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Validation of Nijmegen-Bethesda assay modifications to allow inhibitor measurement during replacement therapy and facilitate inhibitor surveillance
Journal article   Open access   Peer reviewed

Validation of Nijmegen-Bethesda assay modifications to allow inhibitor measurement during replacement therapy and facilitate inhibitor surveillance

C H Miller, S J Platt, A S Rice, F Kelly, J M Soucie and Hemophilia Inhibitor Research Study Investigators
Journal of thrombosis and haemostasis, Vol.10(6), pp.1055-1061
06/2012
DOI: 10.1111/j.1538-7836.2012.04705.x
PMCID: PMC4477703
PMID: 22435927
url
https://doi.org/10.1111/j.1538-7836.2012.04705.xView
Published (Version of record) Open Access

Abstract

As part of a pilot U.S. inhibitor surveillance project initiated at the Centers for Disease Control and Prevention (CDC) in 2006, a centralized inhibitor measurement was instituted. To validate a modified method for inhibitor measurement suitable for surveillance of treated and untreated patients. In all, 710 subjects with hemophilia A were enrolled; 122 had a history of inhibitor (HI). Nijmegen-Bethesda assay (NBA) results on 50 split specimens shipped on cold packs and frozen were equivalent (r=0.998). Because 55% of 228 initial specimens had factor (F)VIII activity (VIII:C) present, a heat treatment step was added. Heating specimens to 56°C for 30 min and centrifuging removed FVIII, as demonstrated by a reduction of VIII:C and FVIII antigen to <1 U dL(-1) in recently treated patients. Among specimens inhibitor-negative before heating, one of 159 with negative HI and five of 30 with positive HI rose to ≥ 0.5 Nijmegen-Bethesda units (NBU) after heating. Correlation of heated and unheated inhibitor-positive specimens was 0.94 (P=0.0001). The modified method had a coefficient of variation (CV) for a 1 NBU positive control of 10.3% and for the negative control of 9.8%. Based on results on 710 enrollment specimens, a positive CDC inhibitor was defined as ≥ 0.5 NBU. Results were similar when 643 post-enrollment specimens were included. Of 160 enrolled hemophilia B patients, two had HI. All others had NBU ≤ 0.2 at enrollment. The CDC experience demonstrates that this modified NBA can be standardized to be within acceptable limits for clinical tests and can be used for national surveillance.
 United States Factor VIII - immunology Predictive Value of Tests Hemophilia B - diagnosis Prospective Studies Humans Blood Coagulation Tests - standards Drug Monitoring - methods Antigen-Antibody Reactions Reference Standards Time Factors Protein Denaturation Hemophilia A - diagnosis Factor VIII - therapeutic use Severity of Illness Index Reproducibility of Results Hemophilia B - immunology Enzyme-Linked Immunosorbent Assay Hemophilia A - immunology Hemophilia B - blood Treatment Outcome Calibration Hot Temperature Hemophilia A - blood Drug Monitoring - standards Hemophilia B - drug therapy Artifacts Coagulants - therapeutic use Antibodies - blood Coagulants - immunology Hemophilia A - drug therapy

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