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Reticulocyte counting in thalassaemia.

The thalassaemias are a heterogeneous group of genetic haemoglobin disorders. The use of the Sysmex R- 1000 instrument in their study during the last 5 years has proved valuable. 1 Reticulocyte percentage and absolute counts were estimated in heterozygous beta-thalassaemia, in beta thalassaemia intermedia and in sickle beta thalassaemia and were compared with normal controls. Reticulocyte maturation subpopulations (high, middle and low fluorescence ratio) were assessed and compared with those of other haematological disorders. Red cell size and non-specific auramine-O binding were shown to be factors affecting mature red cell autofluorescence. 2 Nucleated red blood cells (NRBC) interfere with leucocyte counts in most haematology analysers. The upper particle count (UPP), provided by the R-1000 with modified fluorescence amplification voltage, appeared to produce a direct NRBC count in beta-thalassaemia intermedia when compared to NRBC counts assessed indirectly. 3 Erroneous platelet counts are reported by most haematology analysers in thalassaemia intermedia (especially in haemoglobin H disease) due to extensive microcyte-platelet interference and cause problems in diagnosis and management. Platelet counts provided by the R-1000 instrument in such patients were comparable to counts assessed by microscopy. Flow cytometric analysis by the Sysmex R-1000 instrument is useful in thalassaemia syndromes not only for providing precise reticulocyte counts and reticulocyte maturation data, but for direct NRBC counting and accurate platelet enumeration in cases of thalassaemia intermedia.

Anemia, Sickle Cell↗

Methodology-dependent variations in reticulocyte counts using a manual and two different flow cytometric procedures.

This paper describes a comparison between the microscopic brilliant cresyl blue and the flow cytometric thiazol orange (FACScan) and auramine O (Sysmex) methods for enumeration of reticulocytes. The mean intra-assay coefficients of variation for the microscopic, FACScan and Sysmex methods were established to be 33.9 and 5% respectively. A rather poor correlation was observed between the microscopic count and both flow cytometric methods (FACScan r = 0.61; Sysmex r = 0.57). However, the correlation between the flow cytometric methods was satisfactory (r = 0.79). Reference ranges for the reticulocyte count, corresponding with the 2.5th and 97.5th percentile, were determined for the microscopic (8-30/1000), FACScan (11-27/1000) and Sysmex (8-18/1000) methods. Sysmex R-3000 methodology definitely revealed the lowest and narrowest reference range. In conclusion, because of higher reproducibility, flow cytometric analysis of reticulocytes is an attractive alternative procedure for the microscopic enumeration method.

Flow Cytometry↗

Carbon monoxide production and reticulocyte count in normal women. Effect of contraceptive drugs and smoking.

Endogenous production of carbon monoxide (VCO), serum bilirubin (SB) and reticulocyte (retic) count have been followed during the menstrual cycle in 12 women (4 of them smokers) without contraceptive drugs and in 10 women (5 of them smokers) on contraceptive drugs. During the progesterone phase (Pph), VCO in the non-smokers was 18.2 +/- 2.1 mumol/mmol total body heme (TBH) and day (mean +/- 1 S.E.M.), compared with 10.1 +/- 1.0 during the estrogen phase (Eph). In the smokers, VCO was 10.4 +/- 1.9 during Pph and 12.5 +/- 0.7 during Eph. SB for the non-smokers was 5.2+/- 1.0 mug/ml during Pph and 3.9 +/- 0.5 during Eph, compared with 3.2 +/- 0.8 and 3.9 +/- 1.2, respectively, for smokers. The retic count for the non-smokers was 64.1 +/- 4.7 times 10(3)/mul during Pph and 17.7 +/- 5.1 during Eph. In the smokers the corresponding counts were 93.6 +/- 15.9 and 60.8 +/- 7.6. These results confirm earlier reports that VCO is increased during Pph and indicate that the rise could be due to a change in red cell catabolism, since it is concomitant with a significant increase in retic counts, which has not been found in earlier works on VCO, but which is known from reticulocyte studies. Furthermore, the results may indicate that persistent smoking affects the metabolism of CO and/or heme, since the smokers did not react with any change in VCO during Pph. In subjects using contraceptive drugs, VCO was significantly higher (14.9 +/- 0.6 on combined estrogen-gestagen drugs and 14.5 +/- 1.1 on continuous gestagens, respectively) than during Eph in non-smokers (10.1 +/- 1.0 mumol/mmol TBH and day), which might be taken as the baseline group. This indicates that the increased VCO in subjects on contraceptive drugs may be related to the activity of gestagens, which were concluded in all contraceptive drugs tested.

Adult↗