The IGRA procedure includes:
Method
Detection of the interferon-gamma (IFN-g) produced during the test. One commercial test (QFT) uses an enzyme-linked immunosorbent assay (ELISA) method to measure soluble cytokines, while the other method (T-Spot) employs enzyme-linked immunospot assay (ELISPOT) method to stain the interferon-gamma in T cells. These newer generation assays have improved specificity by decreasing false positives caused by interference from the BCG vaccine or nontuberculosis mycobacteria.
Note: Processing, testing, and interpretation steps are detailed and complex. Many public health and reference laboratories offer the test, but smaller medical laboratories may not have adequate staff or budget to devote to this technically challenging, laborious, and expensive test.
Procedure (Note - This synopsis is intended to provide a basic understanding of the IGRA procedure; to provide detail is beyond the scope of this course. For detail, please consult individual vendor package inserts.)
- Both methods require fresh blood drawn into a lithium heparin tube. (Note that sodium citrate is also acceptable for the ELISPOT method.)
- Blood samples must be mixed with antigens and controls.
- At this time, both IGRA methods utilize synthetic peptides (mycobacterial proteins) representing ESAT-6 and CFP-10; these proteins stimulate the patient cells in heparinized whole blood.
- ELISA and ELISPOT use 96-well microplates. A series of steps involve the addition of conjugate and substrate reagents, wash steps.
- Interferon-gamma (IFN-g) is released by the patient's T-cell lymphocytes.
- Measurement of the IFN-g is performed. In the QuantiFERON® method, the amount of IFN-g released in response to the M. tuberculosis antigens and control substances is measured quantitatively, utilizing a standard curve. Numerical values include responses to two controls (nil and mitogen). In the SPOT® TB method, the number of interferon-gamma producing cells (precipitate appearing as spots) are measured/counted. (Both quantitative and qualitative results should be reported along with interpretation.)
Advantages
- A single visit to the lab blood draw site is required. (IGRA test has been associated with higher levels of LTBI treatment completion and compliance [Stockbridge et al., 2020].)
- Results may be available within 24 hours.
- No booster phenomena is present.
- No errors like those associated with TST placement and reading.
- BCG vaccination does not usually cause a false positive as in TST. (This makes IGRA the preferred test for anyone who has received a BCG vaccine.)
Disadvantages
- QFT whole blood samples must be processed within 16 hours, while T-Spot blood cells must be processed within 8-32 hours. (This is because white blood cells must be fresh and viable.)
- Preanalytic errors in collection and transport, analytic errors in running the test, or postanalytic errors in the interpretion of the assay can be problematic.
- IGRAs are relatively new, and limited data exist on the use of IGRAs to predict the progression to TB disease.
- Limited data on children under the age of 5 years, individuals recently exposed to M. tuberculosis, and immunocompromised.
- Limited data is available on serial testing.
