Human blood autoantibodies in the detection of colorectal cancer

Colorectal cancer (CRC) is the second most common malignancy in the western world. Early detection and diagnosis of all cancer types is vital to improved prognosis by enabling early treatment when tumours should be both resectable and curable. Sera from 3 different cohorts; 42 sera (21 CRC and 21 matched controls) from New York, USA, 200 sera from Pittsburgh, USA (100 CRC and 100 controls) and 20 sera from Dundee, UK (10 CRC and 10 controls) were tested against a panel of multiple tumour-associated antigens (TAAs) using an optimised multiplex microarray system. TAA specific IgG responses were interpo- lated against the internal IgG standard curve for each sample. Individual TAA specific responses were examined in each cohort to determine cutoffs for a robust initial scoring method to establish sensitivity and specificity. Sensitivity and specificity of combinations of TAAs provided good discrimination between cancer-positive and normal serum. The overall sensitivity and specificity of the sample sets tested against a panel of 32 TAAs were 61.1% and 80.9% respectively for 6 antigens; p53, AFP, K RAS, Annexin, RAF1 and NY-CO16. Furthermore, the observed sensitivity in Pittsburgh sample set in different clinical stages of CRC;stageI(n=19),stageII(n=40),stageIII(n=34)andstageIV(n=6)wassimilar (73.6%, 75.0%, 73.5% and 83.3%, respectively), with similar levels of sensitivity for right and left sided CRC. We identified an antigen panel of sufficient sensitivity and specificity for early detection of CRC, based upon serum profiling of autoantibody response using a robust multiplex antigen microarray technology. This opens the possibility of a blood test for screening and detection of early colorectal cancer. However this panel will require further validation studies before they can be proposed for clinical practice

The Potential and Challenges of Nanopore Sequencing

A nanopore-based device provides single-molecule detection and analytical capabilities that are achieved by electrophoretically driving molecules in solution through a nano-scale pore. The nanopore provides a highly confined space within which single nucleic acid polymers can be analyzed at high throughput by one of a variety of means, and the perfect processivity that can be enforced in a narrow pore ensures that the native order of the nucleobases in a polynucleotide is reflected in the sequence of signals that is detected. Kilobase length polymers (single-stranded genomic DNA or RNA) or small molecules (e.g., nucleosides) can be identified and characterized without amplification or labeling, a unique analytical capability that makes inexpensive, rapid DNA sequencing a possibility. Further research and development to overcome current challenges to nanopore identification of each successive nucleotide in a DNA strand offers the prospect of ‘third generation’ instruments that will sequence a diploid mammalian genome for ~$1,000 in ~24 h.Molecular and Cellular BiologyPhysic

Cyanobacterial lipopolysaccharides and human health – a review

Cyanobacterial lipopolysaccharide/s (LPS) are frequently cited in the cyanobacteria literature as toxins responsible for a variety of heath effects in humans, from skin rashes to gastrointestinal, respiratory and allergic reactions. The attribution of toxic properties to cyanobacterial LPS dates from the 1970s, when it was thought that lipid A, the toxic moiety of LPS, was structurally and functionally conserved across all Gram-negative bacteria. However, more recent research has shown that this is not the case, and lipid A structures are now known to be very different, expressing properties ranging from LPS agonists, through weak endotoxicity to LPS antagonists. Although cyanobacterial LPS is widely cited as a putative toxin, most of the small number of formal research reports describe cyanobacterial LPS as weakly toxic compared to LPS from the Enterobacteriaceae. We systematically reviewed the literature on cyanobacterial LPS, and also examined the much lager body of literature relating to heterotrophic bacterial LPS and the atypical lipid A structures of some photosynthetic bacteria. While the literature on the biological activity of heterotrophic bacterial LPS is overwhelmingly large and therefore difficult to review for the purposes of exclusion, we were unable to find a convincing body of evidence to suggest that heterotrophic bacterial LPS, in the absence of other virulence factors, is responsible for acute gastrointestinal, dermatological or allergic reactions via natural exposure routes in humans. There is a danger that initial speculation about cyanobacterial LPS may evolve into orthodoxy without basis in research findings. No cyanobacterial lipid A structures have been described and published to date, so a recommendation is made that cyanobacteriologists should not continue to attribute such a diverse range of clinical symptoms to cyanobacterial LPS without research confirmation

LASER-INDUCED FLUORESCENCE DETECTION OF RHODAMINE-6G AT 6 X 10-15 M

We have demonstrated sensitive fluorescence detection of rhodamine-6G in aqueous solutions down to approximately 10(-15) M with a detection limit of 6 x 10(-15) M. The working curve was observed to be linear over four orders of magnitude and could be extended to higher concentrations. Single molecules of rhodamine-6G had a transit time of 1.8 ms through the 11 pL probe volume. At 2 x 10(-14) M, the probability of a molecule being in the 11 pL probe volume is approximately 0.1. Analysis of the data indicates that the signal-to-noise for the detection of a single molecule of rhodamine-6G is approximately 1.open1113sciescopu

SINGLE-MOLECULE DETECTION OF RHODAMINE-6G IN ETHANOLIC SOLUTIONS USING CONTINUOUS WAVE LASER EXCITATION

The ultimate in analytical sensitivity is the ability to detect analytes on a single-molecule level. Laser-induced fluorescence (LIF) detection of single molecules in solution is hampered by specular, Rayleigh, and Raman scattering that contribute significantly to the background. In order to observe individual fluorescent molecules as they transit the laser beam in the presence of large backgrounds, it is necessary to detect a large number of photons per molecule. One method to increase the number of photons per event is to increase the residence time of the molecule in the laser beam. However, with long residence times, photostability sets an upper limit on the number of times the molecule can be cycled between the ground and first excited singlet state. We have observed the passage of individual rhodamine 6G (R-6G) molecules in ethanol (EtOH). The use of EtOH as a solvent allows one to obtain nearly 2 orders of magnitude more photons per molecule than may be obtained in H2O. Observation of single molecular events of R-6G in EtOH is substantiated by autocorrelation analysis and from shifts in the histograms of the frequency of photoelectron counts. Results from Monte Carlo simulations also support our experimental results.X11101sciescopu