Paving the way for establishing a Reference Measurement System for standardization of plasma Prothrombin Time: harmonizing the Manual Tilt Tube method.

BACKGROUND: International Normalized Ratio (INR) is traceable to World Health Organization (WHO) International Standards for thromboplastins. International Standards must be used with a Manual Tilt Tube technique (MTT) for prothrombin time (PT) determination. An important part of the total variability of INR is due to poor harmonization of MTT across WHO reference laboratories. OBJECTIVES: To determine the origins of PT differences between operators performing MTT, and to develop a harmonized MTT. METHODS: Two workshops were held where WHO reference laboratory operators could compare their PTs using MTT and the same equipment. A harmonized MTT was used by 7 operators in the second workshop. RESULTS: Differences have been observed in tilting frequency, and in the height of pipetting plasma in the test tube. At the beginning of the first workshop the tilting cycle time varied between 1.1 and 2.7 s. The mean PT of normal plasma obtained by pipetting plasma at the top of the tube was 14.3 s but was 12.9 s when plasma was pipetted at the bottom of the tube. When using the harmonized MTT for WHO International Standard rTF/16, the differences between operators were not greater than 1.1 s in normal plasma, and not greater than 1.3 s in patient plasma with average INR of 3.0. INR between-operator coefficient of variation was 2.3 %. CONCLUSION: Application of a harmonized MTT in three reference laboratories resulted in substantial reduction of between-operator variation of PT and INR. The harmonized MTT is proposed as Candidate Reference Measurement Procedure.