Biodeterioration of cinematographic cellulose triacetate by Sphingomonas paucimobilis using indirect impedance and chemiluminescence techniques

The bacterium Sphingomonas paucimobilis, isolated from cinematographic films in an earlier project, was able to biodeteriorate the cellulose triacetate material (acetylation degree of 2.7). Film colonization was monitored by the indirect impedance technique and the production of carbon dioxide. The presence of ruggedness and irregularities on the surface of the film, produced when the plasticizer was extracted, accelerated the biodeterioration of the material. In contrast, cinematographic films with their layered structure made of photographic gelatine emulsion were protected and no colonization was observed. The cinematographic film without photographic emulsion reached a 5% level of biodeterioration after six weeks of incubation, confirming the possibility of biodeterioration of archival cinematographic materials if conservation conditions are not adequate. Through viscosity measurements, a decrease in relative viscosity was observed on the biodeteriorated sample, with respect to the original material, confirming a lower molecular weight as a result of enzymatic activity of S. paucimobilis. Also, using chemiluminescence, the film surface oxidation on the biodeteriorated sample was observed. This technique was very sensitive in detecting material oxidation by reactive oxygen species generated by bacteria, and could be useful to study microbiological biodeterioration in polymeric materials.     

Luminol-enhanced chemiluminescence by Listeria monocytogenes: A test for rapid assessment of antimicrobial agents

Abstract Listeria monocytogenes produces chemiluminescence in brain heart infusion broth at 37°C in the presence of carbonate ions and acetaldehyde. This phenomenon can be enhanced by the use of luminol rather than acetaldehyde. Furthermore, there is direct relationship between the extent of growth and the level of luminescence which culminates at the end of the exponential growth. This property was used to study the susceptibility of this bacterium to two antiseptics, cetrimonium bromide and chlorhexidine, and to two antibiotics, ampicillin and chloramphenicol. Inhibition of chemiluminescence was proportional to the antimicrobial agents' concentrations and was complete at their minimal inhibitory concentrations.     

Effects of glucose, anoxia, and adriamycin on the chemiluminescence of Ehrlich Ascites cells

Glucose and anoxia accelerate the photocount due to luminescence of Ehrlich Ascites cells. Adriamycin also has this effect if glucose is present. Comparison with a chemical standard combined with estimates of cellular and population transmittance yield a photon generation rate of at least 10.s⁻¹.cell⁻¹ in the presence of 10⁻² M glucose, and twice this with anoxic conditions or 10⁻⁵ M adriamycin. Effects of adriamycin on Ehrlich Ascites cell respiration may depend on the presence of glucose.     

Molecular markers for plant breeding: comparisons of RFLP and RAPD genotyping costs

Three molecular marker protocols, chemiluminescent restriction fragment length polymorphisms (c-RFLPs), radioactivity-based restriction fragment length polymorphisms (r-RFLPs), and randomly amplified DNA polymorphisms (RAPDs) were compared in terms of cost and time efficiency. Estimates of cost of supplies and time requirements were obtained from simulations of maize (Zea mays L.) genotyping experiments utilizing protocols currently in use. The increase in total cost with increasing numbers of individuals genotyped and markers analyzed is higher for RAPDs than for RFLPs. RAPDs were generally found to be more cost and time efficient for studies involving small sample sizes, while RFLPs have the advantage for larger sample sizes. Because of the shorter exposure times involved, c-RFLPs require less time than r-RFLPs to obtain a given amount of information. Variations in the protocols, such as number of re-uses of Southern blots or cost of Taq DNA polymerase per reaction of amplification, also affect the relative merits of RAPDs and RFLPs. Two examples were analyzed where molecular markers are used: a germ plasm survey and quantitative trait loci (QTL) mapping in a segregating population. No protocol was found to be the most cost and time efficient over the entire range of sample sizes and number of marker loci studied.     

A chemiluminescent metalloimmunoassay based on silver deposition on colloidal gold labels

A sensitive chemiluminescent (CL) immunoassay of human immunoglobulin (IgG) which combined the inherent high sensitivity of CL analysis with the dramatic signal amplification of silver precipitation on colloidal gold tags was developed. First, the sandwich-type complex was formed in this protocol by the primary antibody immobilized on the polystyrene wells, the analyte in the sample, and the secondary antibody labeled with colloidal gold. Second, the colloidal gold was treated by an Ag(+) reduction solution, which resulted in the catalytic precipitation of silver on the surface of colloidal gold. Third, a large number of Ag(+) were oxidatively released in HNO(3) solution from the silver metal anchored on the sandwich-type complexes and then the human IgG was indirectly determined by a sensitive combined CL reaction of Ag(+)-K(2)S(2)O(8)-Mn(2+)- H(3)PO(4)-luminol. The chemiluminescence intensity depends linearly on the logarithm of the concentration of human IgG over the range of 0.02-50ngml(-1) and detection limit (3sigma) is 0.005ngml(-1) (i.e., approximately 3x10(-14)M, 3amol in 100-mul sample). This assay has been successfully applied to the determination of human IgG in human serum samples and showed great potential for numerous applications in immunoassay.     

