Comparative Analysis of Assays for Detection of Cell-Mediated Immunity Toward Cytomegalovirus and M. tuberculosis in Samples From Deceased Organ Donors

Abstract

Cell-mediated immunity assays could be valuable for risk assessment of organ donors, but no data exist on their feasibility in deceased donors. In this study, 105 deceased donors (52.3 AE 16.9 years) were screened at the time of organ procurement. Pathogen-specific stimulation was performed using a cytomegalovirus (CMV) lysate, tuberculin (purified protein derivative [PPD]) and soluble Mycobacterium tuberculosis-specific ESAT-6/CFP-10 proteins in combination with an inhouse fluorescence-activated cell sorting (FACS) assay or commercial assay formats (QuantiFERON-CMV/TB for ELISA, T-SPOT.TB for ELISPOT). CMV-IgG antibody titers were determined as gold standard for CMV infection; 51.4% of samples were CMV seropositive. Indeterminate results were observed in 47.6% of ELISA, 12.5% of FACS and 0% of ELISPOT assays. Agreement with serology was highest for FACS (95.6%, k ¼ 0.91), followed by ELISPOT (84.0%, k ¼ 0.68) and ELISA (80.0%, k ¼ 0.60). Agreement between ELISA and serology increased if the CMV lysate was used as stimulus (96.7%, k ¼ 0.92). Among the T cell assays, agreement between ELISPOT and FACS was highest (k ¼ 0.70). PPDpositive results among valid samples differed between assays (26.5% for ELISA, 23.1% for FACS and 50.5% for ELISPOT); 2.0% were QuantiFERON-TB positive, 3.3% were ESAT-6/CFP-10-positive in FACS and 13.4% were positive in the T-SPOT.TB assay. In conclusion, cellular immunity may be analyzed from samples of deceased donors, although the assays differ in the rate of positivity and indeterminate results.

Superior Sensitivity of Ex Vivo IFN-γ Release Assays as Compared to Skin Testing in Immunocompromised Patients

“…In line with other studies (8,26), this may also result in a higher percentage of indeterminate results in transplant recipients using the ELISA. Interestingly, similar within-test agreements were also found after comparative analysis of M. tuberculosis-or CMV-specific immune responses using the ELISA, ELISPOT and the FACS assay in samples from deceased donors (9). This indicates that the results of this study may also be extended to other antigen-specificities and to other vulnerable patient groups.…”

“…Detection of antigen-specific cellular immunity using flow-cytometry Stimulation-induced antigen-specific CD4 T cell frequencies were determined based on intracellular cytokine staining from heparinized whole blood with titered amounts of PPD (7.32 mg/mL, Statens Serum Institute RT50), soluble ESAT-6 and CFP-10 (10 mg/mL each) as previously described (9)(10)(11). Stimulation with phosphate buffered saline (diluent) and 2.5 mg/mL Staphylococcus aureus Enterotoxin B (SEB, Sigma, Deisenhofen, Germany) served as negative and positive controls, respectively.…”

Assessment and prevention of cytomegalovirus infection in allogeneic hematopoietic stem cell transplant and in solid organ transplant: A multidisciplinary consensus conference by the Italian GITMO, SITO, and AMCLI societies

“…In the cases of suspected passive antibody acquisition, CMI assays may be useful in establishing true immunologic status . This can be considered a feasible practice in recipients and in live donors, while in deceased donors indeterminate results may occur …”

Risk of tuberculosis after lung transplantation: the value of pretransplant chest computed tomography and the impact of mTOR inhibitors and azathioprine use

“…Donor TST status is not generally known and it is not possible to perform a TST in the context of a donation. We are unable to validate the performance of new techniques based on the detection of gamma‐interferon production in response to M. tuberculosis antigens (interferon‐gamma release assays) in this population . Newer TB diagnostics are needed to detect the presence of TB in potential donors.…”