Comparison of the sensitivity of NAT using pooled donor samples for HBV and that of a serologic HBsAg assay.

Background: Studies were conducted using samples from early and late-stage HBV-infected persons to determine the pool size at which PCR had better sensitivity than a sensitive HBsAg chemoluminescence immunoassay (CLIA-HBsAg).

Study Design And Methods: HBV seroconversion panels were tested for HBsAg by CLIA and for HBV DNA by nested PCR (95% hit rate: 100 copies/mL); PCR was carried out at various dilutions. HBV serologically positive samples that were detected from the simultaneous screening of 540,161 routine whole-blood donations using CLIA-HBsAg and agglutination assays were also characterized for additional markers of HBV infection.

Superoxide generation from human polymorphonuclear leukocytes by liposome-encapsulated hemoglobin.

We investigated the interactions between liposome-encapsulated hemoglobin (Neo Red Cells: NRC) and human polymorphonuclear leukocytes as assessed by superoxide generation. NRC triggered superoxide generation from neutrophils in a dose-dependent manner. Empty liposomes also induced superoxide production of neutrophils.

Effects of poly(ethyleneglycol)-modified hemoglobin vesicles on agonist-induced platelet aggregation and RANTES release in vitro.

We studied the effects of hemoglobin-vesicles modified with PEG (PEG-HbV), a type of liposome-encapsulated hemoglobin (LEH), on human platelet functions in vitro. The effect of a low concentration of PEG-HbV (Hb; 5.8 mg/dl) was assessed by examining an agonist-induced aggregation response, and that of relatively high concentrations of PEG-HbV (Hb; 0.

Inactivation of parvovirus B19 in coagulation factor concentrates by UVC radiation: assessment by an in vitro infectivity assay using CFU-E derived from peripheral blood CD34+ cells.

Background: Nonenveloped and thermostable viruses such as parvovirus B19 (B19) can be transmitted to patients who are receiving plasma-derived coagulation factor concentrates treated by the S/D method for inactivating enveloped viruses. Therefore, it is important to develop and validate new methods for the inactivation of nonenveloped viruses.

Study Design And Methods: Suspensions of B19 in coagulation factor concentrates (FVIII) were irradiated with UVC light.

CrkL is an adapter for Wiskott-Aldrich syndrome protein and Syk.

Wiskott-Aldrich syndrome (WAS) and X-linked thrombocytopenia are caused by mutations of the WAS protein (WASP) gene. WASP may be involved in the regulation of podosome, an actin-rich dynamic cell adhesion structure formed by various types of cells. The molecular links between WASP and podosomes or other cell adhesion structures are unknown.

Involvement of reactive oxygen species in hemoglobin oxidation and virus inactivation by 1,9-dimethylmethylene blue phototreatment.

The participation of reactive oxygen species (ROS) in virus inactivation by 1,9-dimethylmethylene blue (DMMB) phototreatment in stroma-free hemoglobin (SFH) was investigated with the use of scavengers, quenchers and enhancer. Virus (R17 bacteriophage) photoinactivation by either activated monomer or dimer DMMB was suppressed by sodium azide (singlet oxygen quencher) and promoted by the substitution of H2O for deuterium oxide (D2O), which is known to prolong the lifespan of singlet oxygen. There was no or little effect of mannitol (hydroxyl radical scavenger) and superoxide dismutase (superoxide scavenger) on the photoinactivation.

Wiskott-Aldrich syndrome protein and platelets.

Wiskott-Aldrich syndrome (WAS) and X-linked thrombocy topenia (XLT) are caused by mutations of the WAS protein (WASP) gene. The manifestations of the classic WAS phenotype consist of immunodeficiency, eczema and thrombocytopenia. However, thrombocytopenia and small platelets are the only consistent features of WAS and XLT.

Serum-free coculture system for ex vivo expansion of human cord blood primitive progenitors and SCID mouse-reconstituting cells using human bone marrow primary stromal cells.

