What do hemolyzed whole-blood specimens look like? Analysis with a CellaVision DM96 automated image analysis system.

We planned an original study to investigate the morphological changes caused by spurious hemolysis of whole-blood samples, analyzed using an automated image analysis system. Seven whole-blood specimens anticoagulated with EDTA were pooled and divided in two aliquots. The former was left untreated, whereas the latter was subjected to mechanical hemolysis by forced aspiration with an insulin syringe. The complete blood cell count was performed on a Sysmex XE-2100, and the aliquots were then processed with CellaVision DM96. In spuriously hemolyzed samples, the main findings included a rarefaction of erythrocytes, the presence of a remarkable number of cellular debris, a greater degree of microcytosis and anisocytosis, the appearance of band neutrophils, a shift of values between lymphocytes and monocytes, and an increase in smudge cells, artifacts, and large platelets. The results of this study demonstrate for the first time that blood cell morphology may be consistently biased in spuriously hemolyzed whole blood and that the use of an automated image analysis system such as the CellaVision DM96 may be a suitable approach to identify spurious hemolysis in whole-blood specimens.

Evaluation of mean platelet volume with four hematological analyzers: harmonization is still an unresolved issue.

The clinical applications of mean platelet volume (MPV) have recently broadened far beyond the differential diagnosis of platelet (PLT) disorders to embrace diagnosis and prognostication of a variety of thrombotic conditions. As the potential usefulness of this simple and inexpensive parameter may be challenged by instrument heterogeneity, we investigated the degree of analytical quality and interinstrument comparability. One hundred consecutive inpatient samples were simultaneously assessed on Abbott Sapphire, Mindray BC6800, Siemens Advia 2120, and Sysmex XE5000. The within-run imprecision of the four hematological analyzers was also assessed according to the Clinical and Laboratory Standards Institute document EP5-A2. The imprecision of PLT count ranged between 1.4 and 4.3%, and hence was always within the desirable quality specifications. The within-run imprecision of MPV ranged between 1.1 and 3.8%, and hence was also within the desirable quality specifications. The optical and impedance measurements displayed excellent correlations. Overall, the PLT count exhibited a modest instrumental variation, with bias always within the desirable quality specifications. A large bias was instead recorded for MPV, with between-instrument variations exceeding the desirable quality specifications in five out of six interinstrumental comparisons. No significant correlation was also observed between PLT count and MPV with any of the instruments tested. These results attest that although there is an optimal degree of analytical quality and comparability for PLT counting among different hemocytometers, the harmonization of MPV is poor, thus making the adoption of universal cutoffs virtually impossible.

Lack of harmonization of red blood cell distribution width (RDW). Evaluation of four hematological analyzers.

OBJECTIVES: To assess analytical imprecision and comparability of red blood cell distribution width (RDW) on Abbott Sapphire, Mindray BC6800, Siemens Advia 2120 and Sysmex XE-5000. DESIGN AND METHODS: Within-run imprecision was assessed on three pools and comparability using 132 inpatient samples. RESULTS: The imprecision of RDW was comprised between 0.3 and 1.2%, but the values exhibited broad variation among different analyzers, with bias exceeding the desirable quality specifications. CONCLUSIONS: Harmonization of RDW is still an unmet need.

Evaluation of the fully automated hematological analyzer Sysmex XE-5000 for flow cytometric analysis of peritoneal fluid.

Although microscopy still represents the gold standard for cytometric analysis of peritoneal fluids, automated flow cytometry may improve throughput and accuracy. We evaluated the performance of total nucleated cell (TNC), white blood cell (WBC), polymorphonuclear cell (PMN), and mononuclear cell (MONO) counts of Sysmex XE-5000 on peritoneal fluids. The imprecision was excellent, being always lower than 11%, whereas linearity studies yielded correlation coefficients of 1.00 for all parameters. The carryover was always lower than 0.2%. The comparison between XE-5000 and microscopic analysis of 117 ascitic fluids yielded correlation coefficients always greater than 0.96, with mean biases <11/µL. The diagnostic accuracy versus manual microscopy was greater than that of XE-2100, especially at thresholds for septic ascites (100 versus 98% for ≥500 WBC/µL; 98 versus 93% for ≥250 PMN/µL). The correlation with manual microscopy for macrophages and mesothelial cell count was also higher for XE-5000 than for XE-2100 (0.63 versus 0.55). The results of this evaluation show optimal performance of XE-5000 for routine analysis of ascitic fluids, which are combined with the advantages of automated analysis such as high throughout, shortened turnaround time, no need of sample preparation and trained staff, reduced sample volume, and less likelihood of transcriptional errors.

Evaluation of white blood cell count in peritoneal fluid with five different hemocytometers.

OBJECTIVES: Evaluation of automated flow cytometric analysis of white blood cell (WBC) count in peritoneal fluids. METHODS: One hundred peritoneal fluids were analyzed with manual microscopy, Sysmex XE-2100 and XE-5000, Siemens Advia 2120, Mindray BC-6800, Abbott Sapphire. RESULTS: High correlations (0.978 to 0.999) and modes biases (-132 to 80 WBC/mm(3)) were found. Agreement at septic peritonitis cutoff ranged between 96% and 99%. CONCLUSIONS: These hemocytometers display acceptable performance for WBC screening in peritoneal fluids.

Studies on in vitro hemolysis and utility of corrective formulas for reporting results on hemolyzed specimens.

INTRODUCTION: Spuriously hemolyzed specimens are the most common preanalytical problems in clinical laboratories. Corrective formulas have been proposed to allow the laboratory to release test results on these specimens. This study aimed to assess the influence of spurious hemolysis and reliability of corrective formulas. MATERIALS AND METHODS: Blood collected into lithium heparin vacuum tubes was divided in aliquots and subjected to mechanical injury by aspiration with an insulin syringe equipped with a thin needle (30 gauge). Each aliquot (numbered from "#0" to "#5") was subjected to a growing number of passages through the needle, from 0 to 5 times. After hematological testing, plasma was separated by centrifugation and assayed for lactate dehydrogenase (LD), aspartate aminotransferase (AST), potassium and hemolysis index (HI). RESULTS: Cell-free hemoglobin concentration gradually increased from aliquot #0 (HI: 0) to #5 (HI: 76 +/- 22, cell-free hemoglobin approximately 37.0 g/L). A highly significant inverse correlation was observed between HI and red blood cell count (RBC), hematocrit, mean corpuscular volume (MCV), LD, AST, potassium, whereas the correlation was negative with mean corpuscular hemoglobin (MCH). No correlation was found with hemoglobin, platelet count and glucose. A trend towards decrease was also observed for white blood cells count. The ANCOVA comparison of analyte-specific regression lines from the five subjects studied revealed significant differences for all parameters except potassium. In all circumstances the sy,x of these equations however exceeded the allowable clinical bias. CONCLUSIONS: Mechanical injury of blood, as it might arise from preanalytical problems, occurs dishomogeneously, so that corrective formulas are unreliable and likely misleading.