Complete Blood Count as point of care testing QBC STAR™: Preliminary evaluation.

INTRODUCTION: Point of care testing (POCT) represents a valuable option when laboratory data shall be urgently available for timely clinical management, with a turnaround time (TAT) that is unfeasible using conventional laboratory instrumentation. This study was aimed to compare the performance of QBC STAR™ compared to Sysmex XN-module and the reference optical microscopy (OM) assessment. MATERIAL AND METHODS: One hundred peripheral blood samples, collected in K3 EDTA tubes, and 50 capillary blood samples obtained by finger stick were analyzed with QBC STAR™, Sysmex XN-module, and OM. Data were compared with Passing-Bablok regression and Bland-Altman plots. RESULTS: The Passing-Bablok regression analysis (QBC STAR™ capillary sample vs XN-module) yielded slopes comprised between 0.30 and 1.37, while the intercepts ranged between -17.57 and 232.6. Bland-Altman plots yielded relative bias comprised between -4.87% (for MN QBC STAR™ capillary sample vs XN-module) and 27% (PLT QBC STAR™ capillary sample vs XN-module). A significant bias was found for all parameters except MN and WBC, RBC in all and pediatric samples, and HB in adults samples. CONCLUSION: The results of this analytical evaluation suggest that QBC STAR™ may not be the ideal tool for performing complete blood count analysis for diagnostic purposes, while it could be more useful in urgent/emergent conditions, such as for rapid monitoring of some hematological parameters (eg, WBC and HB).

What is the normal composition of pericardial fluid?

OBJECTIVE: Biochemical and cytological pericardial fluid (PF) analysis is essentially based on the knowledge of pleural fluid composition. The aim of the present study is to identify reference intervals (RIs) for PF according to state-of-art methodological standards. METHODS: We prospectively collected and analysed the PF and venous blood of consecutive subjects undergoing elective open-heart surgery from July 2017 to October 2018. Exclusion criteria for study enrolment were evidence of pericardial diseases at preoperatory workup or at intraoperatory assessment, or any other condition that could affect PF analysis. RESULTS: The final study sample included 120 patients (median age 69 years, 83 men, 69.1%). The main findings were (1) High levels of proteins, albumin and lactate dehydrogenase (LDH), but not of glucose and cholesterol (2) High cellularity, mainly represented by mesothelial cells. RIs for pericardial biochemistry were: protein content 1.7-4.6 g/dL PF/serum protein ratio 0.29-0.83, albumin 1.19-3.06 g/dL, pericardium-to-serum albumin gradient 0.18-2.37 g/dL, LDH 141-2613 U/L, PF/serum LDH ratio 0.40-2.99, glucose 80-134 mg/dL, total cholesterol 12-69 mg/dL, PF/serum cholesterol ratio 0.07-0.51. RIs for pericardial cells by optic microscopy were: 278-5608 × 106 nucleated cells/L, 40-3790 × 106 mesothelial cells/L, 35-2210 × 106 leucocytes/L, 19-1634 × 106 lymphocytes/L. CONCLUSIONS: PF is rich in nucleated cells, protein, albumin, LDH, at levels consistent with inflammatory exudates in other biological fluids. Physicians should stop to interpret PF as exudate or transudate according to tools not validated for this setting.

Multicenter evaluation of analytical performances of platelet counts and platelet parameters: Carryover, precision, and stability.

