Performance evaluation of platelet counting by novel fluorescent dye staining in the XN-series automated hematology analyzers.

BACKGROUND: Conventional automated hematology analyzers have limitations in platelet measurements such as poor accuracy and precision in the low count range and interference by nonplatelet particles. In order to improve it, the newly developed XN-Series automated hematology analyzers (Sysmex Corporation, Kobe, Japan) have been installed with a new dedicated channel for platelet analysis (PLT-F), which is based on a fluorescence flow cytometry method with uses of a novel fluorescent dye specifically staining platelets. We evaluated the basic performance of this new PLT-F channel. METHODS: Basic performance of the PLT-F channel in within-run reproducibility and assay linearity was studied using standard methods. Correlation was studied between PLT-F and a conventional automated hematology analyzer (XE-2100) and immunoplatelet analysis using anti-CD61 monoclonal antibody (Cell-Dyn Sapphire; Abbott Laboratories). The assay interference by nonplatelet particles such as fragmented red and white blood cells was evaluated by using clinical samples, respectively, from burn injury and acute leukemia. RESULTS: Basic performance of the PLT-F platelet counting was satisfactory in within-run reproducibility, linearity and correlation with the conventional analyzer. The correlation was satisfactory also with the immunoplatelet analysis, even for samples from a patient with burn injury, and those with white blood cell fragments displayed, platelet abnormal flag and low platelet counts (<50 × 10(9)/l). CONCLUSION: The platelet counting performance of the PLT-F channel of the XN Series had improved accuracy and precision in the low range and in abnormal samples, avoiding the interference by nonplatelet particles.

Morphological definition of CD71 positive reticulocytes by various staining techniques and electron microscopy compared to reticulocytes detected by an automated hematology analyzer.

BACKGROUND: The enumeration of peripheral blood reticulocytes plays an important part in clinical hematology. Although reticulocyte enumeration is currently performed with visible dyes such as New Methylene Blue (NMB), fluorescent dyes, or anti-CD71 (transferrin receptor) antibody, it has not been demonstrated whether the reticulocytes detected in each method are the same or not. METHODS: We prepared the reticulocyte rich fraction with density gradient centrifugation, stained with both anti-CD71 and Sysmex's fluorescent stain RET SEARCH (II), and detected the cells by both confocal laser scanning microscopy and flow cytometry. We also stained the reticulocyte rich fraction and the CD71+ reticulocytes with NMB and compared them by microscopy. We also observed the CD71+ reticulocytes by electron microscopy. RESULTS: Almost all CD71+ reticulocytes were intensely stained with both NMB and RET SEARCH (II). These cells were therefore classified as highly immature reticulocytes. During the stages of reticulocyte maturation, the expression of CD71 antigen decreased prior to the reduction of the reticular structures. The electron microscopic observation showed that CD71+ reticulocytes had some typical morphological characteristics found in highly immature reticulocytes. CONCLUSIONS: The CD71+ reticulocytes consisted of highly immature reticulocytes and were not equal to the reticulocytes defined with NMB or RET SEARCH (II).

Effects of biological variations on platelet count in healthy subjects in China.

The effects of biological variations on platelet counts were investigated in 694 healthy subjects aged 18 to 60 years living in three cities including Chengdu (Sichuan Province), Suzhou (Jiangsu Province) and Harbin (Heilongjang Province) in China. Platelet counts in healthy subjects were significantly lower in Chengdu (52-202 x 10(9)/L) and Suzhou (60-259 x 10(9)/L) than in Harbin (154-348 x 10(9)/L) (p <0.0001), but the mean platelet volume (MPV) determined concurrently was negatively correlated with platelet count, the MPV values were significantly higher in Chengdu (11.8-15.6 fl) and Suzhou (10.9-15.8 fl) than in Harbin (9.5 approximately 12.9 fl) (p < 0.0001). Platelet counts were significantly higher in summer (73-289 x 10(9)/L) than in winter (52-202 x 10(9)/L) (p <0.0001), but the MPV values were lower in summer (11.2-14.7 fl) than in winter (11.8-15.6 fl) (p <0.05) in Chengdu. Platelet associated immunoglobulin (PA-IgG) in Chengdu was revealed to be significantly higher in the low platelet count group (<150 x 10(9)/L, 13.5 +/- 7.1 ng/10(7) PLT) than in the normal platelet count group (> or =150 x 10(9)/L, 8.3 +/- 2.7 ng/10(7) PLT) (p <0.0001). Similar results were observed in Suzhou for the reticulated platelet ratio, which was significantly higher in the low platelet count group (19.5 +/- 7.1%) than in the normal platelet count group (11.6 +/- 2.7%) (p <0.01). The bleeding time in Chengdu showed a significantly longer time in the low platelet count group (8.6 +/- 2.3 min) than in the normal platelet count group (6.0 +/- 1.2 min)(p <0.01). With regard to the effects of lipids on platelet counts, the HDL values were significantly higher in the normal platelet count group (1.60 +/- 0.76 mmol/L) than the low platelet count group (1.23 +/- 0.31 mmol/L) (p <0.01); but no significant differences in cholesterol and triglycerides values between the normal and low platelet count groups (p >0.05) were recorded. These findings suggest that the platelet counts could be greatly influenced in healthy subjects by biological variations such as geographical, seasonal, and lipid variations.

Studies on reference intervals for platelet counts in three cities in China and one in Japan.

Hematological parameters including platelet counts, etc. were determined in 1,140 healthy subjects living in four cities: Suzhou (Jiangsu Province), Chengdu (Sichuan Province) and Harbin (Heilongjang Province) in China, and Kobe in Japan. Then, the reference intervals for platelet counts were calculated and compared. The reference interval for platelet count of subjects aged between 18 and 60 years was 60-259 x 10(9)/L in Suzhou and 52-202 x 10(9)L in Chengdu, and subjects with platelet counts of 100 x 10(9)/L or less accounted for about 30% of the subjects examined in these cities. The reference intervals in Harbin and Kobe were within the range of 150-350 x 10(9)/L, and no subject having a platelets count of 100 x 10(9)/L or less was detected. Mean platelet volume (MPV) determined concurrently was negatively correlated with platelet count, and the reference intervals for MPV in Chengdu and Suzhou were higher than those in Harbin and Kobe.