Differential leucocyte subpopulation analysis of leucodepleted red cell products.

Donor leucocytes are responsible for many adverse transfusion effects. Clinical reactions may be attributed to specific leucocyte subsets. In this study leucocyte subpopulations were identified and quantified pre- and post-leucodepletion by integral filtration using novel Optipac(R) configurations incorporating either WBF-1 (Pall Medsep) or RS2000 (Asahi) whole blood filters. Leucocytes were analysed by flow cytometry using direct, four-colour, membrane immunofluorescence with monoclonal antibodies specific for CD 3, 14, 16, 19 and 45. Filtration reduced the leucocyte load by 3-4 log10, consistently giving products with < 2 cells microL-1. Subset distributions were also affected with the proportion of neutrophils and monocytes increased and the lymphocyte/monocyte ratio inverted. These effects were independent of the preprocessing hold conditions, filter used and buffy coat (BC) removal. All filtered red cell products contained 75-80% neutrophils, 16-20% monocytes and 2-7% lymphocytes. Results presented here demonstrate that whole blood filtration, and BC removal, significantly reduce the content and substantially alter the subpopulation distribution of the donor leucocytes remaining in leucodepleted red cell products.

Prevention of transfusion-transmitted cytomegalovirus infection.

Cytomegalovirus (CMV) is a double-stranded DNA virus which can be transmitted by blood transfusion. Its seroprevalence in adults ranges from 40% to 100% depending on geographical and socioeconomic conditions. Seropositive individuals have latent CMV infection with viral DNA present in peripheral blood leucocytes. CMV can be associated with considerable morbidity and mortality in susceptible individuals, e.g. CMV-seronegative bone marrow allograft patients. Evidence, from a number of reports, suggests that provision of leucodepleted blood components may be as effective as the use of components from CMV-seronegative donors in preventing CMV infection and disease. This is relevant in the UK because Blood Transfusion Services are implementing universal leucodepletion of cellular blood components to minimize the theoretical risk of transmission of new variant Creutzfeldt-Jakob disease. This review examines data on the biology of CMV, discusses options for testing and summarizes the impact of CMV-seronegative and leucodepleted blood components on transfusion-transmitted CMV.

Evaluation of plasma and red cells obtained after leucocyte depletion of whole blood.

We evaluated whole blood integral filtration to produce leucocyte-depleted red cells and plasma by using the WBF1 whole blood filter (Pall Medsep). Whole blood units were filtered after either warm (2-4 h at room temperature) or cold (12-24 h at 4 degrees C) holds. Filtered and control units were processed using either a bottom-and-top or top-top method. Red cells were tested weekly for 6 weeks, and plasma 3 monthly for 12 months. All filtered red-cell packs contained < 5 x 10(6) leucocytes/unit with 71 of 72 containing < 1 x 10(6) leucocytes/unit. No clinically significant differences in red-cell storage parameters were seen, although haemolysis was less and pO2/pCO2 values were better maintained in filtered units. Plasma units contained < 2.5 x 10(3) leucocytes/unit with no significant loss of factor VIII except in the warm hold units processed by the top-top method. There was no evidence of complement or coagulation activation with significant removal of preformed C3a in cold hold units. Plasma storage parameters were maintained at control levels for 12 months.

Human cytomegalovirus infection and systemic lupus erythematosus.

OBJECTIVE: Viruses are considered possible aetiologic agents of autoimmune disease. Evidence suggests that human cytomegalovirus (HCMV) may be a pathogenetic factor in systemic lupus erythematosus (SLE). We undertook a seroepidemiological study to determine whether HCMV infection is increased in patients with SLE. METHODS: Sero-epidemiologic data, indicative of virus prevalence, were obtained by enzyme immunoassay. RESULTS: Eighty-eight of 97 serum samples (90.7%) taken from adult patients with SLE were seropositive for HCMV. By contrast, HCMV was detected in only 32 of 50 (64.0%) adult patients with rheumatoid arthritis (RA) and 42 of 97 (43.3%) normal controls. The odds ratio for HCMV prevalence in SLE/normal controls was 14.53 (95% CI is 6.39 to 33.04). For comparison, data for herpes simplex virus-I (HSV-I) seropositivity were obtained from the same three groups. Seventy-eight patients with SLE (80.4%), 40 patients with RA (80.0%) and 57 normal controls (58.8%) were seropositive for this closely-related herpesvirus. CONCLUSION: The data shows a specific and highly significant association between infection with HCMV and a clinical diagnosis of SLE.

Evaluation of a new, integral, whole blood filter (RS2000) system for prestorage leucodepletion of SAG-M red cells.

Residual donor leucocytes are responsible for many adverse transfusion reactions. Prestorage leucodepletion may ameliorate these effects and enhance product quality. We studied a bottom and top (BAT) system incorporating an integral filter for whole blood leucodepletion. Our evaluation assessed leucodepletion efficiency as well as in vitro SAG-M red cell quality and storage characteristics. Sixty-six units of blood were collected; test units into the Optipac-pLuS system and controls into the standard triple pack configuration. Test units were held for 4-6 h at room temperature (rt) or 12-18 h at 4 degrees C. The mean leucocyte counts for the SAG-M red cells in the quality and storage trial were 0.6 x 10(6) (rt hold), 0.05 x 10(6) (4 degrees C hold) and 2500 x 10(6) (controls). We observed no significant differences between the groups for Na+, ATP, 2,3-DPG, glucose, lactate and pH during the 49 d storage. The control group, however, showed a greater increase in haemolysis and K+ with time. Autologous in vivo 24 h red cell recovery, after 42 d storage, was > 75%. Adjustment of processing parameters in subsequent studies gave leucodepleted SAG-M red cells with minimal cell loss (9.19%) plus acceptable haemoglobin content (46-76 g/U) and haematocrit (54-62%). This system achieved > 3.5 log leucodepletion with all but one unit containing < 1 x 10(6) leucocytes. The product quality is good and the system suitable for routine use in blood centres.

Morphology and distribution of gp52 on extracellular human cytomegalovirus (HCMV) supports biochemical evidence that it represents the HCMV glycoprotein B.

Glycoprotein gp52 exists within the mature human cytomegalovirus (HCMV) envelope in heterodimeric, disulphide-linked complexes with glycoproteins gp95 and gp130. Biochemical studies involving immunoprecipitations and Western blots have demonstrated that gp52 is the glycoprotein B (gB) homologue of HCMV but that gp95 and gp130 are probably separate gene products. The distribution of this putative gB on extracellular HCMV particles was revealed by high resolution electron microscopy of preparations labelled with a monoclonal antibody, F5, directly coupled to colloidal gold. F5-gold probes, specific for HCMV gp52, bind to the distal end of 12 nm long, slender spikes projecting from virion and dense body envelopes. Labelled spikes were most often present in closely packed, homogeneous clusters and were frequently present on envelope protrusions. The degree of labelling on individual HCMV particles was highly variable. Both the morphology and distribution of HCMV gp52 show strong similarity with that previously reported for the gB of herpes simplex virus. Other morphologically distinct spikes occur in the HCMV envelope but these were not recognized by F5-gold probes.