Which Tests to Perform?

There is a logical sequence in selecting laboratory tests to aid in the diagnosis of Lyme disease. Much like the hepatitis model, antigen is present early after initial infection. Using PCR, Schmidt, et al.³² have detected B. burgdorferi in urine within the first few weeks of infection. Their published success rate is greater than 90%. IGeneX has detected antigen in the urine as early as three days after a tick bite.

Later there is an antibody response in about 70% of patients. IgM appears first, followed by IgG. Even in the presence of IgM (detected by Western Blot), IGeneX has detected B. burgdorferi in serum and plasma using the multiplex PCR.

There are two different types of antibody assays: the ELISA and the Western Blot. The ELISA is a poor assay, with marginal sensitivity. The Western Blot is more sensitive. The increased sensitivity of the Western Blot is analogous to a mountain where the base is a Western Blot and the summit is an ELISA. The Western blot has considerably more sensitivity because it provides detection before the peak of the response.

The Western Blot is a qualitative assay based upon the individual visualization of a patient's unique antibody response against the various Borrelia antigens. This type of assay is not restricted by the sensitivity and specificity concerns as the ELISA. An ELISA assay with a quantitative cut-off, and which is not necessarily specific to only the most unique antigens of B. burgdorferi, needs to consider population aspects of false positives (specificity) and false negatives (sensitivity). A positive or equivocal ELISA must always be followed with the corresponding (IgM or IgG) Western Blot. IGeneX will run the ELISA only when a Western Blot is ordered with it.

Figure 11a. Time course of development of antibodies and appearance of antigen

These relationships can be depicted in the hypothetical model of an "idealized" B. burgdorferi infection. After the first year, or with persistent or recurrent disease, this model may not be valid. Panel testing (as used with other diseases, such as hepatitis and thyroid disease) gives a much more complete picture for diagnosis and is ultimately more economical for the patient.

Recurrent or persistent infection offers a unique diagnostic problem, since the IgG response may be absent in more than 50% of patients.^3-5 The IgM Western blot may be helpful for some patients with persistent disease, but care must be taken to rule out cross-reactions due to other flagellar diseases, rheumatoid factor, and perhaps mononucleosis.^13,15

Figure 11b. Time course of development of antibodies and appearance of antigen

During persistent or recurring disease, assays that detect antigen may be particularly useful.²⁰ The Lyme Dot Blot Assay for antigen detection is a good diagnostic assay, especially during the start up phase with new antibiotics, which seems to enhance antigenuria.¹⁶

The antigen tests (the Lyme Dot Blot Assay and the Reverse Western Blot) and the multiplex PCR can be complementary to one another. The genomic PCR requires at least one recoverable bacterium, or at least the DNA from one. The antigen assays and the plasmid PCR, on the other hand, need only pieces of antigen. Studies at the Rocky Mountain National Laboratory have shown that pieces of antigen are more common in the urine than are whole or semi-whole B. burgdorferi.²³ IGeneX is currently using the multiplex PCR, which simultaneously detects genomic and plasmid DNA in urine, serum, and whole blood.

Neurological symptoms due to Lyme disease should be studied by examining the cerebral spinal fluid. It may be necessary to perform all of the above assays, including the multiplex PCR, IgM and IgG Western Blots, and antigen capture (LDA), on the CSF.

IGeneX, Inc.
795 San Antonio Rd., Palo Alto, CA 94303 USA
Tel. 650.424.1191 / 800.832.3200 Fax. 650.424.1196