Lyme Disease: The ELISA Test and Ontario Health

Let’s break this down a bit. Four almost three months I have been having mysterious symptoms of the unexplained. I found a tick in my nose. Ontario Health ELISA returned negative for Lyme’s Disease (or antibodies associated with Borellia burgdorferi) but the tick itself came back positive. On the advice of my ND I sent bloodwork to California to IgeneX, Inc. for further testing as they do not utilize ELISA (non-specific, broadspectrum analysis) but focus on Immunofluorescence Assay (IFA), Western Blot (WB), Protein Chain Reaction (PCR) – serum and wholeblood, and they do a CD57 for the low cost of $600.13 USD. IgeneX is also rated the leading laboratory on clinical testing for Lyme’s and its associates. Big price tag, but thank goodness this is 1) tax deductible and 2) covered under my spousal benefits.

Before I get to my results let’s return to first year chemistry – CHEM1050.

The ELISA Test

The enzyme-linked immunosorbent assay (ELISA) is a test that uses antibodies and colour change to identify a substance. I should have paid more attention to those painful Tuesday evening chem labs because this is what we did for weeks on end. It was also what everyone ELSE in my lab was doing at graduate school. Damn it! There is a practical application beyond plant pathology?!!

This is what the ELISA plates look like in laboratory application. You can either determine how much antibody is in a sample, or you can determine how much protein is bound by an antibody. I will explain antibody detection as this is what Ontario Health does for its Lyme tests. Typical plates are 96-well which permits a high output of results.


96-well ELISA plate

The bottom of each well is coated with a protein to which will bind the antibody you what you want to measure. Whole blood is allowed to clot and the cells are centrifuged (spun quickly) out to obtain the clear serum with antibodies (called primary antibodies). The serum is incubated in a well, and each well contains a different serum. A positive control serum and a negative control serum would be included among the 96 samples being tested. After some time, the serum is removed and weakly adherent antibodies  washed off with buffer solution. A few more chemical washes and incubation time leaves the enzymes colourized. The plate is then put into an optical reader to measure the colour produced; the amount of colour produced is proportional to the amount of antibody bound to the proteins on the bottom of the wells.  But because it can sometimes provide false-positive results, it’s not used as the sole basis for diagnosis – right??

Each lab/province/country will have a different threshold for which they consider a positive or negative test. This test may not be positive during the early stage of Lyme disease as ELISA tests tend to show a lot of false negative results (Luger et al 1990). Golightly et al (1990) saw a lack of sensitivity with a 70% false-negative rate in early Lyme disease and from 4% to 46% with late manifestations of Lyme disease. Ontario Health acknowledges this flaw in its testing procedures:

ELISA testing is highly sensitive but not specific, and therefore serves as an ideal screening test to detect antibodies to B. burgdorferi. Since it has lower specificity, it can produce false-positive results and may cross react with antibodies that are produced as a result of other infections including those caused by other spirochetes.  (Sider et al, 2012).

Despite such a known false-negative assay, ELISA continues to be the primary screening test for Lyme’s Disease in Ontario, and only if it produces positive results will a Western Blot assay be completed to determine whether the sample is positive or negative.

Blood tests may be negative in patients with early Lyme disease (for example when a rash is present) or in patients who have had antibiotic treatment. The accuracy of blood tests increases as the infection progresses, although it is recognized that a very small proportion of patients with later-stage Lyme disease may test negative. The stage of infection and the possible impact of treatment on the outcomes of blood testing should be taken into consideration during diagnosis. (Public Health Canada, 2013).

For illustrative use.

Sample processed ELISA plate for illustrative use.

The other thing to remember is that antibodies require an incubation period in the host and can occur anywhere between 3 days to 3 months, to years. So yes the primary test with ELISA may be negative, but as soon as a doctor sees that negative result it registers as negative ALWAYS due to their myopic view on the world. And it is very difficult to convince some doctors otherwise, that retesting should occur or that the test may be producing false negatives. Sadly, our medical system relies very heavily on this single test.

I had my bloodwork sent through to Life Labs Inc via my GP, and it came back negative, they also confirmed with the GP that they only do an ELISA as per Ontario Health protocol. They will not do further testing (i.e. Western Blot) unless it is positive. However the tick that was found in my nose (!!) was positive.

This tells me several things:

  1. The tick I found in my nose was positive as a carrier of Lyme Disease (B. burgdorferi) – definitive positive
  2. The tick may not have transferred the bacteria into my body (Ontario Health states it requires 24-48 hours to do this) but I err on the side of caution.
  3. The tick did have time to transfer bacteria into my system; there was not enough antibody available to be detected.
  4. The serum was contaminated.
  5. Ontario Health/Life Labs uses the same protocol of ELISA as primary screening – this could be a false negative.
  6. THIS tick is a red herring.

As a secondary assessment after lots of research I went with IgeneX despite its high price tag. They do not use ELISA in any of their protocol. However, Ontario Health warns against testing privately or out of country:

The results from [private] laboratories that are not using validated tests can lead to misdiagnoses that can be harmful to patients, to the extent that appropriate diagnoses and treatment can be delayed or precluded. The WB interpretations used by these laboratories, based on their own internal validation studies, are much more liberal than interpretations recommended based on CDC guidelines. (Sider et al, 2012).

So Health Canada and Ontario Health acknowledge this flaw, yet continue to rely on it for screening. This is not surprising because both government bodies fail to acknowledge that Lyme’s Disease is even a problem in Canada. But one must use reason with Ontario Health/Health Canada clinical studies. They have publicly stated (although difficult to find the reports) that the ELISA produces many false-negatives. Funny enough, the Canadian Lyme Disease Association does not directly endorse the Canadian Health system for LD testing (you’d think they would have faith in our health system). If you take a walk over to their website they recommend two laboratories (in the USA) which provides more accurate testing, IgeneX being one of them. Neither one of them utilizes ELISA as a primary screening test due to its insensitive protocol and ability for misdiagnosis.

So at this point, I do not put my full trust in either one of the testing systems. In a few weeks I will be retested with Ontario Health and at the end of September I will retest with IgeneX as I received some inconclusive results.



Golightly MG, Thomas JA, Viciana AL. (1990). The laboratory diagnosis of Lyme Borreliosis. Lab Med; 21: 299-304.

Luger SW, Krauss E. (1990). Serologic tests for Lyme disease: interlaboratory variability. Arch Intern Med; 15: 761-763.

Public Health Canada. (2013). Lyme Disease Frequently Asked Questions. Queen’s Printer, Ottawa. Retrieved from:

Sider D, Patel S, Russell C, Jain-Sheehan N and Moore S. (2012). Technical Report: Update on Lyme Disease Prevention and Control. Ontario Public Health. Retrieved from:


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