Objective: In an attempt to maintain and expand human stem cells, many investigators have used xenogeneic, especially murine, stromal cells and fetal calf serum. Because of the possible transmission of infectious diseases, however, the safety of the delivery of grafts expanded in culture using xenogeneic cells and serum has been debated. Using primary human marrow stromal cells, we established a novel serum-free culture system to expand human primitive progenitors and transplantable stem cells.

Thrombopoietin and interleukin-2 induce association of CRK with STAT5.

Crk (Crk I and II) proteins and closely related CrkL are adapters which are commonly involved in various signaling processes in various cells, and these proteins share many ligands. Whether they have redundant or distinct physiologic roles is unclear. By coprecipitation and far Western blotting analysis, we demonstrate that Crk (I/II) binds to tyrosine phosphorylated STAT5 in cells stimulated by cytokines such as thrombopoietin (TPO) and interleukin-2 (IL-2).

Elucidation of the HIV-1 virucidal mechanism of methylene blue photosensitization and the effect on primary isolates.

The antiviral activity for primary isolates of human immunodeficiency virus (HIV) type 1 of a combination of methylene blue and light irradiation was investigated, in comparison with their virucidal effects on laboratory-adapted HIV-1. The antiviral mechanism was evaluated in terms of reverse transcriptase activity and viral RNA in the same viral stock. Despite a marked reduction in RNA (>3.

Removal of parvovirus B19 from hemoglobin solution by nanofiltration.

Parvovirus B19 (B19), which may contaminate red cell components for blood transfusion, is known to be resistant to several viral inactivation methods. To increase the safety of hemoglobin solutions as a source of red cell substitutes, we investigated the removal of parvovirus B19 from hemoglobin solution using nanofiltration. The hemoglobin solution spiked with parvovirus B19 was tangentially filtered using the BMM-35 filter (mean pore size of 35 nm) followed by BMM-15.

Comparison of sensitivity to ultraviolet B irradiation between human lymphocytes and hematopoietic stem cells.

To investigate the clinical applicability of prophylaxis of post-transplant graft-versus-host disease by UV-B irradiation of stem cell preparations, the UV-B sensitivities of human lymphocytes and primitive hematopoietic progenitors were compared. The mononuclear cell fractions (MNC) derived from human cord blood and granulocyte-colony-stimulating factor-mobilized peripheral blood were used. After UV-B irradiation, lymphocyte proliferation ability, hematopoietic colony-forming cells, and apoptotic cells were analyzed.

Elimination of both cell-free and cell-associated HIV infectivity in plasma by a filtration/methylene blue photoinactivation system.

Background: Methylene blue phototreatment effectively inactivates cell-free viruses in plasma while maintaining coagulation activities. However, this treatment is considered to be less effective for cell-associated virus inactivation. This report describes a new virus elimination system designed to eliminate cell-associated viruses with a cell-removal filter followed by methylene blue photoinactivation of cell-free viruses in plasma.

Signal transduction in primary cultured human erythroid cells.

Development of erythrocytes is a complex process governed by multiple cytokines. Colony assays have revealed the physiologic importance of these cytokines, although biochemical studies of highly purified human colony-forming unit-erythroid (CFU-E) generated in vitro from CD34+ cells have only recently begun. Studies from our groups and others suggested that signal transduction in primary erythroid cells differs considerably from that in cell lines or primary cells from other species.

Lipopolysaccharide-triggered desensitization of TNF-alpha mRNA expression involves lack of phosphorylation of IkappaBalpha in a murine macrophage-like cell line, P388D1.