INTRODUCTION: The correctness of the results of automated platelet analysis is still highly debated. The aim of this multicenter study, conducted according to international guidelines, was to verify the analytical performance of nine different types of hematology analyzers (HAs) in the automated platelet analysis. METHODS: Four hundred eighty-six peripheral blood samples (PB), collected in K3 EDTA tubes, were analyzed by ABX Pentra, ADVIA2120i, BC-6800, BC-6800 Plus, Cell-DYN Sapphire, DxH800, XE-2100, XE-5000, XN-20 with PLT-F App. Within-run imprecision and between-run imprecision were carried out using PB and material control, respectively. The carryover, low limit of quantification (LoQ), and the PB stability were evaluated. RESULTS: The carryover was absent for all HAs. The LoQ of PLT ranged between 2.0 (Cell-Dyn Sapphire) and 25.0 × 109 /L (ADVIA 2120i), while immature platelet fraction (IPF) ranged between 1.0 (XN-20) and 12.0 × 109 /L (XE-5000). The imprecision (%CV) increases as the platelet count decreases. No HAs showed desirable CVAPS for PLT counts less than 50.0 × 109 /L, with the exception of Cell-DYN Sapphire (CV 3.0% with PLT-O mean value of 26.7 × 109 /L), XN-20 (CV 2.4% with PLT-F mean value of 21.5 × 109 /L), and BC-6800 Plus (CV 1.9% with PLT-O mean value of 26.5 × 109 /L). The sample stability ranged between under two hours for MPV by ADVIA2120i and 8 hours for other PLT parameters and HAs. CONCLUSION: The findings of this study may provide useful information regarding carryover, precision, and stability of platelet counts and parameters, especially in thrombocytopenic samples. Moreover, the stability of sample for platelet analysis is conditioned by the HA and by temperature and storage time.

Preliminary evaluation of a new flow cytometry method for the routine hematology workflow.

Background In a generalist laboratory, the integration of the data obtained from hematology analyzers (HAs) with those from multiparametric flow cytometry (FMC) could increase the specificity and sensitivity of first level screening to identify the pathological samples. The aim of this study was to perform a preliminary evaluation of a new simple hybrid method (HM). The method was obtained by integration between HAs reagents into FCM, with a basic monoclonal antibodies panel for the leukocytes differential count. Methods Eighty-one peripheral blood samples, collected in K3EDTA tubes, were analyzed by XN-module, and CyFlow Space System, using both standard MoAbs and HM method analysis, and with the optical microscopy (OM). Within-run imprecision was carried out using normal samples, the carryover was evaluated, data comparison was performed with Passing-Bablok regression and Bland-Altman plots. Results The within-run imprecision of HM methods ranged between 1.4% for neutrophils (NE) and 10.1% for monocytes (MO) always equal or lower to the OM. The comparison between HM methods vs. OM shows Passing-Bablok regression slopes comprised between 0.83 for lymphocyte (LY) and 1.14 for MO, whilst the intercepts ranged between -0.18 for NE and 0.25 for LY. Bland-Altman relative bias was comprised between -12.43% for NE, and 19.77% for eosinophils. In all 11 pathological samples the agreement between the methods was 100%. Conclusions The new hybrid method generates a leukocytes differential count suitable for routine clinical use and it is also useful for identifying morphological abnormalities with a reduction in cost and improvement of screening for first level hematology workflow.

Comparison between optical microscopy and automation for cytometric analysis of pericardial fluids in a cohort of adult subjects undergoing cardiac surgery.

AIMS: Limited information is available on number and type of cells present in the pericardial fluid (PF). Current evidence and has been garnered with inaccurate application of guidelines for analysis of body fluids. This study was aimed at investigating the performance of automate cytometric analysis of PF in adult subjects. METHODS: Seventy-four consecutive PF samples were analysed with Sysmex XN with a module for body fluid analysis (XN-BF) and optical microscopy (OM). The study also encompassed the assessment of limit of blank, limit of detection and limit of quantitation (LoQ), imprecision, carryover and linearity of XN-BF module. RESULTS: XN-BF parameters were compared with OM for the following cell classes: total cells (TC), leucocytes (white blood cell [WBC]), polymorphonuclear (PMN) and mononuclear (MN) cells. The relative bias were -4.5%, 71.2%, 108.2% and -47.7%, respectively. Passing and Bablok regression yielded slope comprised between 0.06 for MN and 5.8 for PMN, and intercept between 0.7 for PMN and 220.3 for MN. LoQ was comprised between 3.8×106 and 6.0×106 cells/L for WBC and PMN. Linearity was acceptable and carryover negligible. CONCLUSIONS: PF has a specific cellular composition. Overall, automated cell counting can only be suggested for total number of cells, whereas OM seems still the most reliable option for cell differentiation.

Lack of harmonization in high fluorescent cell automated counts with body fluids mode in ascitic, pleural, synovial, and cerebrospinal fluids.

