Performance Evaluation of the Plateletworks in the Measurement of Blood Cell Counts as compared to the Beckman Coulter Unicel DXH 800
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1 The Journal of ExtraCorporeal Technology Technique Articles Performance Evaluation of the Plateletworks in the Measurement of Blood Cell Counts as compared to the Beckman Coulter Unicel DXH 800 Erick McNair, CCP, PhD, FICA;* A. Mabood Qureshi, MSc, FCACB;* Cara Bally, BSc* *Departments of Pathology and Laboratory Medicine and Surgery, Division of Cardiothoracic Surgery, College of Medicine, University of Saskatchewan, Saskatoon Health Region, Saskatchewan, Canada Abstract: Prior to undergoing cardiac surgery many patients may have impaired platelet function due to platelet inhibition. Point of care testing (POCT) that produces quick results of platelet counts and function allow earlier clinician interpretation, diagnosis and treatment. Before being adopted for routine clinical use, a POCT device s performance must be evaluated by standard laboratory techniques to ensure high quality results. The purpose of this study is to determine the performance of the Plateletworks BC 3200 automated hematology analyzer by correlating its precision, accuracy and linearity for the measurement of blood counts to our hospital central laboratory analyzer (Beckman Coulter Unicel DXH 800). The study utilizes well described methods for Within- Run and Day-to-Day precision, comparison of methods (bias), and linearity. Control samples from the manufacturer were used for the precision studies, blood samples from 115 cardiac surgical subjects were used for comparison of methods and accuracy, and pre diluted control samples from the manufacturer were used for the linearity studies. The precision of the Plateletworks analyzer was acceptable. The overall coefficient of variation (CV) for the measured parameters at all levels of control for Within-Run precision was acceptable ranging from %. Likewise, the CV for the measured parameters at all levels of control for Dayto-Day precision was acceptable ranging from 1.45% to 6.7%. The correlation and accuracy between the two analyzers for the evaluated parameters (platelets, red blood cells, white blood cells, and hemoglobin) was acceptable. The linearity for the measured parameters was also acceptable with a range between %. The performance of the Plateletworks analyzer was acceptable for providing blood cell counts as compared to our central hospital laboratory analyzer. Keywords: performance, point-of-care testing, platelet counts, correlation, precision, accuracy, linearity The ischemic events occurring in acute coronary syndrome (ACS) patients result in platelet aggregation, leading to thrombus formation and plaque instability (1). Percutaneous coronary intervention (PCI) causes further endothelial damage and platelet activation. On the activated platelet, the central sites for the development of the procoagulant process are the surface receptors (2). The use of dual antiplatelet therapy (DAPT) has significantly Received for publication December 16, 2014; accepted July 1, Address correspondence to: Erick McNair, CCP, PhD, FICA, Department of Pathology and Laboratory Medicine, College of Medicine, Health Science Building, Room 2D01, Royal University Hospital, 107 Wiggins Road, University of Saskatchewan, Saskatoon, SK, S7N 5E5, Canada. erick.mcnair@usask.ca The senior author has stated that the authors have reported no material, financial, or other relationship with any healthcare-related business or other entity whose products or services are discussed in this paper. reduced the thrombotic complications associated with the adverse cardiovascular outcomes of PCI (3). However, there is a significant amount of antiplatelet nonresponders that need to be identified through receptor testing (4). Many patients referred for cardiothoracic surgery with cardiopulmonary bypass (CPB) have undergone at least one, if not multiple, PCI procedures and are subsequently administered long-term DAPT (5). In addition, CPB-induced depletion of clotting factors and platelet dysfunction increase the risk of postsurgical bleeding (5). The combined longterm DAPT and the risk of CPB-induced platelet dysfunction may place this population of patients at increased risk for postoperative bleeding. Therefore, a point-of-care testing (POCT) device capable of dual determination of platelet counts and platelet reactivity, which can be used for patients undergoing PCI and those undergoing cardiac surgery, is needed. 113
