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Background and Objectives The haemoglobin level of prospective blood donors is usually performed on blood obtained by from the finger pulp by fingerstick with a lancet and filling a capillary tube with a sample. New noninvasive methods are now available for rapid, noninvasive predonation haemoglobin screening. Materials and Methods Prospective blood donors at our blood centre were tested, in two different trials, as follows: by the NBM 200 (OrSense) test (n = 445 donors) and by the Pronto-7 (Masimo) test (n = 463 donors). The haemoglobin values of each trial and the haemoglobin of finger pulp blood obtained by fingerstick with a lancet (HemoCue) were compared with the haemoglobin values obtained from a venous sample on a Cell Counter (Beckman Coulter). Results Comparison of Beckman Coulter Cell Counter and OrSense and results showed a bias of 0·29 g/dl, the standard deviation of the differences (SDD) of 0·98 and 95% limits of agreement from −1·64 to 2·21, using Bland and Altman statistical methodology. Comparison of Masimo and Beckman Coulter Cell Counter results showed a bias of −0·53 g/dl, SDD of 1·04 and 95% limits of agreement from −2·57 to 1·51. Cumulative analysis of all 908 donors, as tested by the usual fingerstick test showed a bias of 0·83 g/dl, SDD of 0·70 and 95% limits of agreement from −0·54 to 2·20 compared with the Coulter Cell Counter. Compared with the Coulter Counter, the specificity of the methods was 99·5% for fingerstick, 97% for OrSense and 83% for Massimo, and the sensitivity was 99, 98 and 93%, respectively. Conclusions Analysis of finger pulp blood by either direct sampling by fingerstick and Hemocue, or by noninvasive haemoglobin tests does not replicate the results of cell counter analysis of venous samples. Compared with fingerstick, noninvasive haemoglobin tests eliminate pain and reduce stress, but have a lower level of specificity and sensitivity.

Background and objectives Packed red blood cells (pRBCs) contain apoptotic white cells. We studied apoptotic cells in pRBCs after filtration and at various time-points during storage. Materials and methods To maintain the same subset of cells, seven pRBC units were pooled in a single bag and divided equally into seven aliquots. Two series of five experiments were performed: in the first we utilized the Biofil R01 Max filter, and in the second the Pall BPF4 filter was used. One aliquot was immediately leucodepleted while the others were stored at 4 degrees C and filtered on days 3, 7, 10, 14, 21 and 42 of storage. The postfiltration leucocyte counts and apoptotic evaluations were performed by using the Nageotte chamber and flow cytometry. Results The absolute number of residual leucocytes was always less than 0.5 x 106 in each experiment. Nageotte chamber counts showed a greater number of white blood cells than flow cytometry during the 42 days of storage. On day 0, the percentage of apoptotic cells in non-leucodepleted pRBCs was 1.1 +/- 0.4 and 1.2 +/- 0.4, while in filtered pRBCs it was high from day 0, at 53.5 +/- 16.3 and 52 +/- 18.5, respectively, with Biofil and Pall filters. On day 10 of storage, apoptotic cells reached a percentage of 42.5 +/- 15.8 and 41.6 +/- 18.6 in non-leucodepleted pRBCs, while in filtered units an average value of approximately 90% was found with both filters. Conclusions The percentage of apoptotic cells was higher in leucodepleted than in non-leucodepleted pRBCs. After filtration, the degree of apoptosis was already high on day 0, and reached a mean of approximately 90% by day 10. The difference in residual WBC counts between the Nageotte chamber and flow cytometry could be related to the presence of a high percentage of apoptotic cells in filtered blood components, and to the method used to distinguish viable from apoptotic cells.

We performed a flow-cytometric analysis of the expression of D antigen in D+W samples (previously termed Du). We also analysed a series of D-positive and D-negative (cde phenotype) samples to obtain positive and negative controls, respectively. The evaluation was carried out by immunofluorescence and the intensity of positivity was expressed as mean channel value (MCV) of fluorescence. Results demonstrated that D+W samples have lower expression (less than 1 log) than D-positive cases (p > 0.001, Student t test), while cde samples show the same MCVs as negative controls. Moreover, it was also possible to set a grading of D antigen expression and to analyze cases difficult to assess by agglutination only.