INTRODUCTION: Cellular analysis in body fluids (BFs) provides important diagnostic information in various pathological settings. This study was hence aimed at comparing automated cell count obtained with Mindray BC-6800 (BC-BF) vs Sysmex XN-series (XN-BF) and evaluating other quantitative and qualitative information provided by these analyzers in ascitic (AF), pleural (PF), synovial (SF), and cerebrospinal (CSF) fluids. METHODS: Three hundred and fifty-one samples (99 AFs, 45 PFs, 75 SFs, and 132 CSFs) were analyzed in parallel with BC-BF, XN-BF, and optical microscopy (OM). The study also included the assessment of diagnostic agreement among BC-BF, XN-BF, and OM. RESULTS: The comparison of BC-BF vs XN-BF yielded slopes of Passing and Bablok regression always comprised between 0.9 and 1.0 except for EO-BF and HF-BF, whilst the intercepts ranged from -0.4 for MN-BF and 12.0 for PMN-BF. The bias was comprised between -103.3% and 67.1% for HF-BF and EO-BF, respectively. A significant bias was found for TC-BF, WBC-BF, HF-BF (negative bias) and for PMN-BF and EO-BF (positive bias). The agreement (Cohen's kappa) between XN-BF and BC-BF was always ≥0.7, ranging between 0.87 in CSFs and 0.94 in AFs, and that with OM was similar (ie, 0.85 and 0.96). CONCLUSION: The cytometric analysis of BF samples using BC-BF and XN-BF is clinically favorable when appropriately combined with OM. Quantitative and qualitative parameters displayed optimal agreement, whilst instrument-specific cut-offs should be defined and implemented for HF-BF and EO-BF. Further efforts should be made for achieving better harmonization in cytometric analysis of BF samples.

A Preliminary Proposal for Quality Control Assessment and Harmonization of Leukocytes Morphology-structural Parameters (cell Population Data Parameters).

BACKGROUND: The cell population data (CPD) measured by Sysmex XN-9000 can be used for screening many hematological and non-hematological disorders. Since little information is available on harmonization of CPD among different instrumentation and clinical laboratories, this study aimed at assessing the current degree of CPD harmonization between separate Sysmex XN modules allocated to the same laboratory. METHODS: A total number of 78291 data were used for verification of within-run imprecision, analyzers harmonization, reference ranges and assessment of blood sample stability of CPD parameters, including results of daily quality control testing and those generated in samples collected from blood donors and healthy volunteers. RESULTS: Within-run imprecision of CPD parameters ranged between 0.4 and 14.1%. Good agreement was found among five different XN-modules, especially when values were adjusted after calculation of instrument-specific alignment factors. The bias of all parameters remained always lower than the reference change values in samples stored for up to 8 hours, regardless of storage temperature. CONCLUSIONS: The imprecision of CPD parameters was acceptable, except for those reflecting the dispersion of cellular clusters. Due to the lack of reference control materials, we showed that the use of data generated on a large number of normal routine samples (i.e., a Moving Average population) may be a reliable approach for testing analyzers harmonization. Nevertheless, availability of both calibration and quality control materials for these parameters is highly advisable in the future. We finally showed that whole blood samples may be stable for up to 2-4 hours for most CPD parameters.

Short- and medium-term biological variation estimates of red blood cell and reticulocyte parameters in healthy subjects.

BACKGROUND: The integrated evaluation of traditional and innovative red blood cell (RBC) and reticulocyte parameters is a rapid, inexpensive and non-invasive diagnostic tools for differential diagnosis and follow-up of anemia and other pathological conditions needing bone marrow erythropoiesis assessment. Therefore, estimating the biological variation (BV) of these parameters is essential for evaluating the analytical performance of hematological analyzers, and for enabling accurate data interpretation and appropriate clinical management. This study aims to define short- and medium-term BV estimates and reference change value (RCV) of RBC and reticulocyte parameters. METHODS: Twenty-one healthy volunteers participated in the assessment of medium-term BV (blood sampling once/week, five consecutive weeks) and 22 volunteers in the assessment of short-term BV (blood sampling once/day, five consecutive days) using Sysmex XN. Outlier analysis was performed before CV-ANOVA, to determine BV estimates with confidence intervals (CI). RESULTS: Medium- and short-term within-subject BV were between 0.3% and 16.4% and 0.2%-10.4% (MCH and IRF), respectively, whereas medium and short-term between-subjects BV ranged between 0.9% and 66.6% (MCHC and Micro-R) and 1.4%-43.6% (MCHC and IRF), respectively. The RCVs were similar for all parameters in both arms of the study, except for hemoglobin, RDW-CV and MCV. CONCLUSIONS: This study allowed for estimating the BV of many RBC and reticulocyte parameters, some of which have not been currently explored. For RBC, hemoglobin, RDW-CV and MCV it seems advisable to use RCV calculated according to monitoring time and/or differentiated by sex. As regards analytical goals, we suggest using the most stringent targets found in the short-term arm of this study.