2 114 E. MCNAIR ET AL. The dual performance of the Platelet Works BC 3200 automated hematology analyzer (Helena Laboratories, Beaumont, TX) is dependent upon accurate platelet counts. Since this analyzer has not been well studied, the purpose of this investigation is to validate its correlation with our central hospital laboratory analyzer. The objectives of this study are to examine its precision, accuracy, and linearity in measuring blood counts as the initial steps toward verifying its clinical use in our patient population. MATERIALS AND METHODS This study was approved by the University of Saskatchewan Ethics Review Board and received operational approval from the Saskatoon Health Region. All participants signed an informed consent. Table 1 gives a physical description and characteristics of the Platelet Works BC 3200 automated hematology analyzer and the Beckman Coulter Unicel DXH 800 automated hematology analyzer (Beckman Coulter Canada LP, Mississauga, Canada). The gold standard for the measurement of platelet function is light transmittance aggregometry. The disadvantages of this technique are poor reproducibility, requirement of sample preparation, time consumption, and expense (3,6). The Plateletworks BC 3200 (Helena Laboratories) is a point-of-care testing analyzer and diagnostic screening assay. It can be used to assess patient primary hemostasis by measuring the platelet count and examining platelet reactivity by determining the percentage of platelet aggregation or inhibition using three agonists adenosine diphosphate (ADP), collagen, and arachidonic acid (7). The analyzer uses the impedance method to correlate platelet counts from a baseline sample of blood in an ethylenediaminetetraacetic acid (EDTA) tube with platelet counts in a citrate tube after aggregation with either ADP or collagen. The addition of blood to the agonist tubes stimulates platelets to activate and adhere to the tube whereby they are eliminated from the platelet count. The ratio of the activated platelet count to the nonactivated platelet count is a function of the reactivity of platelets in the sample. The percentage of platelet aggregation or inhibition is based on actual baseline and agonist counts (7,8). Although both analyzers produce several measured and calculated parameters, we chose to evaluate the directly measured blood cell counts of platelet (PLT), white blood cell (WBC), red blood cell (RBC), and hemoglobin (Hgb) for this study. Our central hospital laboratory uses a Beckman Coulter Unicel DXH 800 analyzer (Beckman Coulter Canada) that also uses the impedance method (9). To ascertain whether the Plateletworks analyzer had an acceptable performance relative to blood cell counts as compared to the Table 1. A physical description and characteristics of the Beckman Coulter Unicel DXH 800 automated hematology analyzer and the Platelet Works BC 3200 automated hematology analyzer. Specifications Unicel DXH 800 Plateletworks BC 3200 Height cm 45.7 cm Weight kg 25 kg Depth cm 42.5 cm Power Voltage: Voltage: Frequency: Hz Frequency: Hz Input Power: 180 VA Input Power: 180 VA Sample vol. 165 ml 13 ml Cartridge size 5 tubes 4 tubes Analysis time 2 3 minutes 2 minutes Directly measured parameters (operating range) WBC ( /ml)wbc ( /ml) RBC ( /ml)rbc ( /ml) Hgb ( g/dl) Hgb ( g/dl) PLT ( /ml)plt ( /ml) Calculated parameters (ref. range) Lymphocyte ( /ml)lymphocyte ( /ml) Neutrophils ( /ml)granulocyte ( %) Monocyte ( /ml)hct ( %) Eosinophils ( /ml) MCV ( fl) Basophils ( /ml)mch ( pg) HCT ( %) MCHC ( g/dl) MCV (79 99 fl) RDW ( %) MCH (27 32 pg) MPV ( fl) MCHC ( g/dl) RDW ( %) MPV ( fl) NRBC ( +10/mL) NRBC/100 WBC cm, centimeters; HCT, hematocrit; Hz, hertz; kg, kilograms; MCH, mean corpuscular hemoglobin; MCHC, mean corpuscular hemoglobin concentration; MCV, mean corpuscular volume; MPV, mean platelet volume; ml, microliter; RDW, red blood cell distribution width; NRBC, nucleated red blood cells; VA, voltage alternating current; vol, volume.
