Current recommendations for testing blood for evidence of antibodies against Lyme disease include a two-step or two-tier process. Both steps can be done using the same blood sample.
The Two-Tier Testing Decision Tree describes the steps required to properly test for Lyme disease. The first required test is the enzyme immunoassay (EIA) or rarely the indirect immunofluorescence assay (IFA). If this test yields negative results, the provider should consider an alternative diagnosis; or in cases where the patient with has had symptoms for less than or equal to 30 days, the provider may treat the patient and follow up with a convalescent serum. If the first test yields positive or indeterminate results, also known as, “equivocal,” two options are available: 1) If the patient has had symptoms for less than or equal to 30 days, an IgM Western Blot is performed; 2) if the patient has had symptoms for more than 30 days, the IgG Western Blot is performed. The IgM should not be used if the patient has been ill for more than 30 days. It is not recommended to skip the first test and go right to the Western blot. This may increase the frequency of false positive results and potentially lead to misdiagnosis and inappropriate treatment.
Caution regarding testing for Lyme disease: some commercial laboratories that conduct testing for Lyme disease utilize assays whose accuracy and clinical usefulness have not been adequately established. In the United States, FDA has cleared 70 serologic assays to aid in the diagnosis of Lyme disease.
A list of unvalidated tests available as of 2011 include:
- Capture assays for antigens in urine
- Culture, immunofluorescence staining, or cell sorting of cell wall-deficient or cystic forms of B. burgdorferi
- Lymphocyte transformation tests
- Quantitative CD57 lymphocyte assays
- “Reverse Western blots”
- In-house criteria for interpretation of immunoblots
- Measurements of antibodies in joint fluid (synovial fluid)
- IgM or IgG tests without a previous ELISA/EIA/IFA
Understanding Test Results for Infectious Diseases
The illustration depicts the likelihood of false positive and false negative test results based on the prior probability of a disease occurring in a given population. Clinicians should consider the likelihood of disease before performing laboratory testing. The likelihood that a patient has a disease depends on many factors:
- Has a patient been in an area where the disease is found?
- Does the patient have signs and symptoms typical of the disease?
- Does the patient have risk factors for contracting or developing the disease?
In populations where disease is rare or unlikely, testing is likely to lead to false positives more frequently than true positives
Understanding the EIA (enzyme immunoassay)
There are several types of EIAs that are validated and approved by the FDA which include the “ELISA” (enzyme-linked immunosorbent assay and “ELFA” (enzyme-linked fluorescent immunoassay).
EIA tests are designed to be very “sensitive” or, in other words, when they are used properly, almost everyone with Lyme disease will test positive. However, it is possible to test positive with an EIA test even when you do not have Lyme disease. This can occur due to serological cross-reactivity because of other medical conditions, including:
- Tick-borne relapsing fever
- Anaplasmosis (formerly known as granulocytic ehrlichiosis)
- Some autoimmune disorders (e.g., lupus)
- Bacterial endocarditis
- Infection with Helicobacter pylori, Epstein Barr virus, or Treponema denticola (bacteria found in the mouth that can cause gum disease and/or infection after dental procedures)
Therefore it is recommended to verify any “positive” or “equivocal” (indeterminate) EIA results by performing, the “second tier” of the two-tier testing: an immunoblot test such as a Western blot. By performing the Western blot or other FDA-approved type of immunoblot, this enables one to distinguish patients who have Lyme disease from those with other conditions.
Understanding an Immunoblot Test
The immunoblot is a laboratory test that looks for antibodies the body makes against different molecules, or “antigens,” that are part of the Borrelia burgdorferi bacteria. Western Blots were the first type of immunoblot developed for Lyme disease testing.
The Western blot test produces something that looks like a bar code, with several lines or “bands”. Each line represents antibodies to a different component of the bacteria. As with bar codes, the presence of any one or two lines is not particularly meaningful. Instead, it is the combination of multiple, specific lines that identifies the infection as being due to Borrelia burgdorferi.
Immunoblot tests for Lyme disease testing can detect two different classes of antibodies: IgM and IgG. IgM antibodies are made sooner, so testing for them can be helpful for identifying patients during the first few weeks of infection. The downside of testing for IgM antibodies is that they are more likely to give false positive results. Tests for IgG antibodies are more reliable, but can take 4-6 weeks for the body to produce in large enough quantities for the test to detect them.
Remember the Following:
- The immunoblot should not be run without first performing an EIA or IFA.
- The immunoblot should not be run if the EIA or IFA tests are negative.
- A positive IgM immunoblot is only meaningful during the first 4 weeks of illness
- If you’ve been ill for longer than 4-6 weeks and the IgG immunoblot test is negative, it is unlikely that you have Lyme disease, even if the IgM immunoblot is positive.
Note on test result interpretation: It is not correct to interpret a test result that has only some bands that are positive as being “mildly” or “somewhat” positive for Lyme disease. The criterion that requires at least 5 IgG bands reflects the fact that people with Lyme disease have at least 5 antigens (specific molecules) detectable.
American College of Physicians (ACP)- Lyme Self-Assessment Quiz
Here is a link to the ACP Initiative on Lyme disease containing six clinical scenarios regarding the evaluation, diagnosis and treatment of Lyme disease.