OC 26.3 - Novel Flow Cytometric Diagnostic Assay for Primary Immune Thrombocytopenia.

Sunday, July 10, 2022

3:15 PM – 3:30 PM

Location: ExCel Center, ICC Capital Suite Room 10&11

Presentation Format: Virtual

Presenting Author(s)

Philippa J. Woolley, MBChB, MRCP, FRCPath

Haematology specialist registrar
University College London Hospitals NHS Foundation Trust, London, United Kingdom
London, England, United Kingdom

Co-Author(s)

Adela Constantinescu-Bercu, PhD

Research Fellow
Unviersity College London
London, England, United Kingdom
MS

Matthew Stubbs

Consultant Haematologist
University College Hospitals London NHS Foundation Trust
London, England, United Kingdom
Rd

Rens de Groot

Senior Research Fellow
University College London
London, England, United Kingdom
JW

John-Paul Westwood

Consultant Haematologist
University College Hospitals London NHS Foundation Trust
London, England, United Kingdom
YG

Yanping Guo

Flow Cytometry Translational Technology Platform Manager
Univsersity College London
London, England, United Kingdom
Marie Scully, MD

Professor of Haemostasis and Thrombosis
Department of Haematology, University College London Hospitals, London, UK
London, England, United Kingdom

Background: Primary Immune Thrombocytopenia (ITP) remains a diagnosis of exclusion and the lack of diagnostic tests in this field makes conclusive diagnosis difficult.

Aims: A whole blood flow cytometry assay was developed to confirm the diagnosis of ITP and differentiate between immune versus non-immune thrombocytopenia.

Methods: Samples of 50 patients with ITP, 50 normal controls and 30 thrombocytopenic controls were analysed; written consent was obtained. Whole blood was incubated with 3 platelet markers (CD41, CD42b and CD61) and 3 immunoglobulin markers (IgA/IgG and IgM). Sample collection was standardised to 10000 platelet events. Gating for platelet events established CD41/CD42b/CD61 positivity and with-in these gates, IgA and IgG positive events were captured and formed the basis for a multinomial regression model.

Results: The 3 groups were of comparable ages and gender. Patients with ITP had a platelet count range of 3x109/L-213 x109/L and patients with active disease, patients on treatment and patients in remission were included.

Thrombocytopenic controls platelet count ranged from 4 x109/L-96 x109/L and normal controls platelet count ranged from 154 x109/L-474 x109/L. Receive operator curve (ROC) curve analysis confirmed a sensitivity of 87% and specificity of 84% (p=0.0006) and this equated to a positive predictive value of 90% in confirming a diagnosis of ITP (Figure 1).

This assay also confirmed a >99% probability of ITP at an IgG level >10 and a 90% probability at an IgA level of > 18 (Figure 2). In contrast, 97.5% of thrombocytopenic and normal controls had IgG levels < 3.7 and IgA levels of < 11.5 for thrombocytopenic controls and < 3.0 for normal controls.

Conclusion(s): This assay demonstrates the first clinically useful technique for confirming the diagnosis of ITP by whole blood flow cytometry. This assay enables the differentiation of patients with ITP from both normal controls and patients with thrombocytopenia not related to ITP.