Improved Method for the ASTM Platelet and Leukocyte Assay: Use of Minimal Heparinization in a Screening Test for Hemocompatibility of Blood-Contacting Medical Devices

Abstract

Most blood-contacting medical devices must be assessed for potential thrombogenicity prior to regulatory approval. A common assay for screening and qualifying devices involves monitoring the reduction of platelet and leukocyte (P&L) counts in whole blood exposed to the device. We have validated an improved method for assessing a device's effect on platelet activation and surface adhesion, offering significant improvement over the current ASTM F2888-13 method, which uses blood fully anticoagulated by acidified citrate (known to significantly inhibit platelet responsiveness). Our method uses minimal heparinization (final concentration 1 IU/mL) to optimize the response to commonly used control materials: latex, black rubber, and high-density polyethylene (HDPE). We also have shown the assay's capacity to appropriately assess a legally marketed comparator device (LMCD) with a documented clinical history. The test materials were prepared for incubation and allowed to remain in contact with the citrated or heparinized blood for ∼1 h at 37 °C. A complete blood count was performed prior to exposure, and at the end of the incubation period, reductions in P&L counts were recorded. Results from citrate-anticoagulated assay showed only a marginal response to the positive control, black rubber. Using heparinized blood, the assay generated a robust response to the positive controls, the “intermediate scoring” controls, and also assessed a legally marketed and approved device as clearly nonthrombogenic. This modification adds robustness and sensitivity to this quick and inexpensive thrombogenicity assay and should be incorporated into the next ASTM standards.

Platelet and leukocyte count assay for thrombogenicity screening of biomaterials and medical devices: Evaluation and improvement of ASTM F2888 test standard

“…However, the difference was more pronounced for 1 U/ml, suggesting a greater test sensitivity for thrombogenicity evaluation at the lower heparin concentration. Our platelet count results using low‐dose heparin are consistent with a recent study by Schatz et al, 16 in which 1 U/ml heparin anticoagulation substantially improved test sensitivity for discriminating between materials when compared to sodium citrate anticoagulation. Using a heparin concentration below 1 U/ml is not recommended for this assay as pretest blood clot formation was already observed at 1 U/ml heparin for a small portion (< 10%) of blood donors, and the pretest clotting rate would be higher at a lower heparin concentration.…”

The use of scanning electron microscopy and fixation methods to evaluate the interaction of blood with the surfaces of medical devices

“…Careful fixation of blood is essential due to its instability. To ensure a reliable assessment of blood interaction with the tested material, appropriately low concentrations of anticoagulant and stabilizing agents have to be used 7 .…”

The use of scanning electron microscopy and fixation methods to evaluate the interaction of blood with the surfaces of medical devices

“…Careful xation of blood is essential due to its instability. To ensure a reliable assessment of blood interaction with the tested material, appropriately low concentrations of anticoagulant and stabilizing agents have to be used [7].…”