Diagnostic workflow for X‐linked agammaglobulinemia integrating <scp>BTK</scp> expression and B‐cell maturation analysis by flow cytometry
X-linked agammaglobulinemia (XLA) is most often caused by impaired Bruton's tyrosine kinase (BTK) function. Rapid laboratory support is needed when genetic findings are inconclusive or pending. We developed a flow-cytometric assay to quantify intracellular BTK protein in monocytes, aiding the diagnosis of XLA. A stain index (SI) reference range was established using healthy donors (n = 25; SI median 1.14, range 1.01-1.66). ROC analysis defined a diagnostic cutoff of SI = 1.79, providing 95% sensitivity and 100% specificity compared to patients with pathogenic BTK variants (n = 40). Using this cutoff, abnormal BTK expression was detected in 38/40 genetically confirmed XLA patients. All patients with variants of uncertain/likely pathogenic significance (n = 17) had SI values above the cutoff. Bone marrow analysis in a patient subset showed a developmental arrest at the Pre-BI stage. The assay offers a rapid, clinically applicable readout to support XLA diagnosis and carrier evaluation in females, effectively complementing genetic testing.
Authors
- Mariia Fadeeva
- Oyuna Lodoeva (ORCID: https://orcid.org/0000-0002-2874-0014)
- Yulia Rodina (ORCID: https://orcid.org/0000-0001-9857-4456)
- Ирина Абрамова (ORCID: https://orcid.org/0000-0003-2335-0735)
- A Yur'evna Shcherbina
- Daria Yukhacheva
- Natalia Zinoveva (ORCID: https://orcid.org/0000-0002-6926-2055)
- Elena Raykina (ORCID: https://orcid.org/0000-0002-7634-2053)
- Daria Fomina (ORCID: https://orcid.org/0000-0002-5083-6637)
- Anna Roppelt (ORCID: https://orcid.org/0000-0001-5132-1267)
- Elvira Musaeva (ORCID: https://orcid.org/0000-0003-0581-9472)
- Tatiana Sozonova (ORCID: https://orcid.org/0009-0004-0057-2727)
- Viktoriia Vedmedskaia (ORCID: https://orcid.org/0000-0001-7247-4844)
- Aleksey Merkushov (ORCID: https://orcid.org/0000-0001-9517-5046)
- Mariia Igonina (ORCID: https://orcid.org/0009-0006-8667-0671)
- Ekaterina Malakhova (ORCID: https://orcid.org/0000-0002-7334-0706)
- Dmitry Pershin (ORCID: https://orcid.org/0000-0002-6148-7209)
- Anna Mukhina (ORCID: https://orcid.org/0000-0002-3305-1694)
- Amina Kieva (ORCID: https://orcid.org/0000-0003-2467-2840)
- Michael Maschan (ORCID: https://orcid.org/0000-0003-1735-0093)
- Yuri Shlyapnikov (ORCID: https://orcid.org/0000-0001-7800-3525)
Institutions
- Sechenov University (RU)
- Institute of Theoretical and Experimental Biophysics (RU)
- City Children's Hospital No. 22 (RU)
- Russian Children's Clinical Hospital (RU)
- Research Centre for Medical Genetics (RU)
- Dmitry Rogachev National Research Center of Pediatric Hematology, Oncology and Immunology (RU)
- Astana Medical University (KZ)
Publication Details
- Journal
- Cytometry Part B Clinical Cytometry
- Published
- 2026-05-11
- DOI
- https://doi.org/10.1002/cyto.b.70038
- Primary Topic
- Immunodeficiency and Autoimmune Disorders
- Type
- article
- Field-Weighted Citation Impact
- 0.00