Packed red blood cells (pRBCs) contain apoptotic white cells. We studied apoptotic cells in pRBCs after filtration and at various time-points during storage.To maintain the same subset of cells, seven pRBC units were pooled in a single bag and divided equally into seven aliquots. Two series of five experiments were performed: in the first we utilized the Biofil R01 Max filter, and in the second the Pall BPF4 filter was used. One aliquot was immediately leucodepleted while the others were stored at 4 degrees C and filtered on days 3, 7, 10, 14, 21 and 42 of storage. The postfiltration leucocyte counts and apoptotic evaluations were performed by using the Nageotte chamber and flow cytometry.The absolute number of residual leucocytes was always less than 0.5 x 106 in each experiment. Nageotte chamber counts showed a greater number of white blood cells than flow cytometry during the 42 days of storage. On day 0, the percentage of apoptotic cells in non-leucodepleted pRBCs was 1.1 +/- 0.4 and 1.2 +/- 0.4, while in filtered pRBCs it was high from day 0, at 53.5 +/- 16.3 and 52 +/- 18.5, respectively, with Biofil and Pall filters. On day 10 of storage, apoptotic cells reached a percentage of 42.5 +/- 15.8 and 41.6 +/- 18.6 in non-leucodepleted pRBCs, while in filtered units an average value of approximately 90% was found with both filters.The percentage of apoptotic cells was higher in leucodepleted than in non-leucodepleted pRBCs. After filtration, the degree of apoptosis was already high on day 0, and reached a mean of approximately 90% by day 10. The difference in residual WBC counts between the Nageotte chamber and flow cytometry could be related to the presence of a high percentage of apoptotic cells in filtered blood components, and to the method used to distinguish viable from apoptotic cells.

Background and objectives: There is increasing interest in the development of rapid and reliable techniques for human platelet alloantigen (HPA) typing. This study investigates the reliability of flow cytometry for large-scale immunophenotyping of platelet alloantigens. Materials and methods: We used flow cytometry and polymerase chain reaction-site-specific primer (PCR-SSP) for the characterization of the human platelet antigen 1 (HPA-1) mosaic in blood donors. Results: By using specific alloantisera and immunofluorescence labelling 9 (2.6%) out of 351 samples were HPA-1a-negative. To confirm this antigenic phenotype, all of the latter samples were submitted to PCR-SSP analysis, showing an HPA1-b/b genomic pattern. In HPA-1a-positive donors, flow cytometry was unable to distinguish HPA-1a/b heterozygous from HPA-1a/a homozygous subjects who were clearly identified by genotyping. Conclusions: Flow cytometry is a valuable tool for large-scale screening to identify HPA-1a-negative persons, whereas genotyping is the assay of choice for zygosity testing, antenatal diagnosis, and for thrombocytopenic alloimmunized patients.

A functional and phenotypic analysis of the circulating natural killer (NK) cell population was carried out in 9 patients with idiopathic autoimmune hemolytic anemia (IAHA). The NK-cell activity, assessed by a sensitive method based on the inhibition of target clone growth in plasma semisolid medium, was markedly decreased in all patients as compared with normal controls and was not restored by stimulation of the cells with recombinant alpha interferon (alpha-IFN). Analysis of peripheral blood phenotypic markers showed that cells bearing Leu7 and CD16 antigens numbered in the normal range. These findings suggest that IAHA patients exhibit a functional impairment of the NK compartment.

Alterations in the autologous mixed lymphocyte reaction (autologous MLR) have been reported in many autoimmune diseases and in diseases with a derangement of T regulatory function. We have studied autologous MLR in 10 patients with idiopathic autoimmune hemolytic anemia (IAHA). All patients had decreased autologous MLR which averaged 4,106 +/- 1,332 cpm (SEM) compared to 12,153 +/- 4,166 cpm for simultaneously studied controls. A marked decrease in phytohemagglutinin response and an imbalance of T-cell subsets were also observed. These findings suggest a possible abnormality of T immunoregulatory function in IAHA patients.

We tested serum samples from 11 HIV-seropositive and 32 HIV-seronegative Italian patients with hemophilia A or B for the presence of lymphocytotoxic antibodies (LCTAs). No patients had the acquired immune deficiency syndrome and all had received over many years heat-untreated commercial clotting factors. LCTAs directed mainly to B cells, were significantly present in 5/6 HIV-seropositive patients with hemophilia A and in 5/5 HIV-seropositive patients with hemophilia B. The presence of LCTAs in HIV-seronegative hemophiliacs was significantly correlated with both a decrease of T cells and an increase in serum levels of IgG and IgM.