CD69 flow cytometry to complement interferon‐γ release assay for active tuberculosis
Background The interferon‐γ (IFN‐γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However, indeterminate IGRA results due to “high Nil” or “low PHA” responses limit its clinical utility. We developed a novel assay using CD69 flow cytome...
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| Vydané v: | Cytometry. Part B, Clinical cytometry Ročník 102; číslo 6; s. 471 - 486 |
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| Hlavní autori: | , , , |
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| Jazyk: | English |
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Hoboken, USA
John Wiley & Sons, Inc
01.11.2022
Wiley Subscription Services, Inc |
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| Abstract | Background
The interferon‐γ (IFN‐γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However, indeterminate IGRA results due to “high Nil” or “low PHA” responses limit its clinical utility. We developed a novel assay using CD69 flow cytometry (FC) to complement IGRA.
Methods
CD69 FC measures the surface CD69 expression on T cells prior to centrifugation to harvest the plasma for IGRA. T cell responses against Mtb antigen 1 (Ag1) or Ag2 were measured using three‐color FC (CD3, CD4, and CD69) in TB (n = 140) and non‐TB groups (n = 117). The cutoff values of Δ%CD69bright cells (stimulated minus unstimulated) for CD4+ and CD4− T cells were established based on healthy individuals (n = 63). The assay performances of CD69 FC and IGRA were compared.
Results
In subjects with determinate IGRA results (“positive” or “negative”; n = 216), the diagnostic accuracies of CD69 FC (90.3%) and IGRA (87.0%) were not significantly different (p = 0.31). For indeterminate IGRA results (n = 40), CD69 FC attained a diagnostic accuracy of 92.5%. The CD4+/CD4− ratio within CD69bright T cells measured by CD69 FC was significantly higher (p < 0.05) in the active TB group (6.39 ± 132.05; n = 72) than in other CD69 FC‐positive subjects (2.84 ± 15.36; n = 63) (p < 0.05), whereas CD8 responses expected by IGRA (difference of IFN‐γ levels between Mtb Ag tubes) did not differ significantly (0.00 ± 9.18 and 0.00 ± 4.25, respectively, IU/ml; p = 0.58).
Conclusions
We demonstrated the potential of CD69 FC as a simple, rapid assay for clarifying indeterminate IGRA results and identifying active TB. With further improvements, CD69 FC may complement the IGRA to enhance TB risk stratification in the routine diagnostic workup. |
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| AbstractList | BackgroundThe interferon‐γ (IFN‐γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However, indeterminate IGRA results due to “high Nil” or “low PHA” responses limit its clinical utility. We developed a novel assay using CD69 flow cytometry (FC) to complement IGRA.MethodsCD69 FC measures the surface CD69 expression on T cells prior to centrifugation to harvest the plasma for IGRA. T cell responses against Mtb antigen 1 (Ag1) or Ag2 were measured using three‐color FC (CD3, CD4, and CD69) in TB (n = 140) and non‐TB groups (n = 117). The cutoff values of Δ%CD69bright cells (stimulated minus unstimulated) for CD4+ and CD4− T cells were established based on healthy individuals (n = 63). The assay performances of CD69 FC and IGRA were compared.ResultsIn subjects with determinate IGRA results (“positive” or “negative”; n = 216), the diagnostic accuracies of CD69 FC (90.3%) and IGRA (87.0%) were not significantly different (p = 0.31). For indeterminate IGRA results (n = 40), CD69 FC attained a diagnostic accuracy of 92.5%. The CD4+/CD4− ratio within CD69bright T cells measured by CD69 FC was significantly higher (p < 0.05) in the active TB group (6.39 ± 132.05; n = 72) than in other CD69 FC‐positive subjects (2.84 ± 15.36; n = 63) (p < 0.05), whereas CD8 responses expected by IGRA (difference of IFN‐γ levels between Mtb Ag tubes) did not differ significantly (0.00 ± 9.18 and 0.00 ± 4.25, respectively, IU/ml; p = 0.58).ConclusionsWe demonstrated the potential of CD69 FC as a simple, rapid assay for