Human factors in chemical containment laboratory design

A human factors engineering analysis of a chemical containment laboratory was performed to develop appropriate standards for future laboratory design. In order to perform this evaluation, a state-of-the-art facility was studied in depth. Measurements and observations were made of key operating areas. In addition, technicians were observed and interviewed as they performed various tasks. Compiled data were compared to existing ergonomic standards. The occupational implications of instituting ergonomically developed laboratory design standards are as follows: increased worker health, safety and productivity, improved work quality and reduced stress.     

Ultrasensitive analysis of the intestinal absorption and compartmentalization of aluminium in uraemic rats: a 26Al tracer study employing accelerator mass spectrometry

BACKGROUND: Developments in accelerator mass spectrometry (AMS) now permit the determination of femtogram amounts of 26Al in blood and in various tissues with good precision and free of external contamination. METHODS: In the present study we used trace quantities of 26Al to investigate the intestinal absorption and compartmentalization of aluminium in rats with renal failure (Nx, 5/6 nephrectomy) and in pair- fed controls (C). Single oral doses of 20 ng 26Al were administered to six animals in each group and, subsequently, 24-h post-load 26Al was analysed in serum, urine, bone, liver, and spleen by means of AMS. RESULTS: Serum concentrations of 26Al were significantly lower in uraemic rats compared to controls, whereas urinary excretion was comparable (Nx, 7.11 +/- 5.78 pg/day vs C, 9.46 +/- 6.10 pg/day), suggesting a higher fraction of ultrafiltrable serum 26Al in uraemia. The target tissues of cellular transferrin-mediated 26Al uptake, liver and spleen, tended to show a larger degree of aluminium accumulation in controls (0.26 +/- 0.31 pg/g vs Nx, 0.14 +/- 0.10 pg/g and 0.37 +/- 0.27 pg/g vs Nx, 0.25 +/- 0.27 pg/g respectively). In contrast, in bone, a site of extracellular aluminium deposition, 26Al concentrations were more elevated in uraemia (1.22 +/- 0.59 pg/g vs C: 0.68 +/- 0.30 pg/g). Estimated total 26Al accumulation in all measured target tissues was significantly higher in uraemic rats (28.15 +/- 9.90 pg vs C: 17.03 +/- 7.03 pg) and total recovery of 26Al from tissue and urine was 26.58 +/- 6.74 pg in controls and 35.75 +/- 7.03 pg in uraemic animals, suggesting a fractional absorption of 0.133% and 0.175% respectively. CONCLUSIONS: Our data suggest that fractional absorption from a dietary level dose of 26Al is about 0.13%. Compartmentalization occurs in transferrin-dependent target tissues such as liver and spleen; however, in quantitative terms extracellular deposition in bone is more important. Uraemia has a significant effect on the intestinal absorption and compartmentalization of aluminium. It enhances fractional absorption and increases subsequent extracellular deposition of aluminium in bone. However, at the same time uraemia does not increase transferrin-dependent cellular accumulation of aluminium in liver and spleen.      

Immunological memory function of the T and B cell types: distribution over mouse spleen and lymph nodes

Spleens from LAF₁ mice injected intravenously with sheep erythrocytes (SE) are relatively rich in memory T cells early in the immune response (1 to 3 days) and rich in memory B cells as the response progresses (2 weeks or more). Marked cooperation for the secondary immune response in vitro was obtained by combining 10⁶ spleen cells from LAF₁ mice, taken 2 days after intravenous priming with SE, with 10⁷ spleen cells from day 14 primed mice. The results indicate relative deficiencies in the spleen for B memory cells on days 1 to 2 and for T memory cells on day 14 after priming. Day – 14, but not day – 2, immune lymph node (LN) cells could replace the day – 2 spleen cells (anti-Thy 1.2 sensitive) in the in vitro cooperation with day – 14 immune spleen cells. Immune spleen cells taken 4 to 7 days after priming contain more equivalent numbers of B and T memory cells, but 10 to 7 days after transfer of such immune spleen cells without SE into irradiated recipients the T memory cells were again more prominent in lymph node and the B memory cells in spleen as shown by in vitro cooperation studies. These results suggest that during the second week after intravenous injection of SE relatively more T than B memory cells migrate from spleen to lymph node, resulting in an imbalance in the splenic memory cell population favoring B memory cell function.     

Absence of demonstrable phospholipid turnover in B cells stimulated by low mitogenic concentrations of dextran-anti-immunoglobulin conjugates.

Previously we have demonstrated that when anti-immunoglobulin (Ig) is conjugated to high molecular weight dextran (Dex) it stimulates B cell activation at pg/ml concentrations in the absence of detectable phosphoinositide hydrolysis or increases in intracellular ionized calcium. To study carefully whether anti-Ig-Dex recruited a phosphoinositide-dependent pathway of activation, we stimulated B cells that were labeled with 32P and [3H]glycerol with anti-Ig-Dex conjugates at concentrations ranging from 1-1 x 10(-4) micrograms/ml. Thirty seconds to thirty minutes after stimulation lipids were extracted and analyzed by thin layer chromatography and spots correlating with known lipid standards were isolated and counted. There was a four- and tenfold increase in the ratio of 32P/3H incorporated into phosphatidic acid (a metabolite of diacylglycerol) and phosphatidylinositol, respectively, when cells were stimulated with 0.1-1.0 microgram/ml of anti-Ig-Dex for 30 min. Below 1 ng/ml there was no detectable increase in the turnover of these metabolites despite the fact that in parallel cultures B cells were stimulated to proliferate by this concentration of anti-Ig-Dex. To determine whether a cAMP-dependent pathway was recruited by low concentrations of conjugates, we evaluated cAMP levels from B cells that were stimulated with anti-Ig-Dex for 5-60 min using a radioimmunoassay. While cholera toxin stimulated a 50-100-fold increase in the levels of cAMP, we observed no alteration in cAMP in anti-Ig-stimulated cells. These results support and extend our previous findings by demonstrating that B cell activation that is induced by cross-linking of surface Ig may not stimulate phosphoinositide-dependent or cAMP-dependent pathways of activation. Possible alternative mechanisms of activation will be discussed.     

