Asthma Allergy Immunology

Asthma Allergy Immunology

2018, Vol 16, Num, 2     (Pages: 081-089)

Diagnostic Usage of Intracellular Protein Staining by Flow Cytometer in Primary Immune Deficiencies; Marmara Experience

İsmail ÖĞÜLÜR 1, Safa BARIŞ 1, Ahmet ÖZEN 1, Ercan NAIN 1, Ayça KIYKIM 1, Elif KARAKOÇ-AYDINER 1,

1 Marmara Üniversitesi Tıp Fakültesi, Pediatrik Allerji ve İmmünoloji Bilim Dalı, İstanbul, Türkiye

DOI: 10.21911/aai.369
Viewed: 767
Downloaded : 363

Objective: The aim of our study was the optimization and standardization of intracellular dedicator of cytokinesis 8 (DOCK8), LPS-responsive beige-like anchor protein (LRBA), SH2D1A/SLAMassociated protein (SAP) and X-linked inhibitor of apoptosis protein (XIAP) protein expressions in healthy controls with a single flow cytometer protocol and to concomitantly evaluate the possible use of this method for diagnosis.

Materials and Methods: Peripheral blood mononuclear cells were isolated from heparinized blood samples. Protein expressions were analyzed as mean fluorescein intensity difference (ΔMFI) according to the isotype.

Results: ΔMFI values obtained by DOCK8 antibody staining were 21.3±4 in CD3+ T cells and 25±3.3 in CD20+ T cells in healthy controls. These values in patients with DOCK8 deficiency were either very low or completely absent. ΔMFI values obtained by LRBA protein antibody staining were 36±7.7 in healthy controls, while they were at the very low levels of 5.9±1.8 in the LRBA protein deficiency patients. The values obtained by SAP and XIAP antibody staining were 30.2±3 in CD8+ T cells for SAP, 13.9±3.2 in CD3+ T and 14.6±3.5 in CD20+ B cells for XIAP. Since the SAP and XIAP results were not confirmed by gene sequencing, the results were not compared to healthy controls.

Conclusion: Due to its rapid and reliable results in clinically relevant cases for DOCK8 and LRBA deficiencies, analysis of protein expression is primarily suitable to evaluate intracellular staining protocol by flow cytometer. In addition, this particular method could be suitable for patients considered to be SAP and XIAP deficient.

Keywords : Flow cytometer, intracellular protein expression, mean fluorescence intensity, primary immunodeficiency