clarifying indeterminate IGRA results and identifying active TB. With further improvements, CD69 FC may complement the IGRA to enhance TB risk stratification in the routine diagnostic workup. The interferon-γ (IFN-γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However, indeterminate IGRA results due to "high Nil" or "low PHA" responses limit its clinical utility. We developed a novel assay using CD69 flow cytometry (FC) to complement IGRA. CD69 FC measures the surface CD69 expression on T cells prior to centrifugation to harvest the plasma for IGRA. T cell responses against Mtb antigen 1 (Ag1) or Ag2 were measured using three-color FC (CD3, CD4, and CD69) in TB (n = 140) and non-TB groups (n = 117). The cutoff values of Δ%CD69 cells (stimulated minus unstimulated) for CD4 and CD4 T cells were established based on healthy individuals (n = 63). The assay performances of CD69 FC and IGRA were compared. In subjects with determinate IGRA results ("positive" or "negative"; n = 216), the diagnostic accuracies of CD69 FC (90.3%) and IGRA (87.0%) were not significantly different (p = 0.31). For indeterminate IGRA results (n = 40), CD69 FC attained a diagnostic accuracy of 92.5%. The CD4 /CD4 ratio within CD69 T cells measured by CD69 FC was significantly higher (p < 0.05) in the active TB group (6.39 ± 132.05; n = 72) than in other CD69 FC-positive subjects (2.84 ± 15.36; n = 63) (p < 0.05), whereas CD8 responses expected by IGRA (difference of IFN-γ levels between Mtb Ag tubes) did not differ significantly (0.00 ± 9.18 and 0.00 ± 4.25, respectively, IU/ml; p = 0.58). We demonstrated the potential of CD69 FC as a simple, rapid assay for clarifying indeterminate IGRA results and identifying active TB. With further improvements, CD69 FC may complement the IGRA to enhance TB risk stratification in the routine diagnostic workup. The interferon-γ (IFN-γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However, indeterminate IGRA results due to "high Nil" or "low PHA" responses limit its clinical utility. We developed a novel assay using CD69 flow cytometry (FC) to complement IGRA.BACKGROUNDThe interferon-γ (IFN-γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However, indeterminate IGRA results due to "high Nil" or "low PHA" responses limit its clinical utility. We developed a novel assay using CD69 flow cytometry (FC) to complement IGRA.CD69 FC measures the surface CD69 expression on T cells prior to centrifugation to harvest the plasma for IGRA. T cell responses against Mtb antigen 1 (Ag1) or Ag2 were measured using three-color FC (CD3, CD4, and CD69) in TB (n = 140) and non-TB groups (n = 117). The cutoff values of Δ%CD69bright cells (stimulated minus unstimulated) for CD4+ and CD4- T cells were established based on healthy individuals (n = 63). The assay performances of CD69 FC and IGRA were compared.METHODSCD69 FC measures the surface CD69 expression on T cells prior to centrifugation to harvest the plasma for IGRA. T cell responses against Mtb antigen 1 (Ag1) or Ag2 were measured using three-color FC (CD3, CD4, and CD69) in TB (n = 140) and non-TB groups (n = 117). The cutoff values of Δ%CD69bright cells (stimulated minus unstimulated) for CD4+ and CD4- T cells were established based on healthy individuals (n = 63). The assay performances of CD69 FC and IGRA were compared.In subjects with determinate IGRA results ("positive" or "negative"; n = 216), the diagnostic accuracies of CD69 FC (90.3%) and IGRA (87.0%) were not significantly different (p = 0.31). For indeterminate IGRA results (n = 40), CD69 FC attained a diagnostic accuracy of 92.5%. The CD4+ /CD4- ratio within CD69bright T cells measured by CD69 FC was significantly higher (p < 0.05) in the active TB group (6.39 ± 132.05; n = 72) than in other CD69 FC-positive subjects (2.84 ± 15.36; n = 63) (p < 0.05), whereas CD8 responses expected by IGRA (difference of IFN-γ levels between Mtb Ag tubes) did not differ significantly (0.00 ± 9.18 and 0.00 ± 4.25, respectively, IU/ml; p = 0.58).RESULTSIn subjects with determinate IGRA results ("positive" or "negative"; n = 216), the diagnostic accuracies of CD69 FC (90.3%) and IGRA (87.0%) were not significantly different (p = 0.31). For indeterminate IGRA results (n = 40), CD69 FC attained a diagnostic accuracy of 92.5%. The CD4+ /CD4- ratio within CD69bright T cells measured by CD69 FC was significantly higher (p < 0.05) in the active TB group (6.39 ± 132.05; n = 72) than in other CD69 FC-positive subjects (2.84 ± 15.36; n = 63) (p < 0.05), whereas CD8 responses expected by IGRA (difference of IFN-γ levels between Mtb Ag tubes) did not differ significantly (0.00 ± 9.18 and 0.00 ± 4.25, respectively, IU/ml; p = 0.58).We demonstrated the potential of CD69 FC as a simple, rapid assay for clarifying indeterminate IGRA results and identifying active TB. With further improvements, CD69 FC may complement the IGRA to enhance TB risk stratification in the routine diagnostic workup.CONCLUSIONSWe demonstrated the potential of CD69 FC as a simple, rapid assay for clarifying indeterminate IGRA results and identifying active TB. With further improvements, CD69 FC may complement the IGRA to enhance TB risk stratification in the routine diagnostic workup. Background The interferon‐γ (IFN‐γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However, indeterminate IGRA results due to “high Nil” or “low PHA” responses limit its clinical utility. We developed a novel assay using CD69 flow cytometry (FC) to complement IGRA. Methods CD69 FC measures the surface CD69 expression on T cells prior to centrifugation to harvest the plasma for IGRA. T cell responses against Mtb antigen 1 (Ag1) or Ag2 were measured using three‐color FC (CD3, CD4, and CD69) in TB (n = 140) and non‐TB groups (n = 117). The cutoff values of Δ%CD69bright cells (stimulated minus unstimulated) for CD4+ and CD4− T cells were established based on healthy individuals (n = 63). The assay performances of CD69 FC and IGRA were compared. Results In subjects with determinate IGRA results (“positive” or “negative”; n = 216), the diagnostic accuracies of CD69 FC (90.3%) and IGRA (87.0%) were not significantly different (p = 0.31). For indeterminate IGRA results (n = 40), CD69 FC attained a diagnostic accuracy of 92.5%. The CD4+/CD4− ratio within CD69bright T cells measured by CD69 FC was significantly higher (p < 0.05) in the active TB group (6.39 ± 132.05; n = 72) than in other CD69 FC‐positive subjects (2.84 ± 15.36; n = 63) (p < 0.05), whereas CD8 responses expected by IGRA (difference of IFN‐γ levels between Mtb Ag tubes) did not differ significantly (0.00 ± 9.18 and 0.00 ± 4.25, respectively, IU/ml; p = 0.58). Conclusions We demonstrated the potential of CD69 FC as a simple, rapid assay for clarifying indeterminate IGRA results and identifying active TB. With further improvements, CD69 FC may complement the IGRA to enhance TB risk stratification in the routine diagnostic workup. |
| Author | Kim, Shin‐Woo Won, Dong Il Han, Man‐Hoon Kim, Yoonjung |
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| BackLink | https://www.ncbi.nlm.nih.gov/pubmed/36161692$$D View this record in MEDLINE/PubMed |
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| Keywords | interferon-γ release assay flow cytometry CD69 tuberculosis |
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The interferon‐γ (IFN‐γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However,... The interferon-γ (IFN-γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However, indeterminate... BackgroundThe interferon‐γ (IFN‐γ) release assay (IGRA) is widely used to diagnose tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb). However,... |
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| SubjectTerms | Antigens Antigens, Bacterial Assaying CD3 antigen CD4 antigen CD69 CD69 antigen CD8 antigen Centrifugation Complement Diagnostic systems Flow Cytometry Humans Interferon Interferon-gamma Release Tests - methods interferon‐γ release assay Latent Tuberculosis Lymphocytes Lymphocytes T Mycobacterium tuberculosis Tuberculosis Tuberculosis - diagnosis Tubes |
| Title | CD69 flow cytometry to complement interferon‐γ release assay for active tuberculosis |
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