Abstract Number: PB0362
Meeting: ISTH 2021 Congress
Background: The ST Genesia (STG) is a fully automated thrombin generation test (TGT) platform more standardized than the original Calibrated Automated Thrombogram (CAT) and designed to support clinical applications in the lab.
In some clinical indications such as prevention of thrombosis or ExtraCorporeal Membrane Oxygenation, patients may receive unfractionated heparin (UFH). TGT is adversely affected by UFH, thereby it cannot reflect basic haemostatic state (without the influence of heparin) in these patients, requiring UFH neutralization. Polybrene (Pb) at 25µg/mL has been shown to be effective for UFH neutralization in the original CAT system(1).
Aims: Conceive an automated TGT test using the STG-DrugScreen reagent neutralizing UFH and allowing to restore basic TGT parameters.
Select between Pb and Heparinase I (Hep) the best neutralizing agent based on test performances: dose-response effect, neutralization performances on UFH-spiked samples and on board stability.
Methods: Neutralizing agent (Pb or Hep) was added at different concentrations to the reconstituted STG-DrugScreen reagent.
Normal Pooled Plasma (NPP) and twelve donors’ leftover plasma samples were spiked with UFH. NPP was spiked with five different levels of sodium unfractionated heparin (HNa) and two levels of calcium heparin (HCa) to cover the STA-Liquid anti-Xa measuring range of 0-1.08IU/mL (Table 1).
| Performances | Sample | UFH Anti-Xa Concentrations | Neutralizing agent | Analytical Result |
|---|---|---|---|---|
| Dose-response | NPP | ≤ 0.10 IU/mL | Pb: 0 – 210 µg/mL; 6 levels Hep: 0 – 1.6 IU/mL; 7 levels |
Relative deviation to STG-DrugScreen TGT baseline |
| Neutralization performance on spiked NPP | Heparin-spiked plasmas | HNa: 0 – 1.08 IU/mL; 6 levels HCa: 0 – 1.07 IU/mL; 3 levels |
Pb: 0 – 210 µg/mL; 6 levels Hep: 0 – 1.6 IU/mL; 7 levels |
Relative deviation to STG-DrugScreen TGT baseline |
| On-board stability of modified STG-DrugScreen | Heparin-spiked plasmas | HNa: 0 – 0.8 IU/mL; 3 levels |
Pb: 25 µg/mL Hep: 1.2 IU/mL |
Relative deviation of STG-DrugScreen TGT parameters at T4 hours versus T0 |
| Neutralization performance on donor samples | Donors’ leftover plasma samples spiked with HNa | HNa: 0 – 0.8 IU/mL; 12 samples |
Pb: 25 µg/mL | Relative deviation to STG-DrugScreen TGT parameters before versus after heparin neutralization |
Evaluation of neutralizing agents in STG-DrugScreen reagent with citrated plasma samples.
Results: Table 2 provides summary of assay performances.
| Performances | Results |
|---|---|
| Dose-response | Pb had no significant impact for Pb at concentrations < 150µg/mL (< 10%). Hep had minimal impact (< 10%) on TGT of NPP regardless to the tested concentrations. |
| Neutralization performance on heparin-spiked NPP | 25 µg/mL of Pb or 1.2 IU/mL of Hep completely neutralized the effect of both HNa and HCa UFH on TGT. |
| On board stability of modified STG-DrugScreen | With 25 µg/mL of Pb or 1.2 IU/mL of Hep, on board stability relative deviation to reference time was less than 8% for all parameters except 11% for NPP Lag Time with Hep. On board stability with 25 µg/mL of Pb was up to 4h. |
| Neutralization performance on donor samples | Relative deviation to STG-DrugScreen was less than 8% with 25 µg/mL pf Pb, except for one donor sample which shifted to a more rapid TGT up to 14% Lag Time reduction. |
Summary of performance evaluation.
Conclusions: Our results show that 25µg/mL of Pb or 1.2IU/mL of Hep can completely neutralize the anticoagulant effect of UFH up to 1.08IU/mL. At these concentrations, TGT parameters measured in NPP were not impacted. Pb showed slightly better on-board stability than Hep.
These in vitro results indicate that Pb or Hep can be used added to STG-DrugScreen reagent to automatically neutralize UFH for TGT measurement on STG with a four-hour reagent on-board stability. This allows to abolish heparin interference and assess the basic TGT profile of a patient.
(1) Carlo A et al. ISTH 2009 PP-WE-164
To cite this abstract in AMA style:
Fontaine S, Bourdin M, Bildan M-, Kerleau O, Lambert M, Depasse F, Mathieu O. In vitro Neutralization of Unfractionated Heparin for the Measurement of Thrombin Generation with the STG-drugScreen Reagent on ST Genesia [abstract]. Res Pract Thromb Haemost. 2021; 5 (Suppl 1). https://abstracts.isth.org/abstract/in-vitro-neutralization-of-unfractionated-heparin-for-the-measurement-of-thrombin-generation-with-the-stg-drugscreen-reagent-on-st-genesia/. Accessed September 16, 2021.« Back to ISTH 2021 Congress
ISTH Congress Abstracts - https://abstracts.isth.org/abstract/in-vitro-neutralization-of-unfractionated-heparin-for-the-measurement-of-thrombin-generation-with-the-stg-drugscreen-reagent-on-st-genesia/