3 PERFORMANCE EVALUATION OF THE PLATELETWORKS 115 Table 2. The expected results for the external quality control material at the three standardized levels of control. Expected Results from External Quality-Control Material Parameter Units Low Medium High Platelets 10 3 /ml 79 ± ± ± 45 WBC 10 3 /ml 2.5 ± ±.7 21 ± 1.3 RBC 10 6 /ml 2.40 ± ± ±.2 Hb g/dl 6.0 ± ± ± Results are expressed as ±SD. central lab analyzer we measured its 1) precision, 2) accuracy (bias), 3) total error, and 4) linearity. The within-run precision of the Plateletworks analyzer was examined by performing 20 replicate analyses of external quality control material at three standardized levels: low, medium, and high. Table 2 depicts the expected results for the external quality control material at the three standardized levels. The same operator also performed day-to-day precision, which consisted of replicate analysis of the three levels of control over a 20-day period. The quality control material was provided in premixed, equivalently concentrated, vials (same lot number).the mean (m), standard deviation (SD), and coefficient of variation (CV) were calculated. The accuracy of the Plateletworks analyzer was measured by means of a comparison evaluation. The closer the agreement between the measured values produced by the Plateletworks analyzer and those of the central lab analyzer, the greater the accuracy of the analyzer. This was accomplished by obtaining 6 ml of whole blood from 115 patients undergoing cardiac surgery at the following time intervals: pre-cpb, during-cpb, and post-cpb. Blood was aliquoted into EDTA tubes immediately after samples were collected. Samples were transported to the central lab (within minutes) for the simultaneous measurement of PLT, WBC, RBC, and Hgb counts on the Plateletworks and central lab analyzers. The Bias was detected by calculating the average difference of the means (observed expected). The resultant correlations were ranked on the basis of standard ranges as follows: excellent for Spearman rho (r s ) =.93.99, good for (r s ) =.80.92, fair for (r s ) =.59.79, and poor for (r s ) <.59 (10). Total error was calculated using the following formula: bias + 3 +SD (10). Linearity was assessed with a commercially available kit (R&D Systems, Minneapolis, MN) that contained prediluted (5, 10, 20, 50, 70, and 100%) samples (PLT, WBC, RBC, and Hgb). The measured parameters for linearity were as follows: 1) WBC low-level linearity, 2) WBC linearity, 3) platelet low-level linearity, 4) platelet linearity, 5) RBC low-level linearity, 6) RBC linearity, 7) Hgb lowlevel linearity, and 8) Hgb linearity. Linearity was evaluated by performing linear regression analysis between the values provided by the manufacturer (expected) and the measured (obtained) values from the Plateletworks analyzer. There were four runs and six levels of dilution for each measured parameter. All statistical analyses were performed using the Statistical Package for the Social Sciences (SPSS) version 13 (SPSS Inc., Chicago, IL) and Microsoft Office Excel 2008 (Microsoft Corp., Redmond, WA). RESULTS The within-run and day-to-day precision data are shown in Table 3. For all levels of control, the within-day replicate analysis demonstrated a low intra-assay CV for the following: Hb ranging from.65% to 1.1%, RBCs ranging from.97% to 1.61%, WBCs ranging from 1.23% to 6.4%, and platelets ranging from 2.38% to 5.13%. The overall CVs with-in day for the measured parameters at all levels of control were acceptable. For all levels of control, the dayto-day replicate analysis demonstrated a low CV for the following: Hgb ranging from.65% to 1.3%, RBCs ranging from.99% to 1.83%, WBCs ranging from 1.4% to 6.7%, and platelets ranging from 2.56% to 6.69%. For all levels of control material, the day-to-day CVs for the measured parameters were below 3.7% except for platelets in the low control with a CV of 6.7%, and WBCs in the low control Table 3. The within-day and day-to-day precision data of the measured parameters for the three levels of Control. Low Within-Run Medium High Low Day-to-Day Medium High Mean Mean Platelets SD SD CV CV Mean Mean WBC SD SD CV CV Mean Mean RBC SD SD CV CV Mean Mean Hb SD SD CV CV