Innovative haematological parameters for early diagnosis of sepsis in adult patients admitted in intensive care unit.

AIMS: This study was aimed to investigate the role of erythrocyte, platelet and reticulocyte (RET) parameters, measured by new haematological analyser Sysmex XN and C reactive protein (CRP), for early diagnosis of sepsis during intensive care unit (ICU) stay. METHODS: The study population consisted of 62 ICU patients, 21 of whom developed sepsis during ICU stay and 41 who did not. The performance for early diagnosing of sepsis was calculated as area under the curve (AUC) of receiver operating characteristics curves analysis. RESULTS: Compared with CRP (AUC 0.81), immature platelet fraction (IPF) (AUC 0.82) showed comparable efficiency for identifying the onset of sepsis. The association with the risk of developing sepsis during ICU stay was also assessed. One day before the onset of sepsis, a decreased of RET% was significantly associated with the risk of developing sepsis (OR=0.35, 95% CI 0.14 to 0.87), whereas an increased of IPF absolute value (IPF#) was significantly associated with the risk of developing sepsis (OR=1.13, 95% CI 1.03 to 1.24) 2 days before the onset of sepsis. The value of CRP was not predictive of sepsis at either time points. CONCLUSIONS: IPF# and RET% may provide valuable clinical information for predicting the risk of developing sepsis, thus allowing early management of patients before the onset of clinically evident systemic infections.

Preliminary evaluation of UF-5000 Body Fluid Mode for automated cerebrospinal fluid cell counting.

BACKGROUND: Cellular analysis of cerebrospinal fluid (CSF) provides important diagnostic information in various medical conditions. The aim of this study was to evaluate the application of Sysmex UF-5000 body fluid mode in cytometric analysis of CSF compared to Light Microscopic (LM). METHODS: Eighty-one consecutive CSF samples were analyzed by UF-5000 body fluid mode and by LM. The study also included the evaluation of: limit of Blank (LoB), limit of Detection (LoD), limit of Quantitation (LoQ), carryover and linearity. RESULTS: For total nucleated cells (TNC-UF) and white blood cells (WBC-UF) LoB, LoD and LoQ were 1×106cells/L, 1.8×106cells/L and 1.9×106cells/L respectively. For red blood cells (RBC) LoB was 2×106cells/L, LoD was 3.5×106cells/L and LoQ was 14×106cells/L respectively. Linearity was excellent, carryover was negligible. The agreement between UF-5000 body fluid mode parameters and manual cell counts was good in all CSF samples with bias ranged between -0.5 and 25.1×106cells/L. The ROC curve analysis showed an area under curve of 0.99 for both TNC-UF and WBC-UF parameters. CONCLUSIONS: The UF-5000 body fluid mode offers rapid and accurate counts in clinically relevant concentration ranges, replacing the LM for most samples. However, in samples with abnormal cell counts or with abnormal scattergram the need for microscopic review remains.

Short- and medium-term biological variation estimates of leukocytes extended to differential count and morphology-structural parameters (cell population data) in blood samples obtained from healthy people.

