Intra-operative support via augmented reality techniques in endoscopic surgery.

Computer assisted operation planning systems are gaining increasing recognition in the field of surgery. These systems offer new possibilities for preparing an intervention, with the goal of reducing the amount of expensive operating-room time required for the intervention. The safest and most effective surgical approach should always be selected, but it is often difficult to transfer the output of the planning system to the intra-operative situation so that the planning results can be considered during the actual intervention. At the Fraunhofer Institute for Computer Graphics (IGD) in Darmstadt and the Centre for Advanced Media Technology (CAMTech) in Singapore methods are being developed to bridge the gap between the external planning session and the intra-operative case: Augmented Reality (AR) techniques are used to overlay preoperative scanned image data, as well as results of the planning session, on the operation field.     

Development of an ELISA for detection of an organophosphorus compound using monoclonal antibodies

This paper describes a specific and highly sensitive ELISA system using monoclonal antibodies in order to assay an organophosphorus compound. The soman derivative methyl phosphonic acid, p-aminophenyl 1,2,2,-trimethyl-propyl diester (MATP) served as model substance. In order to obtain antibody-producing hybridomas BALB/c mice were immunized with MATP linked onto human serum albumin (HSA). The spleen cells of immunized mice were fused with syngenic plasmacytomas of the non-producer-line X63Ag8.653 with the aid of polyethylene glycol. To eliminate undesirable cross-reaction, common screening procedures were modified by directly coating the ELISA plates with hapten. Five out of 15 positive cell-lines were cloned by limiting dilution and further propagated. The respective immunoglobulin class and subclass of the obtained monoclonal antibodies was determined. Four of which were identified as IgG₁, the other as IgG_2a. After enrichment of antibodies in ascites and their isolation by protein A-sepharose, the affinity of various monoclonal antibodies was estimated in competitive inhibition enzyme immunoassay (CIEIA) by measuring the IC₅₀ rates of free MATP. The rates were found to lie between 2.5 × 10^–6 mol/l and 4.3 × 10^–4 mol/l MATP. The IC₁₀ rate for detectable MATP concentration was 5.4 × 10^–7 mol/l MATP. Test duration was 280 min. The reactivity of the monoclonal antibodies with structurally related substances was used to check their specificity. Cross-reaction turned out to be negative. In order to develop a direct competitive ELISA, MATP was linked to horse radish peroxidase (HRPO) by adding a spacer. This helped to reduce total duration to 40 min. The detection level was further reduced to 1.3 × 10^–7 mol/l MATP (corresponding to 975 pg/25 l test-buffer) using the monoclonal antibody F71D7. Likewise, MATP was detected in goat serum, chicken serum, rabbit serum, milk and company's water in concentrations between 2.1 × 10^–7 mol/l (IC₁₀, company's water) and 4.9 × 10^–8 mol/l (IC₁₀, milk).     

Change in sympathetic activity,cardiovascular functions and plasma hormone concentrations due to cold water immersion in men

The purpose of this study was to determine whether or not repeated short-term cold water immersions can induce a change in the activity of the sympathetic nervous system and, consequently, in cardiovascular functions in healthy young athletes. Changes in some plasma hormone concentrations were also followed. A single cold water immersion (head-out, at 14°C, for 1 h) increased sympathetic nervous system activity, as evidenced by a four-fold increase (P < 0.05) in plasma noradrenaline concentration. Plasma adrenaline and dopamine concentrations were not increased significantly. Plasma renin-angiotensin activity was reduced by half (P < 0.05) during immersion but plasma aldosterone concentration was unchanged. Stimulation of the sympathetic nervous system during immersion did not induce significant changes in heart rate, but induced peripheral vasoconstriction (as judged from a decrease in skin temperature) and a small increase (by 10%) in systolic and diastolic blood pressures. No clear change in reactivity of the sympathetic nervous system was observed due to repeated cold water immersions (three times a week, for 6 weeks). Neither the plasma renin-angiotensin activity, aldosterone concentration nor cardiovascular parameters were significantly influenced by repeated cold water immersions. A lowered diastolic pressure and an increase in peripheral vasoconstriction were observed after cold acclimation, however. Evidently, the repeated cold stimuli were not sufficient to induce significant adaptational changes in sympathetic activity and hormone production.     

CrELISA: a fast and robust enzyme-linked immunosorbent assay bypassing the need for purification of recombinant protein

A multitude of antigens has been recently identified by screening of cDNA expression libraries derived from human tumors with autologous sera. Using a phage autoantibody assay and small panels of sera derived from cancer patients or controls it has been shown that some of these antigens display cancer-associated autoantibody responses. The diagnostic and prognostic significance of these potentially cancer-related autoantibodies remains unclear until large-scale assays are developed and serological data are available for hundreds of cancer patients and controls. The major bottleneck for the development of large-scale assays are the cloning, expression and the purification of each of the respective antigens. Due to these limitations and despite the potential clinical relevance large-scale autoantibody tests are established for only a few of these tumor antigens. Here we describe an enzyme-linked immunosorbent assay, Crude lysate ELISA (CrELISA), suitable for antigens identified by expression screening based on crude lysates of antigen-expressing bacteria. This assay permits sensitive and specific autoantibody seroscreening without the need of laborious and time-consuming cloning, expression and purification of recombinant proteins. CrELISA is robust and provides a versatile high throughput procedure for the rapid evaluation of multiple antigens in large-scale serology.     

