What lab work should be ordered for the diagnosis of Lyme disease?

Laboratory Testing for Lyme Disease Diagnosis

For Lyme disease diagnosis, a two-tiered testing approach should be used, starting with a sensitive enzyme immunoassay (EIA) or immunofluorescent antibody (IFA) test, followed by a Western blot only if the first test is positive or equivocal. 1

Standard Two-Tiered Testing Approach

First-Tier Test

• Enzyme-linked immunosorbent assay (ELISA) or Indirect fluorescent antibody (IFA) test
  • Sensitivity: 85-100%
  • Specificity: 79-95%
  • Results reported as positive, negative, or equivocal/borderline 1
  • Most laboratories use whole-cell sonicate preparation of B. burgdorferi as antigen 2
  • Newer EIAs using VlsE lipoprotein or C6 peptide antigens are gaining popularity due to higher specificity 2

Second-Tier Test (Only if First-Tier is Positive or Equivocal)

• Western immunoblot (Western blot)
  • Specificity: >95% across all stages of disease 1
  • Interpretation criteria:
    • IgM Western Blot: ≥2 of 3 specific bands (21-24,39,41 kDa)
    • IgG Western Blot: ≥5 of 10 specific bands (18,21-24,28,30,39,41,45,58,66,93 kDa) 1
  • Important: IgM Western blot is not clinically interpretable after a patient has had 6-8 weeks of symptoms 2

Clinical Considerations

When Laboratory Testing Is Not Needed

• Patients with classic erythema migrans (EM) rash in an endemic area can be diagnosed clinically without laboratory confirmation 1
  • EM is defined as a gradually expanding annular lesion >5 cm in diameter
  • Approximately 70-80% of persons with Lyme disease have EM

Timing of Testing

• Early Lyme disease (acute phase): Two-tiered testing has limited sensitivity (30-40%) 3
• Convalescent phase (3-4 weeks later): Sensitivity improves to approximately 60-65% 4
• Late/disseminated Lyme disease: Both standard two-tiered testing and C6 EIA have excellent sensitivity (near 100%) 5

Alternative Testing Approaches

• Two-EIA algorithm: Using a whole-cell sonicate EIA followed by C6 peptide EIA (instead of Western blot)
  • Similar sensitivity to C6 testing alone
  • Better specificity than C6 alone
  • Equal specificity to standard two-tiered testing
  • May be easier to implement than Western blot 5

Special Situations

• PCR testing of synovial fluid: May be useful for suspected Lyme arthritis (sensitivity >75%) 1
• Intrathecal antibody testing: Helpful in suspected neuroborreliosis, particularly in regions with high seroprevalence 1

Common Pitfalls and Caveats

1. False negatives in early disease:

   • Early antibiotic treatment can blunt or prevent antibody response 1
   • Consider follow-up testing in 2-3 weeks if symptoms persist and clinical suspicion remains high 1

2. False positives:

   • A single positive IgG antibody band with an overall negative test result should not be interpreted as evidence of Lyme disease 1
   • Cross-reactivity can occur, especially with the 41-kDa band (flagellin protein) 1
   • In low-prevalence areas, positive predictive value of testing is poor (fewer than 20% of positive tests represent true Lyme disease) 6

3. Inappropriate testing:

   • Western blot should not be performed except as a reflex test after an initial EIA 2
   • Routine testing is not recommended for patients with typical amyotrophic lateral sclerosis, multiple sclerosis, Parkinson's disease, dementia, new-onset seizures, or psychiatric illness 1

4. Persistent symptoms after treatment:

   • A "post-treatment" Lyme disease syndrome may occur after appropriate antibiotic therapy
   • Persistent symptoms do not respond to long-term antibiotic therapy based on randomized-controlled trial data 2
   • Additional antibiotic therapy is not recommended for persistent or recurring nonspecific symptoms following recommended treatment 1

Remember that serological results must always be interpreted in the context of clinical signs and symptoms, and the predictive value of testing depends on the prevalence of Lyme disease in the region.