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Simple flow cytometric protocol of CD4+/CD8+ lymphocyte ratio assessment in bronchoalveolar lavage fluids from patients with interstitial lung diseases.
Anal Quant Cytol Histol. 2011 Oct; 33(5):289-96.AQ

Abstract

OBJECTIVE

To validate the fast and accurate flow cytometric (FCM) protocol using blood-standardized antibodies for alveolar lymphocyte subtyping with respect to standard immunocytochemistry (IC).

STUDY DESIGN

FCM and IC were applied to immunophenotype T cell subsets in bronchoalveolar lavage (BAL) fluids from patients with interstitial lung diseases. Diagnostic BAL specimens from 50 patients with suspected sarcoidosis, idiopathic pulmonary fibrosis, and hypersensitivity pneumonitis were evaluated by both IC and FCM. In FCM, CD4+ and CD8+ T cells were identified by light scatter gating with CD3 selection using basic tricolor cytometer.

RESULTS

Relative amounts of CD4+, CD8+ T cells, and CD4+/CD8+ ratios demonstrated by the FCM showed excellent, significant correlations with IC results. FCM values did not differ significantly from IC results. However, the sensitivity and specificity of conventional IC staining were not sufficient to assess CD4+/ CD8+ ratio in most idiopathic pulmonary fibrosis cases. Additionally, performing IC immunophenotyping in BAL samples with low lymphocyte content introduced a remarkable error into CD4+/CD8+ ratio assessment.

CONCLUSION

FCM allowed reliable, precise, and fast T-cell subset measurement in all BAL samples, overcoming the IC disadvantages. Our validated FCM protocol provides diagnostically relevant CD4+/CD8+ ratio determination by simple light scatter gating strategy with CD3 selection.

Authors+Show Affiliations

Laboratory of Molecular Diagnostics and Immunology and Department of Pathomorphology, National Institute of Tuberculosis and Lung Diseases, Warsaw, Poland.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Clinical Trial
Journal Article
Research Support, Non-U.S. Gov't
Validation Study

Language

eng

PubMed ID

22611757

Citation

Szpechcinski, Adam, et al. "Simple Flow Cytometric Protocol of CD4+/CD8+ Lymphocyte Ratio Assessment in Bronchoalveolar Lavage Fluids From Patients With Interstitial Lung Diseases." Analytical and Quantitative Cytology and Histology, vol. 33, no. 5, 2011, pp. 289-96.
Szpechcinski A, Kopinski P, Giedronowicz D, et al. Simple flow cytometric protocol of CD4+/CD8+ lymphocyte ratio assessment in bronchoalveolar lavage fluids from patients with interstitial lung diseases. Anal Quant Cytol Histol. 2011;33(5):289-96.
Szpechcinski, A., Kopinski, P., Giedronowicz, D., Rozy, A., Jagus, P., Szolkowska, M., & Chorostowska-Wynimko, J. (2011). Simple flow cytometric protocol of CD4+/CD8+ lymphocyte ratio assessment in bronchoalveolar lavage fluids from patients with interstitial lung diseases. Analytical and Quantitative Cytology and Histology, 33(5), 289-96.
Szpechcinski A, et al. Simple Flow Cytometric Protocol of CD4+/CD8+ Lymphocyte Ratio Assessment in Bronchoalveolar Lavage Fluids From Patients With Interstitial Lung Diseases. Anal Quant Cytol Histol. 2011;33(5):289-96. PubMed PMID: 22611757.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Simple flow cytometric protocol of CD4+/CD8+ lymphocyte ratio assessment in bronchoalveolar lavage fluids from patients with interstitial lung diseases. AU - Szpechcinski,Adam, AU - Kopinski,Piotr, AU - Giedronowicz,Dorota, AU - Rozy,Adriana, AU - Jagus,Paulina, AU - Szolkowska,Malgorzata, AU - Chorostowska-Wynimko,Joanna, PY - 2012/5/23/entrez PY - 2012/5/23/pubmed PY - 2012/6/13/medline SP - 289 EP - 96 JF - Analytical and quantitative cytology and histology JO - Anal Quant Cytol Histol VL - 33 IS - 5 N2 - OBJECTIVE: To validate the fast and accurate flow cytometric (FCM) protocol using blood-standardized antibodies for alveolar lymphocyte subtyping with respect to standard immunocytochemistry (IC). STUDY DESIGN: FCM and IC were applied to immunophenotype T cell subsets in bronchoalveolar lavage (BAL) fluids from patients with interstitial lung diseases. Diagnostic BAL specimens from 50 patients with suspected sarcoidosis, idiopathic pulmonary fibrosis, and hypersensitivity pneumonitis were evaluated by both IC and FCM. In FCM, CD4+ and CD8+ T cells were identified by light scatter gating with CD3 selection using basic tricolor cytometer. RESULTS: Relative amounts of CD4+, CD8+ T cells, and CD4+/CD8+ ratios demonstrated by the FCM showed excellent, significant correlations with IC results. FCM values did not differ significantly from IC results. However, the sensitivity and specificity of conventional IC staining were not sufficient to assess CD4+/ CD8+ ratio in most idiopathic pulmonary fibrosis cases. Additionally, performing IC immunophenotyping in BAL samples with low lymphocyte content introduced a remarkable error into CD4+/CD8+ ratio assessment. CONCLUSION: FCM allowed reliable, precise, and fast T-cell subset measurement in all BAL samples, overcoming the IC disadvantages. Our validated FCM protocol provides diagnostically relevant CD4+/CD8+ ratio determination by simple light scatter gating strategy with CD3 selection. UR - https://www.unboundmedicine.com/medline/citation/22611757/Simple_flow_cytometric_protocol_of_CD4+/CD8+_lymphocyte_ratio_assessment_in_bronchoalveolar_lavage_fluids_from_patients_with_interstitial_lung_diseases_ L2 - https://medlineplus.gov/interstitiallungdiseases.html DB - PRIME DP - Unbound Medicine ER -