4 116 E. MCNAIR ET AL. Figure 1. The correlation, mean, bias, and R-value for PLT and WBC. The results are expressed as ±SD ml, microliter; R, Pearson correlation coefficient; g/dl, grams per deciliter. WBC, white blood cell. with a CV of 6.7%. The overall ranking of CVs for the measured parameters at all levels of control was acceptable. The accuracy of the Plateletworks analyzer was measured by means of a comparison evaluation. Linear regression was used to produce the graphical plot of the Plateletworks values vs. the central lab values. The correlation, mean, bias, and R-value for PLT, WBC, RBC, and Hgb are shown in Figures 1 and 2. This data demonstrates that the correlation between the Plateletworks and the central lab analyzer was acceptable for the measured parameters (.97.99). The total errors for platelets, WBC, Hgb, and RBC were 2.3%, 1.3%, 2.6%, and 1.3% respectively, which demonstrated values within the documented allowable error (10,11). The measure parameters displayed acceptable correlation between the Plateletworks and the central hospital laboratory analyzer. The linearity for the measured variables was acceptable with obtained R values (Pearson Correlation) ranging between.998 and.999. The slopes of the regression curve produced values which ranged between.953 and 1.03, and the intercept with values ranging between.0008 and (all graphs not shown). Table 4 shows the platelet low-level linearity for the measured levels of dilution and the correlation of expected mean (prediluted samples) vs. observed mean (Plateletworks analyzer). At low levels of dilution (5% and 10%) there was decreased agreement. Overall, the linearity for the measured parameters was acceptable with a range between 98% and 100%. DISCUSSION The course of patient treatment is dependent upon the precision and accuracy of results from analyzers of the central laboratory and POCT devices. To date, there have Figure 2. The correlation, mean, bias, and R-value for RBC and Hgb. The results are expressed as ±SD. RBC, red blood cell; Hgb, hemoglobin; ml, microliter; R, Pearson correlation coefficient; g/dl, grams per deciliter.
5 PERFORMANCE EVALUATION OF THE PLATELETWORKS 117 Table 4. The platelet low-level linearity for the measured levels of dilution and the correlation of expected mean (pre-diluted samples) vs. observed mean (Plateletworks analyzer). Dilution (%) Run 1 Run 2 Run 3 Run 4 Obtained Mean Expected Mean Difference ml, microliter. been no independent reports on precision and accuracy with relationship to blood cell counts of this POCT device with a central laboratory analyzer. Platelet counts during cardiac surgery with CPB are significantly reduced because of hemodilution. Thus, the ability to measure low blood count levels is of importance. The CV has been the most important statistic to determine the precision of experimental errors (12,13). Based upon the previously discussed values, the overall precision of the Plateletworks analyzer was acceptable for within-day and day-to-day precision. The comparison of methods of tests showed acceptable correlation to the central lab analyzer with R-values ranging from.97 to.999 for the measured parameters. The total error for all measured parameters fell within the limits of allowable error (10,11). The platelet low-level linearity had acceptable correlation with an R-value of.999. However, the difference between the obtained and expected mean values for 5% and 10% dilution ranged from 4.8 to 3.8 indicating diminished agreement at extremely low levels of platelets (less than /ml). Thus, caution may be needed when interpreting and treating results with very low platelet values. The prevention of thrombotic complications in patients with high-risk coronary artery disease has traditionally been treated with DAPT consisting of either clopidogrel and aspirin or prasugrel and aspirin. The American College of Cardiology Foundation/American Heart Association s (ACCF/AHA) Guidelines for the Management of Patients with Unstable Angina/Non-ST-Elevation Myocardial Infarction has recommended the withdrawal of clopidogrel for 5 days and prasugrel for 7 days before a planned coronary artery bypass graft (CABG) surgery to allow recovery of platelet function (3). However, it has been demonstrated that in selected patients, platelet function may resume before the suggested 5-day discontinuation period (14). Furthermore, it has been shown that preoperative platelet function monitoring in patients on DAPT requiring CABG decreases the suggested preoperative waiting period