Distinct types of T-cell help for the induction of a humoral immune response to Streptococcus pneumoniae

Studies have indicated that purified soluble polysaccharide antigens can elicit T cell-independent Ig responses in vivo, although these responses can be modulated by T cells in a noncognate manner. Relatively little is known, however, concerning the parameters that regulate polysaccharide-specific, as well as protein-specific, Ig isotype responses to an intact extracellular bacterium. Using the murine in vivo humoral response to intact Streptococcus pneumoniae as a model it can be shown that CD4+ T-cell receptor alphabeta+ T cells deliver help for both polysaccharide- and protein-specific Ig responses. However, these responses differ fundamentally in their mechanism of action.     

Early treatment with ganciclovir to prevent cytomegalovirus disease after allogeneic bone marrow transplantation.

BACKGROUND. Cytomegalovirus (CMV) infection is a major cause of morbidity and mortality after allogeneic bone marrow transplantation. We conducted a controlled trial of ganciclovir for the early treatment of CMV infection in asymptomatic recipients of bone marrow transplants whose surveillance cultures for CMV became positive. METHODS. Bone marrow--allograft recipients who were seropositive for CMV antibodies or who received seropositive marrow were screened for CMV excretion by culture of throat swabs, blood, urine, or bronchoalveolar-lavage fluid. In this double-blind trial, 72 patients who had marrow engraftment and were excreting virus were randomly assigned to receive either placebo or ganciclovir (5 mg per kilogram of body weight twice a day for one week, followed by 5 mg per kilogram per day) for the first 100 days after transplantation. Patients were followed for the development of biopsy-confirmed CMV disease, ganciclovir-related toxicity, and survival. RESULTS. Between assignment to the study drug and day 100 after transplantation, CMV disease developed in only 1 of the 37 patients assigned to receive ganciclovir (3 percent), but in 15 of the 35 patients assigned to receive placebo (43 percent, P less than 0.00001). The ganciclovir recipients had rapid suppression of virus excretion; 85 percent had negative cultures after one week of treatment, as compared with 44 percent of the placebo group (P = 0.001). The principal toxic reaction was neutropenia; 11 ganciclovir recipients had an absolute neutrophil count below 0.75 x 10(9) per liter, as compared with 3 placebo recipients (P = 0.052). Treatment was discontinued in 11 ganciclovir recipients and 1 placebo recipient because of neutropenia (P = 0.003). After treatment was stopped, the neutrophil count recovered in all patients. Overall survival was significantly greater in the ganciclovir group than in the placebo group both 100 days and 180 days after transplantation (P = 0.041 and 0.027, respectively). CONCLUSIONS. Early treatment with ganciclovir in patients with positive surveillance cultures reduces the incidence of CMV disease and improves survival after allogeneic bone marrow transplantation.     

A novel tetrameric gp350 as a potential Epstein–Barr virus vaccine

Infectious mononucleosis and B-cell transformation in response to infection with Epstein–Barr virus (EBV) is dependent upon binding of the EBV envelope glycoprotein gp350 to CD21 on B-cells. Gp350-specific antibody comprises most of the EBV neutralizing activity in the serum of infected patients, making this protein a promising target antigen for a prophylactic EBV vaccine. We describe a novel, tetrameric gp350-based vaccine that exhibits markedly enhanced immunogenicity relative to its monomeric counterpart. Plasmid DNA was constructed for synthesis, within transfected CHO cells, of a tetrameric, truncated (a.a. 1–470) gp350 protein (gp350^1–470). Tetrameric gp350^1–470 induced ∼20-fold higher serum titers of gp350^1–470-specific IgG and >19-fold enhancements in neutralizing titers at the highest dose, and was >25-fold more immunogenic on a per-weight basis than monomeric gp350^1–470. Further, epidermal immunization with plasmid DNA encoding gp350^1–470 tetramer induced 8-fold higher serum titers of gp350^1–470-specific IgG relative to monomer. Tetrameric gp350^1–470 binding to human CD21 was >24-fold more efficient on a per-weight basis than monomer, but neither tetramer nor monomer mediated polyclonal human B-cell activation. Finally, the introduction of strong, universal tetanus toxoid (TT)-specific CD4+ T-cell epitopes into the tetrameric gp350^1–470 had no effect on the gp350^1–470-specific IgG response in naïve mice, and resulted in suppressed gp350^1–470-specific IgG responses in TT-primed mice. Collectively, these data suggest that tetrameric gp350^1–470 is a potentially promising candidate for testing as a prophylactic EBV vaccine, and that protein multimerization, using the approach described herein, is likely to be clinically relevant for enhancing the immunogenicity of other proteins of vaccine interest.