Phenotypic and genotypic analyses of enterohemorrhagic Escherichia coli O145 strains from patients in Germany

Enterohemorrhagic Escherichia coli (EHEC) strains of serogroup O145 are emerging as causes of diarrhea and the hemolytic-uremic syndrome. However, there have been few genetic analyses of this EHEC group. We investigated the serotypes, virulence genes, plasmid profiles, pulsed-field gel electrophoresis (PFGE) patterns, and genetic variability of the fliC and eae genes in 120 EHEC O145 strains isolated from cases of hemolytic-uremic syndrome (n = 24) or diarrhea (n = 96) in Germany between 1996 and 2002. Three isolates belonged to serotype O145:H28, one to serotype O145:H25, and 116 were nonmotile (O145:H(-)). One hundred fourteen of the nonmotile strains shared fliC restriction fragment length polymorphism (RFLP) patterns identical to that of the O145:H28 strains. The remaining two nonmotile strains displayed a fliC-RFLP pattern identical to that of the O145:H25 strain. Each of the 117 strains with the fliC-RFLP(H28) pattern harbored eae gamma, whereas the three strains with the fliC-RFLP(H25) pattern possessed eae beta. Five different stx genotypes, six combinations of plasmid-encoded putative virulence genes, 29 plasmid profiles, and 47 PFGE types were identified. Strains within some of the PFGE types could be further subtyped by means of distinct plasmid profiles. These data demonstrate that the EHEC O145 serogroup is comprised of two different serotypes that possess distinct eae types. The heterogeneity of EHEC O145 strains at the chromosomal and plasmid level, in particular the high diversity in PFGE patterns, provides a basis for molecular subtyping of these pathogens.     

Strong Association Between Shiga Toxin-Producing Escherichia coli O157 and Virulence Genes stx 2 and eae as Possible Explanation for Predominance of Serogroup O157 in Patients with Haemolytic Uraemic Syndrome

The aim of this study was to investigate the association between Shiga toxin-producing Escherichia coli (STEC) O157 and the simultaneous presence of the virulence genes stx ₂ and eae in STEC from patients with gastroenteritis. In Germany, the proportion of serogroup O157 is substantially higher among STEC isolates from patients with haemolytic uraemic syndrome (HUS) than among STEC isolates from patients with gastroenteritis. The reason for this is unknown. Independent of serogroup, the virulence genes stx ₂ and eae have been associated with severe disease. Data collected in 2000–2001 from a Germany-wide laboratory-based surveillance system for STEC-associated gastroenteritis in patients <15 years were analysed. Overall, 18% of the STEC isolates belonged to serogroup O157. Compared with non-O157 strains, 0157 isolates were strongly associated with the simultaneous presence of both an stx ₂ gene and the eae gene (OR, 76; 95%CI, 27–230). Within the subset of STEC isolates that carried both virulence genes, 60% belonged to serogroup O157, a proportion similar to that found in STEC isolates from pediatric patients with HUS in Germany and Austria (67%, P=0.35). These data suggest that the more frequent carriage of both virulence genes, i.e. stx ₂ and eae, forms the basis of why STEC O157 predominates in patients with HUS.     

Detection of the organophosphorus nerve agent soman by an ELISA using monoclonal antibodies

The development of a specific and sensitive immunologic ELISA detection system for methylphosphonoflouridic acid. 1,2,2-trimethylpropylester (soman) by the use of monoclonal antibodies (MAbs) is described. The monoclonal antibodies F71D7, F71H10, F71B12 and F71H9 originally produced against the soman derivative methyl phosphonic acid,p-aminophenyl 1,2,2-trimethylpropyldiester (MATP) also reacted with soman in a previously developed, direct competitive ELISA. After optimizing the ELISA system by varying the reaction mixture and the solvents for the organophosphate, 5.0×10^–7 mol/l soman (80% purity), e.g. 2.5 ng or 2 ng pure soman per 25 l test buffer, could be detected after a total test duration of 40 min. A shortening of the incubation time to 10 min resulted in a drop of sensitivity to 1.8×10^–6 mol/l soman. Various alcohols which may be used as extraction media for soman from various materials (isopropanol, ethanol and methanol) were shown to inhibit peroxidase activity and thereby reduce the sensitivity of the test. However, the influence of alcohols decreased with the shortening of incubation time. All monoclonal antibodies showed little cross reactivity to sarin and no cross reactivity to tabun and VX. Judging on the reactivity of the MAbs with MATP and soman oxidazed by 1,2-dihydrobenzol, some reactivity with some other (non-toxic) soman analogues containing the same pinacolyl group can be expected. There was no evidence for stereoselectivity of the MAbs tested. Finally, soman could be detected in different biological samples like human serum, goat serum, rabbit serum, chicken serum, milk, and tap water in concentrations between 1.3×10^–6 and 2.0×10^–6 mol/l.     

Immunobiology of experimental leishmaniasis

Self-cure versus uncontrolled disease progression in experimental murine cutaneous leishmaniasis depends upon a delicate interplay among various activated cells of the host's immune system. Susceptibility or resistance to infection with Leishmania major is correlated with the ability of different inbred strains of mice to produce the characteristic spectra of lymphokines upon infection. Appropriate experimental interventions now allow the modulation of these reponses, providing the possibility to render genetically susceptible mice resistant to infection and, vice versa, to cause genotypically healer strains to express a non-healer phenotype. These experimental manipulations have proven to be powerful tools in the dissection of the underlying immune mechanisms and cellular parameters responsible for susceptibility and resistance, and will perhaps allow the identification of molecules of parasite origin that induce deleterious immune responses to infection with Leishmania, and thus to exclude them from future vaccines. More importantly, rational immune intervention could permit the diversion of established host-damaging immune responses to host-protective immunization.