The impact of sample site and storage on thromboelastography values
Authors: Mari Tuovila, Tiina Erkinaro, Eeva‐Riitta Savolainen, Paivi Laurila, Pasi Ohtonen, Tero Ala‐Kokko
Journal: American Journal of Hematology (2017)

Thromboelastography (TEG) provides information on the entire clotting system by measuring viscoelastic properties of whole blood during the coagulation process. The advantage of TEG over routine laboratory tests is the rapid assessment of coagulation and fibrinolysis while considering the role of all blood components. Since the use of TEG is expanding outside the operating rooms, we intended to get further evidence whether the use of different sample sites (arterial-venous) result in similar TEG values. Furthermore, we assessed the effect of citrate storage on TEG values. After approval by the Ethical Committee of the Northern Osthrobothnia Hospital District, we recruited 51 ASA II-IV surgical patients aged 18-85 years in Oulu University Hospital in 2013-2014. Written informed consent was obtained from all subjects. Patients with a known pre-existing bleeding tendency or who were taking aspirin, an antiplatelet agent, warfarin, or any other drug to influence coagulation were excluded. On arrival into the operating room, four arterial and four venous samples were drawn from each patient into 2.7-mL Vacutainer tubes containing 3.2% citrate before an intravenous line was inserted to avoid any influence of intravenous infusion or drugs on our results. Arterial samples were obtained from an arterial cannula inserted in the radial artery and venous samples were simultaneously drawn by a direct venipuncture from the antebrachial vein. The first sample from each site was discarded leaving three arterial and three venous samples for analysis. The Vacutainer tubes were labeled in the order of collection and compared respectively. Blood samples were analyzed 30 min, 60 min, and 120 min after collection. All samples were stored at room temperature and re-calcified before analysis. No other laboratory testing were taken. The TEG analyses were performed according to manufacturer's guidelines by a single trained study nurse. Kaolin was used as an initiator in each test. Two computerized TEG® analyses (TEG 5000 Thromboelastograph® Hemostasis System, Haemonetics Corporation, Niles, USA) with four separate channels were used. TEG parameters used for this study were reaction time (R) and maximum amplitude (MA). Patients acted as their own controls, ie blood samples were compared only with individual's own samples. The data was analyzed using SPSS for Windows (IBM Corp., released 2012: IBM SPSS Statistics for Windows, Version 21.0. Armonk, NY: IBM Corp.). The Bland–Altman method was used to quantify the mean bias between measurements with 95% limits of agreement and the percentage errors (mean, median) were calculated. The main results are shown in Table 1. Altogether, 306 blood samples were collected from 51 patients undergoing elective surgery. There were 25 men and 26 women with a mean age of 65 years (range: 30-81 years). The ASA risk score was 2 in 35 patients, 3 in 15 patients and 4 in 1 patient. The patients had the following comorbidities: hypertension (44%), coronary heart disease (8%), diabetes (20%), asthma (24%), and obesity (43%). The following surgical procedures were performed: Laparoscopic splenectomy (1), small bowel resection (1), cholecystectomy (2), laparoscopic fundoplication (2), laparoscopic rectocolpopexy (1), spondylodesis (18), decompression (10), laminectomy (4), foraminotomy (1), microdiscectomy (1), and robotic assisted prostatectomy (10). Forty-six complete series (3 arterial and 3 venous samples) were obtained. In five patients, at least one trace was excluded due to technical TEG channel failure. The TEG trace was abandoned if there was not any numeric value for the analysis. The other traces of these patients were used in the analysis. A total of 239 samples were used for comparisons. The mean R in all samples was 8.2 min (range 3.2-15.6 min, SD 1.8) and the mean MA was 65.2 mm (range 49.3-77.0 mm, SD 6.1). For arterial–venous comparison, 30 min samples were used. The mean percentage error between sample pairs was 17.1% (median 9.1%) for R and 5.5% (median 3.5%) for MA. The Bland–Altman figures were created for quantifying the bias between arterial and venous samples, Supporting Information Figure 1. The figure represents larger variation between arterial and venous samples in R in patients with short R time. The effect of storage was assessed by comparing T60 min and T120 min samples to T30 min samples. In arterial samples, the mean percentage error between sample pairs varied from 11 to 12% for R (median 7.9-8.7%) and from 3.9 to 4.0% for MA (median 2.9-3.0%). In venous samples the mean percentage error between sample pairs varied