Establishing local reference intervals for full blood count and white blood cell differential counts in Cape Town, South Africa.

BACKGROUND: Accurate laboratory reference intervals (RIs) are essential to differentiate between health and disease. There are variations in haematological indices within populations relating to gender, age, ethnicity and environment. Iron deficiency is common, has a wide range of clinical morbidities and affects red cell indices. Locally derived RIs for full blood count (FBC) parameters are needed for the Western Cape region of South Africa, after the exclusion of iron deficiency. In addition, information regarding the prevalence of iron deficiency in first-time blood donors would inform blood transfusion services regarding policies to screen for and treat iron deficiency. OBJECTIVES: To establish locally derived RIs for FBC and white blood cell (WBC) differential count parameters in healthy adults in the Cape Town area, by including first-time blood donors and excluding those with iron deficiency and thalassaemic indices. These new locally established RIs could update those in use by the local National Health Laboratory Service. A secondary objective was to establish the prevalence of iron deficiency in first-time blood donors. This would inform blood donation policies regarding screening and appropriate iron supplementation in high-risk groups prior to blood donation. METHODS: This was a prospective, descriptive study with direct convenience sampling. Participants were prospective voluntary blood donors aged between 18 and 60 years, presenting for first-time blood donation. Ethnicity was self-identified. Participants who tested positive for HIV or hepatitis B and/or C viruses were excluded. Prospective participants with iron deficiency, defined by serum ferritin levels below the RI, and those with red cell indices suggestive of an underlying thalassaemia trait were excluded. FBC samples were analysed using a Sysmex XN-1000 cell counter. Statistical non-parametric methods were used to calculate the RIs, according to international guidelines. RESULTS: Of the 774 participants screened, 82 (11%) had iron deficiency and were excluded. Six hundred and sixty-two patients were included for analysis, 409 (62%) female and 253 (38%) male. The majority of the participants, 348 (53%), were between 20 and 29 years of age, with a mean age of 29 years for females and 28 years for males. Participants comprised a mix of the various ethnic groups residing in Western Cape Province. The mean haemoglobin concentration for females was lower than that for males (p<0.0001). There were significant gender differences for total WBC count, absolute neutrophil count and platelet count, with females having higher counts than males. CONCLUSIONS: Locally established, population-specific RIs are essential for the accurate interpretation of haematological indices. This study established locally derived gender-specific RIs for the Cape Town region, after exclusion of iron deficiency. These new RIs have implications for the accurate diagnoses of cytopenias, cytoses and other blood count abnormalities. Iron deficiency is common in first-time blood donors, and screening for iron deficiency using point-of-care testing should be considered.

The "vanishing counting bead" phenomenon: effect on absolute CD34+ cell counting in phosphate-buffered saline-diluted leukapheresis samples.

BACKGROUND: Using a single-platform protocol to count absolute CD34+ hematopoietic precursor cell (HPC) levels with different reference microbeads, we recorded occasionally artifactually high CD34+ HPC counts in some leukapheresis bags, whereas dual-platform calculations were always consistent. Abnormal countings were observed only when phosphate-buffered saline (PBS)-diluted leukapheresis samples were vortexed before analysis. A large series of blood samples analyzed similarly for CD34+ and CD4+ absolute counts did not show any sample or vortexing effect. With the volumetric absolute counting cytometer Partec-PAS, lower counts were also observed when different reference beads were vortexed before the instrument checking procedures. The counting abnormality was caused by a drop in microbead concentration (the "vanishing bead phenomenon"). This phenomenon reduced the total and relative bead event number in experimental and routine samples and in calibration procedures. This altered the bead denominator used to calculate absolute CD34+ HPC levels and it also reduced the concentration of standard calibration beads. METHODS: Using the Partec-PAS to measure volumetrically the actual bead concentration, we studied the vanishing bead phenomenon. Different types of counting and reference microbeads were resuspended in media with or without proteins or cells. Replicates were submitted either to gentle manual mixing or to vortexing before counting. RESULTS: Vortex agitation almost invariably induced the vanishing bead phenomenon when beads were resuspended in saline media or when an insufficient protein concentration was present, such as in diluted leukapheresis samples. Different bead types showed various degrees of sensitivity to vortexing. The bead disappearance was not caused by bubble formation or disruption. The addition of small amounts of protein completely prevented the vanishing bead phenomenon. The causative effect of the electrostatic charging of tube induced by vortexing is hypothesized. CONCLUSIONS: Sample suspensions containing counting beads for single-platform analysis must be resuspended in media with protein supplements to prevent the vanishing bead phenomenon and to ensure accurate counting.

Monitoring universal leucodepletion performance: the current issues within components production and testing.

The UK NBS completed the introduction of universal leucodepletion in November 1999. This involved the implementation of new methods of production, testing and monitoring, the development of which is still ongoing. Whilst important lessons have been learnt, much remains to be achieved in respect of quality improvement through standardisation of procedures, processes and product modifications to improve safety/efficacy of therapeutic blood components. Since the introduction of universal leucodepletion, many changes in component production and testing have occurred. The primary goal of this report is to review the level of standardisation and harmonisation currently achieved for low leucocyte counting within the NBS and discuss remedial actions undertaken to improve the standard of blood component production and quality monitoring and to highlight some of the related issues which are currently being worked on. The following issues are discussed.

New reference values for routine blood samples and human neutrophilic lipocalin during third-trimester pregnancy.

Reference values are usually based on blood samples from healthy men or non-pregnant women. Blood samples from pregnant women may be compared with these reference values. Correct references for pregnancy can be extremely important for clinical decisions such as ablatio placentae, appendicitis, premature rupture of membranes and preeclampsia. Previous studies of normal variations during third-trimester pregnancy are incomplete. Blood samples during pregnancy weeks 33, 36 and 39 as well as 1-3 h postpartum were collected from pregnant women with dietary iron supplement and at least one previous pregancy without a history of hypertension or preeclampsia. When the sampled values were compared with the present reference values from men and non-pregnant women, the following differences were found during normal pregnancy: Haemoglobin and ferritin were reduced, CRP was slightly elevated, WBC (white blood cell count) and HNL (human neutrophilic lipocalin) were elevated during pregnancy and significantly increased postpartum. Albumin was reduced. ALT and AST were slightly elevated and GGT was unchanged during pregnancy. ALP, D-dimer and fibrinogen were elevated. Uric acid increased during the third trimester and thrombocyte count decreased. Separate reference values for pregnant women are essential for correct diagnostic decisions during third-trimester pregnancy. Elevated levels of D-dimer do not necessarily indicate ablatio placentae. A diagnosis of progressive preeclampsia cannot be based on increasing uric acid levels and reduced platelet count in a stable clinical condition. HNL signals activation of neutrophilic granulocytes and can thereby offer a helpful tool for diagnosing infection during pregnancy and postpartum.