Selected Pool of Peptides from ESAT-6 and CFP-10 Proteins for Detection of Mycobacterium tuberculosis Infection

Abstract

We have validated a new test for detecting Mycobacterium tuberculosis infection. A pool of synthetic peptides derived from ESAT-6 and CFP-10 proteins was used to detect the number of specific gamma interferonproducing T cells by means of an enzyme-linked immunospot assay. Sixty-eight individuals positive for M. tuberculosis infection, either human immunodeficiency virus-seropositive or -seronegative, were studied. The test results were highly specific (87.5%) and sensitive (93.1%), more so than a classical lymphoproliferative assay (specificity and sensitivity of 77.27%), opening new possibilities for diagnosis and screening of tuberculosis. Moreover, the test allowed us to distinguish individuals infected with M. tuberculosis from those vaccinated with BCG.

Comparison of an In-house and a Commercial RD1-based ELISPOT-IFN- Assay for the Diagnosis of Mycobacterium tuberculosis Infection

“…All subjects underwent previous informed consent, the Mantoux TST, our in-house ELISPOT-IFN-γ assay, and the commercial T-SPOT.TB assay (Oxford Immunotec, Oxford, UK; currently approved for use in Europe and awaiting US Food and Drug Administration approval). [6][7][8] The origins and characteristics of the patients and their history of BCG vaccination (documented by charts in 15 individuals and proved by a scar in 23 individuals) are shown in table 1. All subjects were classified for TB infection on the basis of the 5-point scale of the American Thoracic Society (ATS).…”

“…6 The responses to the commercial assay were considered positive when the number of spots in the test wells (against at least one of the two tested antigens) was ≥6 times the number of spots in the negative control wells or, if the negative control had more than 6 SFCs, at least twice that number. The discrepancy between the cutoff for the two tests could depend on the different reagents and materials used in each assay (e.g., capture and detection mAbs, different concentrations of the reagents, type of plates) and their optimization.…”

“…6 Briefly, the PBMCs (2 x 10 5 cells/well) were seeded in duplicate in 96-well plates (MAIPS4510, Millipore, Bedford, MA) precoated with an anti-IFN-γ capture monoclonal antibody (mAb) (clone B-B1, Diaclone, Besançon, France). A pool of six >70% purified, 20 amino acid-long synthetic M. tuberculosis peptides (MTP) (Primm Srl, Milan, Italy) derived from the sequences of the ESAT-6 and CFP-10 proteins were used at a final concentration of 2 µg/ml/peptide to detect a specific response (ESAT-6 protein peptide 71-90 was added to the original pool of five peptides).…”

“…ESAT-6 and CFP-10 proteins that encompass multiple epitopes 6 in subjects classified as having TB infection. The test proved to be highly specific (88%) and sensitive (93%).…”

A systematic review of rapid diagnostic tests for the detection of tuberculosis infection

“…Many papers did not meet several of the study selection criteria. The bibliography of a recent systematic review 52 revealed six potentially relevant references, [248][249][250][251][252][253] which had to be excluded as they did not meet our inclusion criteria. A list of excluded studies with reasons is available.…”