from 14.1 to 14.7% (median 10.1-11.4%) for R and from 4.9 to 5.3% (median 3.3-4.0%) for MA. Bland–Altman figures illustrating the impact of storage on R and MA are given in Supporting Information Figures 2 and 3. Previously, only a few studies have compared TEG parameters of arterial and venous blood samples with conflicting results.1-3 It has been suggested that the blood sample site might cause differences in TEG values.2, 3 In our study, the mean bias between arterial and venous blood samples (Ta30 vs. Tv30) was low. However, in some sample pairs, the variation in R was large, resulting in a high mean percentage error of 17%. Yet, the median percentage error was 9%, reflecting the significance of these single sample pairs to our results. A larger variation in R between arterial and venous blood samples was detected in patients with a short R time suggesting rapid initiation of the coagulation process. Others have suggested that arterial and venous coagulation differences detected in previous studies could be explained by shear stress. A shorter diameter of the medical device used for blood sample collection may result in faster (shorter R) and stronger (larger MA) coagulation.4 However, conflicting results have also been published.3 In our study, we did not investigate the effect of shear stress on TEG values. However, blood sample collection from an arterial line yielded slightly more stable TEG values. This probably reflects variability concerning blood sample collection by venipuncture rather than shear forces. Citrated blood samples were used in our series while previous studies investigating arterial–venous differences in TEG values used native blood samples.1-4 Furthermore, none of these studies used direct venipuncture as a sampling method. Therefore, methodological differences may partly explain the discrepancies. Other studies have not reported the effect of the duration of R on repeated TEG analyses. The large variation in R in patients with short R values in our series reflects the importance of prompt processing of the sample after re-calcification of citrated blood samples. There have been reports of a tendency toward acceleration of coagulation in TEG traces of citrated blood samples over time.5 In our series, however, storage had no significant effect on R or MA values. We were not able to detect any tendency toward hypercoagulation during 120 min of storage. This discrepancy may be due to the initiator used. Earlier studies demonstrated that using kaolin as an initiator in re-calcified citrated blood yields more stable TEG values compared with other initiators.6 The TEG analysis is a sensitive method which is highly dependent on manual processes. The generally accepted principles of sample collection and handling have to be carefully followed. Under or overfilling of the Vacutainer tube affects the citrate concentration in the sample. Furthermore, this can lead to inadequate re-calcification of the sample, which may influence the initiation of the coagulation, ie R. Also, any delay or error in sample preparation (eg, pipetting) may affect R time. In conclusion, there was more variation in R than in MA in all sample pairs at each time point. Arterial samples yielded more stable TEG values. However, the variation in venipuncture samples was acceptable. When citrate containing blood samples are used, it is important to make sure the sample is processed adequately. A slightly prolonged R may be due to inadequate re-calcification and the clinical situation should be considered. Citrated blood samples can be stored up to 120 min. This encourages locating the analyzer in a central laboratory, to expand the use of TEG outside of the operating rooms. The authors appreciate the expertise of study nurse, Sinikka Sälkiö, in sample logistics and the TEG method. Nothing to report. Additional Supporting Information may be found online in the supporting information tab for this article. FIGURE S1 Bland–Altman figure of (a) reaction time (R) and (b) maximum amplitude (MA) in citrated blood samples after 30 min storage: difference (venous – arterial) versus average of venous and arterial samples with 95% LoA. FIGURE S2 Bland–Altman figure of reaction time (R) in citrated blood samples after 30 min vs. 60 min and 30 min vs. 120 min storage: difference (60 min – 30 min) and difference (120 min – 30 min) versus respective averages, with 95% LoA. Arterial samples in (a) and (b), venous samples in (c) and (d). FIGURE S3 Bland–Altman figure of maximum amplitude (MA) in citrated blood samples after 30 min vs. 60 min and 30 min vs. 120 min storage: difference (60 min – 30 min) and difference (120 min – 30 min) versus respective averages, with 95% LoA. Arterial samples in (a) and (b), venous samples in (c) and (d). 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