and results in similar bleeding as compared to those patients without DAPT (15). This suggests that the fixed waiting period usually accompanied by in-hospital withdrawal from DAPT may not be beneficial to patient well-being or the health-care system because of the increased costs associated with increased length of stay. Therefore, a POCT device that can provide the dual performance of platelet counts and testing of platelet receptor inhibition could be a valuable tool to the clinician by providing information that may shorten the patient waiting time for CABG. CONCLUSION The Plateletworks POCT analyzer assesses patient primary hemostasis by measuring platelet counts and testing platelet reactivity within minutes. It produces acceptable precision, accuracy, and linearity for the blood cell counts measured in this study as compared to the central hospital laboratory analyzer (Beckman Coulter DXH 800) except at very low platelet values. This analyzer may be an effective tool for use in the cardiac interventional and surgical settings. However, we suggest that the next step in the verification of this analyzer is to determine its performance in measuring the percentage of platelet aggregation or inhibition of receptors using agonists as compared to other reference methods. REFERENCES 1. Mangiacapra F, Barbato E. Clinical implications of platelet-vessel interaction. J Cardiovasc Transl Res. 2013;6: Pollack CV Jr, Antman EM, Hollander JE focused update to the ACC/AHA guidelines for the management of patients with STsegment elevation myocardial infarction: Implications for emergency department practice. Ann Emerg Med. 2008;52: Wright RS, Anderson JL, Adams CD, et al ACCF/AHA focused update incorporated into the ACC/AHA 2007 Guidelines for the Management of Patients with Unstable Angina/Non-ST-Elevation Myocardial Infarction: A report of the American College of Cardiology Foundation/American Heart Association Task Force on Practice Guidelines developed in collaboration with the American Academy of Family Physicians, Society for Cardiovascular Angiography and Interventions, and the Society of Thoracic Surgeons. J Am Coll Cardiol. 2011;57:e Tantry US, Gurbel PA. Antiplatelet drug resistance and variability in response: The role of antiplatelet therapy monitoring. Curr Pharm Des. 2013;19: Mercado N, Wijns W, Serruys PW, et al. One-year outcomes of coronary artery bypass graft surgery versus percutaneous coronary intervention with multiple stenting for multisystem disease: A meta-analysis of individual patient data from randomized clinical trials. J Thorac Cardiovasc Surg. 2005;130:512 9.
6 118 E. MCNAIR ET AL. 6. Michelson AD. Platelet function testing in cardiovascular diseases. Circulation. 2004;110:e Helena Laboratories. Platelet Works: Platelet Function Kits. Available at: Accessed January Enriquez LJ, Shore-Lesserson L. Point-of-care coagulation testing and transfusion algorithms. Br J Anaesth. 2009;103(Suppl 1):i Beckman Coulter Canada. Available at: Accessed January Burtis CA, Ashwood ER. Evaluation of methods. In: Burtis CA, Ashwood ER, Border B, Tietz NW (eds). Teitz Fundamentals of Clinical Chemistry, 5th ed. Philadelphia, PA: WB Saunders Company, 2001: Data Innovation. Available at: Accessed May Jensen AL, Kjelgaard-Hansen M. Method comparison in the clinical laboratory. Vet Clin Pathol. 2006;35: Arnhold E, Milani KF. Rank-ordering coefficients of variation for popping expansion. Acta Sci Agron. 2011;33: Gurbel PA, Bliden KP, Butler K, et al. Randomized double-blind assessment of the ONSET and OFFSET of the antiplatelet effects of ticagrelor versus clopidogrel in patients with stable coronary artery disease: The ONSET/OFFSET study. Circulation. 2009;120: Mahla E, Suarez TA, Bliden KP, et al. Platelet function measurementbased strategy to reduce bleeding and waiting time in clopidogrel-treated patients undergoing coronary artery bypass graft surgery: The timing based on platelet function strategy to reduce clopidogrel-associated bleeding related to CABG (TARGET-CABG) study. Circ Cardiovasc Interv. 2012;5:261 9.
NOTE: This table will be discontinued after this lot.
AS037-011 Rev. 11/14 ASSAY VALUES AND EXPECTED RANGES QCP DATA MONTHS: DEC, JAN, FEB Beckman Coulter STKS / MAXM / HMX LEVEL 1 + Lot No.: Exp. Date: LOT 871086 Parameter Mean Range WBC 10 3 /µl 4.0 ± 0.6
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