Activation of nuclear factor kappaB (NF-kappaB) is thought to be required for cytokine production by lipopolysaccharide (LPS)-responsive cells. Here, we investigated the contribution of NF-kappaB in preventing LPS-induced transcription of the tumor necrosis factor alpha (TNF-alpha) gene in a murine macrophage cell line, P388D1, when tolerance was induced in the cells with a short exposure to a higher dose of LPS. Electrophoretic mobility shift assays with the kappaB elements of the murine TNF-alpha promoter and enhancer revealed that nuclear mobilization of heterodimers of p65/p50, c-rel/p50 and p65/c-rel, and homodimers of p65 was markedly reduced in LPS-tolerant cells, whereas that of p50 homodimers was only slightly increased.

Stromal cell-independent differentiation of human cord blood CD34+CD38- lymphohematopoietic progenitors toward B cell lineage.

To study the cytokine regulation of early stages of human B-lymphopoiesis, we developed a stroma-free two-step culture system. Single human cord blood CD34+CD38- cells were individually cultured by micromanipulation with interleukin (IL)-3, stem cell factor (SCF), fIt3 ligand (FL), IL-6 and granulocyte colony-stimulating factor (G-CSF). About 10% of the cells formed primary colonies, which were individually tested for myeloid and B-lymphoid potentials by reculturing aliquots of the primary colony cells into secondary myeloid and B-lymphoid cultures.

Efficiency of donor screening for human parvovirus B19 by the receptor-mediated hemagglutination assay method.

Background And Objectives: Mass screening for parvovirus B19 (B19) by receptor-mediated hemagglutination assay (RHA) may be inadequate to eliminate the virus from plasma pools. We characterized B19 carriers detected in blood donor screening by RHA to explain why some carriers were not detected by RHA.

Materials And Methods: Donor plasma was screened for B19 by RHA, B19 DNA by nested polymerase chain reaction (PCR) assay, and anti-B19 by enzyme immunoassay.

Analysis of soluble interleukin 6 receptor in cerebrospinal fluid in inflammatory and non-inflammatory conditions.

The objective of this study was to investigate the pathophysiological roles of soluble interleukin 6 receptor (sIL-6R) in cerebrospinal fluid (CSF). CSF was obtained from patients suspected with meningitis. Eight patients without any meningeal signs or symptoms were enrolled as controls.

Virus photoinactivation in stroma-free hemoglobin with methylene blue or 1,9-dimethylmethylene blue.

Photoinactivation of vesicular stomatitis virus (VSV) in stroma-free hemoglobin (SFH) was carried out using methylene blue (MB) or 1,9-dimethylmethylene blue (DMMB). The VSV was more sensitive to inactivation by 660 nm light with 1 microM DMMB than with the same concentration of MB. Under conditions that inactivated 6 log10 of VSV, the methemoglobin content (Met-Hb[%]) and P50 of hemoglobin were changed by 1 microM MB phototreatment but were not changed by 1 microM DMMB phototreatment.

Photoinactivation of vesicular stomatitis virus with fullerene conjugated with methoxy polyethylene glycol amine.

Fullerene is known to effectively produce mainly singlet oxygen by absorbing light energy, and therefore may be useful as a virucidal photosensitizer. This study was designed to investigate the virucidal activity of a water-soluble fullerene derivative, which is conjugated with methoxy polyethylene glycol amine to enhance its water solubility. Vesicular stomatitis virus (VSV) was inactivated upon illumination with this water-soluble fullerene, in a concentration- or dose-dependent manner.

[Set-up of standard value by sex and age of biochemical tests derived from blood donors].

We set up the standard value of 7 biochemical tests from the data of approximately 237,000 blood donors who are healthy at least objectively. The abnormal data were excluded by Grubbs Smirnov method, and then converted by the log equation in order to simulate to normal distribution. Then the reference interval was set at mean +/- 2SD.

[Quality control of cryopreserved cord blood cells for transplantation].

Quality control (QC) of cryopreserved cord blood (CB) cells is very important for the safety of transplantation. In this article, the cell processing of CB cells, the laboratory tests for CB specimens, and the system of QC in CB bank is discussed.