BACKGROUND: Recent studies showed that some cell population data (CPD) parameters of neutrophils may be useful for diagnosing myelodysplastic syndromes and sepsis, for the differential diagnosis of acute promyelocytic leukemia, and some CPD parameters of lymphocytes may be a valuable tool for preliminary screening of B cell lymphoproliferative disease. Notwithstanding the knowledge, no information has been made available about their analytical quality specification. This study was aimed to define short- and medium-term biological variation (BV) estimates and reference change value (RCV) of leukocyte count, extended leukocyte differential and CPD. METHODS: The study population consisted of 43 healthy volunteers, who participated in the assessment of medium-term (21 volunteers; blood sampling once a week for 5 consecutive weeks) and short-term (22 volunteers; blood sampling once a day for 5 consecutive days) BV, using Sysmex XN. Outlier analysis was carried out before CV-ANOVA, to determine BV estimates and their confidence intervals. RESULTS: The medium-term and short-term within-subject BV (CVI) was comprised between 0.6-19.7% and 0.2-21.9%, whereas the medium-term and short-term between-subjects BV (CVG) was comprised between 1.2-61.5% and 1.1-58.5%. The RCVs were found to be similar for all the parameters, in both arms of the study, except for some CPD parameters. CONCLUSION: This study allowed accurately estimating the BV of many leukocyte parameters, some of which have not been currently explored. The kinetics of some leukocyte turnover suggests the use of short-term BV data for calculating analytical goals and RCV.

Biological variation of platelet parameters determined by the Sysmex XN hematology analyzer.

BACKGROUND: This study was aimed to define the short- and medium-term biological variation (BV) estimates, the index of individuality and the reference change value (RCV) of platelet count, platelet distribution width, mean platelet volume, platelet larger cell ratio, plateletcrit and immature platelet fraction. METHODS: The study population consisted of 43 health subjects, who participated to the assessment of medium-term (21 subjects; blood sampling once a week for 5 consecutive weeks) and short-term (22 subjects; blood sampling once a day for 5 consecutive days) BV study, using Sysmex XN-module. Eight subjects were also scheduled to participate to both phases. The data were subject to outlier analysis prior to CV-ANOVA, to determine the BV estimates with the relative confidence intervals. RESULTS: The medium-term and short-term within-subject BV (CVI) was comprised between 2.3 and 7.0% and 1.1-8.6%, whereas the medium-term and short-term between-subjects BV (CVG) was comprised between 7.1 and 20.7% and 6.8-48.6%. The index of individuality and index of heterogeneity were always respectively <0.6 and >0.63 for all the parameters, in both arms of the study. The RCVs were similar for all parameters, in both arms of the study. CONCLUSION: This study allowed to define the BV estimates of many platelet parameters, some of them unavailable in literature. The kinetics of platelet turnover suggests the use of short-term BV data for calculating analytical goals and RCV. The correct clinical interpretation of platelet parameters also necessitates that each laboratory estimates local RCV values.

Validation rules for blood smear revision after automated hematological testing using Mindray CAL-8000.

BACKGROUND: This article was aimed to test the use of validation rules for blood smear review after automated hematological testing using Mindray CAL-8000 (two hematological analyzers and one autoslider). METHODS: This study was based on 1013 peripheral blood samples (PB) referred for routine hematological testing. Results of testing on CAL-8000 were analyzed using both locally derived and International Consensus Group for Hematology (ICGH) validation rules, and then compared with data obtained by optical microscopy (OM). A workflow analysis was also completed. RESULTS: The overall agreement with locally derived and ICGH criteria was 91% and 85%, but a higher sensitivity was observed for locally derived criteria (0.97 vs 0.95). The percentage of false negative and false positive samples was 2.1% and 7.1% using ICGH criteria, and was 1.4% and 14% using locally defined rules. The throughput of CAL-8000 system was 208 samples/h, with a percentage of OM analysis comprised between 14% and 17%, and sensitivity of 0.97. As regards personnel activity, we estimated 0.8 full-time equivalent (FTE) of technical staff and 0.7 FTE of personnel for clinical validation of data and blood smear review. CONCLUSION: These results show that customization of validation rules is necessary for enhancing the quality of hematological testing and optimizing workflow.

Clinical significance of cell population data (CPD) on Sysmex XN-9000 in septic patients with our without liver impairment.

