Analysis of mono-phosphate nucleotides as a potential method for quantification of DNA using high performance liquid chromatography-inductively coupled plasma-mass spectrometry

The determination of total deoxyribonucleic acid (DNA) concentration is of great importance in many biological and bio-medical analyses. The quantification of DNA is traditionally performed by UV spectroscopy; however the results can be affected greatly by the sample matrix. The proposed method quantifies phosphorus in digested calf thymus DNA and human DNA by high performance liquid chromatography (HPLC) coupled to inductively coupled plasma mass spectrometry (ICP-MS). The method presented showed excellent baseline separation between all 4 DNA mono-nucleotides and 5’UMP. Column recoveries ranging from 95% to 99% for phosphorus resulted in a mass balance of 95% ± 0.5% for standard nucleotides, determined by LC-ICP-MS, compared to total DNA determined by flow injection coupled to ICP-MS (FI-ICP-MS). The ability of LC-ICPMS to act as an internal check that only DNA derived phosphorus was counted in the assay was demonstrated by establishing a mass balance between the total phosphorous signal from undigested DNA and that from the speciated DNA. The method for quantification was evaluated by analysis of NIST SRM 2372; a total speciated DNA recovery of 52.1 ng/μL, compared with an expected value of 53.6 ng/μL, was determined by external calibration. From repeat measurements a mass balance of 97% ± 0.5% for NIST DNA was achieved. The method limits of detection for individual nucleotides were determined between 0.8 to 1.7 μg L-1 (31P) for individual nucleotides by LC-ICP-MS, and 360 ng L-1 for 5’AMP by direct nebulisation

Anti-tumour therapeutic efficacy of OX40L in murine tumour model

OX40 ligand (OX40L), a member of TNF superfamily, is a co-stimulatory molecule involved in T cell activation. Systemic administration of mOX40L fusion protein significantly inhibited the growth of experimental lung metastasis and subcutaneous (s.c.) established colon (CT26) and breast (4T1) carcinomas. Vaccination with OX40L was significantly enhanced by combination treatment with intra-tumour injection of a disabled infectious single cycle-herpes simplex virus (DISC-HSV) vector encoding murine granulocyte macrophage-colony stimulating factor (mGM-CSF). Tumour rejection in response to OX40L therapy required functional CD4+ and CD8+ T cells and correlated with splenocyte cytotoxic T lymphocytes (CTLs) activity against the AH-1 gp70 peptide of the tumour associated antigen expressed by CT26 cells. These results demonstrate the potential role of the OX40L in cancer immunotherapy

Corrected and Republished from: "A Novel, Multiple-Antigen Pneumococcal Vaccine Protects against Lethal Streptococcus pneumoniae Challenge"

Current vaccination against Streptococcus pneumoniae uses vaccines based on capsular polysaccharides from selected serotypes and has led to nonvaccine serotype replacement disease. We have investigated an alternative serotype-independent approach, using multiple-antigen vaccines (MAV) prepared from S. pneumoniae TIGR4 lysates enriched for surface proteins by a chromatography step after culture under conditions that induce expression of heat shock proteins (Hsp; thought to be immune adjuvants). Proteomics and immunoblot analyses demonstrated that, compared to standard bacterial lysates, MAV was enriched with Hsps and contained several recognized protective protein antigens, including pneumococcal surface protein A (PspA) and pneumolysin (Ply). Vaccination of rodents with MAV induced robust antibody responses to multiple serotypes, including nonpneumococcal conjugate vaccine serotypes. Homologous and heterologous strains of S. pneumoniae were opsonized after incubation in sera from vaccinated rodents. In mouse models, active vaccination with MAV significantly protected against pneumonia, while passive transfer of rabbit serum from MAV-vaccinated rabbits significantly protected against sepsis caused by both homologous and heterologous S. pneumoniae strains. Direct comparison of MAV preparations made with or without the heat shock step showed no clear differences in protein antigen content and antigenicity, suggesting that the chromatography step rather than Hsp induction improved MAV antigenicity. Overall, these data suggest that the MAV approach may provide serotype-independent protection against S. pneumoniae

Distribution of landslides and geotechnical properties within the Hampshire Basin

This paper outlines the sedimentary sequences and geotechnical properties of the Hampshire Basin, a basin of filled with 700 m of Palaeogene clays, silts, sands and limestones in southern England. The paper presents results so far of a study to synthesize relevant geological and geotechnical data and relate these to the nature of the landslides in this basin. The study has found that stratigraphic sequences and geotechnical properties vary considerably across the basin owing to basin morphology and depositional environments which are correspond to complex paleogeography and tectonic movements during the Tertiary. Over-consolidated clays with low residual shear strengths are extensive on moderately steep slopes and prone to landsliding, especially on over-steepened coastal sections. Landslides vary from mudflows through mudslides, rotational landslides and minor falls. Landslide characteristics are strongly influenced by lithology but gradient appears to be the controlling factor in many cases. The presences of weak strata (clays, lignite, laminated layers), the pre-existing shear surface, the lithological interface (sand overlying clay) play important roles to locally control the position of the shear surface and the type of movements. At a basin scale, inland landslides are associated with the development of drainage system during and since the Tertiary