Standardization of prothrombin fragment 1.2 measurement: effects of preanalytical variables and calibrator selection.

Author(s): Hursting MJ, Slaughter TF, Stead AG, Szewczyk KM, Witt DJ, Greenberg CS

Publication: Arch Pathol Lab Med, 1998, Vol. 122, Page 31-6

PubMed ID: 9448013 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of blood collection method, type of anticoagulant, and calibrator set on F1.2 measurements using three different commercially available ELISAs. Blood was collected into either sodium citrate, lithium heparin, or a cocktail of including the anticoagulant EDTA, with aprotinin, and PPACK, referred to as EAP. Platelet-poor plasma was prepared within 1 hour, and specimens intended for comparison of the Baxter and OTC assays were stored at 2-8 degrees C for up to 4 hours before assay, while specimens intended for comparison of the Behring and OTC assays were stored at -70 degrees C for an unspecified amount of time, and specimens containing heparin or EAP were mixed with a solution containing heparin, EDTA, and citrate that adjusted specimen pH to 5.8-6.2 prior to frozen storage.

   Summary of Findings:

   The Behring ELISA recommends the use of citrate and the OTC ELISA recommends use of heparin as anticoagulants. Correlations in plasma F1.2 measurements between these two assays, when the recommended anticoagulants were used, were modest for both normal specimens (r=0.49) and those from diseased patients (r=0.48). However, when the same anticoagulant was used for each assay, interassay correlations were higher (r=0.62, citrate; r=0.76, EAP; and r=0.88, heparin). The Baxter ELISA recommends the use of heparin, similar to the OTC ELISA. These two assays showed a very strong correlation when heparinated specimens were used (r=0.94). Interassay and intraassay correlations between anticoagulant pairs were generally higher for the metal-ion chelators, citrate and EAP, as opposed to comparisons with heparin. Collection of specimens through an arterial catheter, rather than by venipuncture, led to very strong correlations between the Behring and OTC assay results when the recommended anticoagulants were used (r=0.96), when citrate was used for both assays (r=0.96), or when heparin was used for both assays (r=0.97). When calibrators from one ELISA were used for a different ELISA, significant differences in F1.2 measurements were observed in all cases.

   Biospecimens

   • Bodily Fluid - Plasma
   • Bodily Fluid - Blood

   Preservative Types

   • Frozen

   Diagnoses:

   • Normal
   • Coronary Artery Disease
   • Not specified
   • Other diagnoses

   Platform:

   Analyte	Technology Platform
   Protein	ELISA

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Acquisition	Method of fluid acquisition	Arterial catheter Venipuncture
   Preaquisition	Diagnosis/ patient condition	Thrombosis Trauma patient Surgical patient Sickle cell disease
   ELISA Specific	Technology platform	Behring Enzygnost F1+2 ELISA OTC Thrombonostika F1.2 ELISA Baxter Prothrombin Fragment F1.2 ELISA
   ELISA Specific	Reaction solution	Behring calibrators OTC calibrators Baxter calibrators
   Analyte Extraction and Purification	Protease inhibitor	Aprotinin No protease inhibitor added PPACKI
   Biospecimen Acquisition	Anticoagulant	PPACK Lithium heparin EDTA Sodium citrate
   Biospecimen Acquisition	Anatomical location of blood draw	Artery Vein

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