RP% Measurement by FCM as a Diagnostic Test for ITP

• Conditions: Immune Thrombocytopenia
• Interventions: Other: RP% Measurement by FCM as a Diagnostic Test for ITP

• Registration Number: NCT02967328
• Lead Sponsor: Shandong University

Brief Summary

Immature platelets-also termed reticulated platelets (RP)-are platelets newly released into the circulation, and have been associated with a variety of pathological bleeding events including primary immune thrombocytopenia (ITP). They can be assessed by flow cytometry (FCM) after staining with thiazole orange (TO) at low concentration and expressed as a fraction of the total platelet count (RP%). The diagnosis of primary ITP is based on differential diagnosis and the measurement of RP% can serve as an alternative diagnostic test that are useful in daily practice. Our study aimed at distinguishing primary ITP from other thrombocytopenic disorders, especially aplstic (hypoplastic) or chemotherapy-induced thrombocytopenia by FCM. The sensitivity and specificity of the assay as well as agreement between RP% measurement and monoclonal antibody-specific immobilization of platelet antigen (MAIPA) were analyzed accordingly.

Detailed Description

The investigators are undertaking a multi-center, prospective blind trial of 500 adults with thrombocytopenic disorders with a platelet count less than 60\*10\^9/L from 4 medical centers in China. In brief, 15 μl aliquots of anti-coagulated whole blood were incubated for 70 min with 5 μl of phycoerythrin-conjugated anti-CD42b monoclonal antibody (BD Pharmingen, Tokyo, Japan) and 1 ml of thiazole orange (Retic-COUNT; Becton-Dickinson, San Jose, CA, USA) diluted 10 times by phosphate-buffered saline. RP% was analyzed on a flow cytometer (FACScan, Becton-Dickinson) by measuring 10,000 events in the CD42b-positive fraction.

Clinical information of all participants including gender, age, platelet count and definitive diagnosis were recorded by an exclusive investigator. RP% results were revealed at the end of recruitment and after all FCM measurements were completed. The agreement between clinical diagnosis and RP% results were analyzed to identify primary ITP.

Study Timeline

• Status Verified: 2016-11
• Study Start: 2016-11
• Study First Submitted: 2016-11-16
• Study First Submitted QC: 2016-11-17
• Last Update Submitted: 2016-11-17
• Study First Posted: 2016-11-18
• Last Update Posted: 2016-11-18
• Primary Completion: 2017-12
• Study Completion: 2017-12

Recruitment & Eligibility

• Status: UNKNOWN
• Sex: All
• Target Recruitment: 500

Inclusion Criteria

1. Untreated adult patients of both gender between the ages of 18 and 75 years
2. Each participant showed a platelet count 60*10^9/L, with or without bleeding manifestations
3. Thrombocytopenic disorders including autoimmune-mediated, aplastic (hypoplastic) or chemotherapy-induced thrombocytopenia

Exclusion Criteria

1. Received high-dose steroids or IVIG within 3 weeks prior to the test
2. Received second-line ITP-specific treatments (eg, cyclophosphamide, 6-mercaptopurine, vincristine, vinblastine, etc) within 3 months prior to the test
3. Current HIV infection, hepatitis B virus or hepatitis C virus infections
4. Severe medical condition (liver and kidney function impairment). Unstable cardiovascular disease or uncontrolled hypertension.
5. Patients who are deemed unsuitable for the study by the investigator

Study & Design

• Study Type: OBSERVATIONAL
• Study Design: Not specified

Arm && Interventions

Group	Intervention	Description
RP% Measurement by FCM as a Diagnostic Test for ITP	RP% Measurement by FCM as a Diagnostic Test for ITP	The investigators are undertaking a multi-center, prospective blind trial of 500 adults with thrombocytopenic disorders with a platelet count less than 60000/uL from 4 medical centers in China. In brief, 15 ul aliquots of anti-coagulated whole blood were incubated for 70 min with 5 ul of phycoerythrin-conjugated anti-CD42b monoclonal antibody (BD Pharmingen, Tokyo, Japan) and 1 ml of thiazole orange (Retic-COUNT; Becton-Dickinson, San Jose, CA, USA) diluted 10 times by phosphate-buffered saline. RP% was analyzed on a flow cytometer (FACScan, Becton-Dickinson) by measuring 10,000 events in the CD42b-positive fraction.

Primary Outcome Measures

Name	Time	Method
Total platelet count (RP%)	The day upon enrollment	We defined an upper limit for healthy control subjects as mean + 3SD in this study.

Trial Locations

Locations (1)

Qilu Hospital, Shandong University; 🇨🇳 Jinan, Shandong, China