Frozen/Thawed Samples Can Replace Fresh Samples for Assignment of ISI to Secondary Thromboplastin Standards for Multiple Reagent/Instrument Combinations: Data to Support Possible Revision of WHO Guidelines

Background: Calibration of thromboplastins is required for accurate calculation of the international normalised ratio (INR). Accurate INR results are required for optimal dosing of vitamin K antagonists. Decreases in vitamin K antagonist usage have made the recruitment of sample sets for international sensitivity index (ISI) calibrations more difficult. A possible solution to this would be to allow the use of frozen-thawed samples in place of fresh plasmas in the calibration of secondary standards.

Objectives: We investigated the effect of freezing and thawing samples before usage in ISI calibrations of secondary standards.

Methods: Multiple reagent/instruments were tested to identify the degree of difference between a fresh sample ISI calibration and one performed on frozen-thawed samples. Where possible, the two ISI calibrations were performed on the same sample set. Alternatively, a separate set of samples from different patients was used.

Results: The difference in ISI values was <3% for those datasets where the same samples were used, and <6% for those datasets where two sample sets were used. Additionally, other parameters required for a valid ISI calibration showed only minor differences-some calibrations showed fewer outliers in the frozen-thawed datasets. Mean normal prothrombin time for the international reference thromboplastins was <3.5% different across four different calibrations (two for rabbit thromboplastin and two for recombinant human thromboplastin).

Conclusions: This modification to the WHO guidelines would facilitate the recruitment of test plasmas in advance of calibration solving the problem of requiring availability of fresh patient samples with a range of INRs in a 5-h window.

Trial registration: Not a part of any clinical trial.