’re referring to. Rapid diagnostic tests (RDTs) for coronavirus work on a similar basis to home pregnancy test (but you don’t have to urinate on these ones). These can function in a few different ways, but a simple description of a common method follows.

Your sample for these tests will usually be a blood sample (from a fingerprick), a saliva sample, or a sample taken using a nasal swab. You add the sample to one end of the test stick. A wick at the other end of the test stick draws the sample through.

As your sample passes up the stick, it passes through an area loaded with inactivated virus. If your sample contains antibodies for the virus, they’ll bind to it, picking it up and carrying it along up the test stick. The virus is also tagged with gold nanoparticles, which play an important part in generating the result.

As the sample continues its journey up the test stick, it approaches the test lines. These lines contain fixed antibodies which grab your antibodies, along with the virus and the gold nanoparticles they’re stuck to. Gold is red in nanoparticle form, and it causes a red line to appear in the test zone. If you don’t have any antibodies to the virus in your sample, there’s nothing for the fixed antibodies to grab, so the test line doesn’t turn red.

RDTs will usually have two test lines; one for IgM antibodies and one for IgG antibodies. There’ll also be a control line; this line uses control antibodies contained on the test stick to check that the test is working correctly. If the control line turns red, the test has functioned correctly. if it doesn’t, we have to do the test again with a different test stick.

RDTs are the quickest test for antibodies, but they are more limited than some of the other tests. They can only tell us if antibodies are present in the sample. They can’t tell us how many antibodies are present or how effective your antibodies are against the virus.

RDTs are also amongst the least accurate of the antibody tests. When it comes to coronavirus RDTs, the World Health Organisation currently recommends against their use.  [Note: this article previously referenced sensitivity figures quoted by the WHO for RDTs, but these were for antigen RDTs, not antibody RDTs].

Neutralisation assays
Unlike RDTs, neutralisation assays are lab-based tests. They also take up to 5 days, the longest out of all of the antibody tests examined here. The delay does come with a benefit, however: they can tell us information about a patient’s antibodies that none of the other tests can.

To carry out a neutralisation assay, your blood sample will be diluted to varying degrees. These different dilutions are then each mixed with a constant concentration of a virus suspension, incubated, then added to host cells in a petri dish. Covered in agar, the samples in the dishes are incubated over several days.

Over this time, the regions of infected cells (plaques) grow. Antibodies to the virus in the patient sample will reduce plaque formation. By comparing the results at different dilutions of the patient sample, we can determine how effective the antibodies to the virus in your sample are. This is useful to know, since antibody effectiveness can vary. If you have antibodies, but they aren’t particularly effective, you could still be at risk of contracting the virus again in the future.

Enzyme-linked immunoabsorbent assays (ELISA)
This is another lab-based test. Like RDTs and neutralisation assays, it can tell us whether antibodies to the virus are present in your sample. Additionally, it can give us an idea of the levels of antibodies in your sample, which can be an important factor in future immunity. There are a few slight variatios of this test – I’ve focused on the ‘indirect’ method here, but there are others that differ in some respects.

Indirect ELISA involves adding the patient sample to a microplate well coated with the deactivated virus or a virus protein. Any antibodies in your sample will bind to these when the microplate contents are incubated.

Subsequently, enzyme-labelled antibodies are added. These antibodies bind to the patient antibodies. After this, a substance that binds to the enzyme (a substrate) is added. This changes colour when it binds to the enzyme, with the intensity of the colour directly linked to the number of antibodies in the patient sample. So, the more antibodies the your sample has, the more intense the colour. If you don’t have antibodies, no colour change will be seen.

ELISA-based tests usually take a few hours to run. They usually have higher sensitivity and specificity than RDTs, meaning fewer false negative and false positive results. Multiple tests can also be carried out at once. However, it doesn’t give us information on the effectiveness of the antibodies in your sample.

Other immunoassays
There are a number of different immunoassay tests which work in a similar manner to ELISA but have some notable differences.
For example, chemiluminescent immunoassays use luminescent (light-producing) substrates instead of colour-changing ones. The amount of light produced is proportional to the level of antibodies in your sample.

A variant of this, the electro-chemiluminescence assay, labels the antibody with a substrate which produces light when an electric current is applied. Again, the level of light produced is proportional to the level of antibodies in your sample.

Approved coronavirus tests
In the UK, there are currently two antibody tests for coronavirus. One, from Roche, is an electro-chemiluminescence assay. This test has a reported specificity of 99.81% and a sensitivity of 100% 14 days after infection confirmation. It can produce results in 18 minutes.

The other antibody test being used is produced by Abbott. It’s a chemiluminescent microparticle assay with a reported specificity of 99.9% and a sensitivity of 100% 14 days after infection confirmation.

Some companies are offering the Abbott test where you can take your sample at home by pricking your finger before sending it off. However, Abbott themselves have stated that their test is not designed to run with fingerprick blood samples. So if you’ve been pondering purchasing one of these test kits, it might be wise to save your money and hold off until the official antibody testing is more widely available.

In the US, several other tests are approved for use, including RDTs and ELISA tests. There’s a run-down of these tests and their sensitivities and specificities (where known) on this page.

So, you’ve taken an antibody test
If you’ve had a coronavirus antibody test and got a result, what does it mean? Well, if you tested negative, it means no antibodies to SARS-CoV-2 were detected. Does that mean you’ve definitely never had the disease? We can’t be 100% certain because there’s always the small risk of a false negative result.

The same is true if you test positive, meaning that you do have antibodies to the disease. Again, there’s always a small risk of a false positive result. Even if the result is correct, we don’t know for definite that having antibodies will definitely prevent you from catching the disease again, or even if they do, how long that immunity might last.

Until we know more, antibody tests aren’t the gateway to a lockdown-free society that some have touted them as. Even with these tests now becoming more widely available, we’re still looking at waiting for a vaccine before we can return to pre-coronavirus life.

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References/further reading