To see if modification of the recently FDA-approved QuantiFERON blood test (MMWR 2003; 52[RR-02]:15) to include newer, quasi-specific tuberculosis (TB) antigens, such as ESAT-6 (Lancet 2000; 356:1099) may allow more specific in vitro testing of mycobacterial immunity.
Thirteen subjects were studied, 2 with TB, 1 after recovery from TB, 1 patient with active M kansasii infection, 3 with latent TB infection (LTBI), and 6 PPD negative healthy persons. Subjects were PPD tested within 4 weeks of being studied, unless they had a previous strong reaction. 5–10 ml venous blood were obtained; 1 ml incubated 24 hr in 24-well plates at 37°C, supernatants removed, and IFN-γ assays performed by a modified QuantiFERON ELISA assay using OptEIA (Pharmingen, SanDiego, CA). All samples were run in duplicate, and replicate assays were performed in 3 subjects to test variability and reproducibility. Preliminary tests were done using diluted specimens in 48-well plates to miniaturize the test for children.
The in vitro data using the modified assay correlated well with the results of PPD skin tests. Use of ESAT-6, as well as antigens prepared from M avium complex(MAC), and from BCG helped distinguish persons with BCG vaccination or nontuberculous mycobacterial (NTM) infection from patients with latent or active TB. The strongest ELISA response to TB antigen came in patients with past or present active TB. One with M kansasii was positive to ESAT-6. Repeat assays in 3 subjects at 1–3 month intervals showed consistency in specificity of responses; diluted specimens in 48-well plates gave satisfactory results as well.
This assay, as modified to include ESAT-6, may distinguish in one visit whether a patient has cellular immunity to MTB, or has PPD reactivity due to previous BCG vaccination or NTM infection.GRANT SUPPORT: None
H. Yeager, None.