BACKGROUND: This study evaluated the clinical significance of cell population data (CPD) parameters obtained on Sysmex XN-9000 in septic patients admitted to intensive care unit (ICU) and stratified according to liver function. METHODS: The study population consisted in 84 patients, 44 of whom did not develop sepsis (NS), whereas the remaining 40 developed sepsis (SE) (n=24) or septic shock (SS) (n=16). Two hundred ostensibly healthy blood donors [healthy subjects (HS)], undergoing routine blood testing before a regular blood donation, were studied. RESULTS: Except for neutrophils and lymphocytes cell size (NE-FCS and LY-Z), all other CPD values were significantly different in ICU patients compared to HS. Neutrophils and monocytes fluorescence intensity (NE-SFL and MO-X) values were significantly higher in SS compared to sepsis and not develop sepsis patients. The value of many parameters was also different according to liver function. Overall, MO-X and neutrophils fluorescence intensity (NE-SFL) exhibited the best performance for diagnosing sepsis in all patients (AUC, 0.75 and 0.72), as well as in those with (AUC, 0.95 and 0.89) or without (AUC, 0.72 for both) liver impairment. These parameters were also significantly correlated with Sequential Organ Failure Assessment (SOFA) score. CONCLUSIONS: This study suggested that some novel CPD parameters (namely NE-SFL and MO-X) may provide useful information for diagnosis and management of sepsis.

Optimization of Cellular analysis of Synovial Fluids by optical microscopy and automated count using the Sysmex XN Body Fluid Mode.

BACKGROUND: This study was planned to assess the impact of pre-treating synovial fluid (SF) samples with hyaluronidase (HY), defining the best procedure for optical microscopy (OM) analysis and evaluating the performance of Sysmex XN-9000 Body Fluid module (XN-BF). METHODS: The cell count by OM was carried out both with and without HY pre-treatment, and using 3 different types of staining reagents. The evaluation of XN-BF included data comparison with OM (100 SFs), carryover, Limit of Blank (LoB), Limit of Detection (LoD), Limit of Quantitation (LoQ) and linearity. RESULTS: Unlike cell count in Burker's chamber and staining with Stromatol, pre-treatment with HY and staining with Methylene Blue and Turk's promoted cell clustering. The SF samples pre-treated with HY displayed excellent morphological quality, contrary to samples without HY pre-treatment. Excellent correlation was found between total cells counting with both OM and XN-BF. Satisfactory agreement was also observed between polymorphonuclear neutrophils compared to XN-BF parameter, whereas mononuclear cell count on XN-BF had suboptimal agreement with OM. The carryover was negligible. The LoB, LoD, LoQ and linearity were excellent. CONCLUSION: XN-BF displays excellent performance, which makes it a reliable and practical alternative to OM for SF samples analysis in clinical laboratories.

Assessment of blood sample stability for complete blood count using the Sysmex XN-9000 and Mindray BC-6800 analyzers.

BACKGROUND: Different hematological analyzers have different analytical performances that are often reflected in the criteria for sample stability of the complete blood count. This study aimed to assess the stability of several hematological parameters using the XN-9000 Sysmex and BC-6800 Mindray analyzers. METHODS: The impact of storage at room temperature and 4°C was evaluated after 2, 4, 6, 8, 24, 36 and 48h using ten normal and 40 abnormal blood samples. The variation from the baseline measurement was evaluated by the Steel-Dwass-Critchlow-Fligner test and by Bland-Altman plots, using quality specifications and critical difference as the total allowable variation. RESULTS: Red blood cells and reticulocyte parameters (i.e. hematocrit, mean corpuscular volume, mean corpuscular hemoglobin concentration, red blood cell distribution width, immature reticulocyte fractions, low-fluorescence reticulocytes, middle-fluorescence reticulocytes, high fluorescence mononuclear cells) showed less stability compared to leukocyte and platelet parameters (except for monocyte count and mean platelet volume). The bias for hematocrit, mean corpuscular volume, mean corpuscular hemoglobin concentration and red blood cell distribution width coefficient of variation was higher than the critical difference after 8h using both analyzers. CONCLUSION: Blood samples measured with both analyzers do not show analytically significant changes in up to 2h of storage at room temperature and 4°C. However, the maximum time for analysis can be extended for up to 8h when the bias is compared to the critical difference.