Influence of beta-glucans on the immune responses of carpet shell clam (Ruditapes decussatus) and Mediterranean mussel (Mytilus galloprovincialis)

The effects of beta-glucans on several immune functions of carpet shell clam (Ruditapes decussatus) and Mediterranean mussel (Mytilus galloprovincialis) hemocytes were determined. Nitric oxide (NO) production increased significantly in beta-glucan treated mussels and clams. In mussels, beta-glucans increased by themselves the release of free oxygen radicals and also were able to enhance the phorbol 12-myristate 13-acetate (PMA) mediated effect on this hemocyte activity. However, high doses of beta-glucans when combined with zymosan decreased this respiratory burst. In clams, hemolymph treated with several doses of beta-glucans limited the growth of the three bacteria, Vibrio algynolyticus (strain TA15), Vibrio splendidus (strain TA2) and Escherichia coli (strain ATCC 13706). This modulation on the antibacterial activity, however, was not observed when mussel hemolymph was incubated with beta-glucans. These results suggest that the immune responses of these animals can be up and down modulated by external stimuli and, although clams and mussels are both relatively closely related species, their behaviour concerning immune responses can be different.     

Determination of uric acid in human urine and serum by capillary electrophoresis with chemiluminescence detection

A simple and sensitive method based on capillary electrophoresis (CE) with chemiluminescence (CL) detection has been developed for the determination of uric acid (UA). The sensitive detection was based on the enhancement effect of UA on the CL reaction between luminol and potassium ferricyanide (K₃[Fe(CN)₆]) in alkaline solution. A laboratory-built reaction flow cell and a photon counter were deployed for the CL detection. Experimental conditions for CL detection were studied in detail to achieve a maximum assay sensitivity. Optimal conditions were found to be 1.0 × 10⁻⁴ M luminol added to the CE running buffer and 1.0 × 10⁻⁴ M K₃[Fe(CN)₆] in 0.2 M NaOH solution introduced postcolumn. The proposed CE-CL assay showed good repeatability (relative standard deviation [RSD] = 3.5%, n = 11) and a detection limit of 3.5 × 10⁻⁷ M UA (signal/noise ratio [S/N] = 3). A linear calibration curve ranging from 6.0 × 10⁻⁷ to 3.0 × 10⁻⁵ M UA was obtained. The method was evaluated by quantifying UA in human urine and serum samples with satisfactory assay results.     

Effect of different inhibitors on the intracellularly and extracellularly generated chemiluminescence induced by formylmethionyl-leucyl-phenylalanine in polymorphonuclear leukocytes. Cellular response in the presence of mannitol,benzoate, taurine,indomethacin and NDGA

When polymorphonuclear leukocytes (PMNL) interact with the soluble stimulus formylmethionyl-leucyl-phenylalanine (FMLP), the cells increase their production of oxidative metabolites. This increased production can be measured as lumino-amplified light emission or chemiluminescence (CL). In the present report, experimental systems which allow a quantitation of extracellularly and intracellularly generated metabolites have been used, and the effect of mannitol, benzoate, taurine, indomethacin and nordihydroguaiaretic acid has been investigated. The presence of the hypochlorous acid scavenger taurine had no effect on the intracellular response, whereas the extracellular response was reduced with around 50%. The hydroxyl radical scavenger mannitol had only minor effects on the response, whereas benzoate, another hydroxyl radical scavenger, reduced the extracellular response with around 50% and the intracellular response with more than 90%. Indomethacin, an inhibitor of arachidonic acid metabolism, did not influence the response, whereas NDGA, also an inhibitor of the arachidonic acid metabolism, totally abolished both the extracellular and the intracellular response. The use of scavengers/inhibitors as a means of determining the mechanisms of light emission, and the origin of chemiluminescence produced by neutrophils stimulated by FMLP is discussed.     

Acridinium ester-labelled DNA oligonucleotide probes

Chemiluminescent acridinium ester derivatives have been synthesized and covalently attached to suitably modified synthetic DNA oligonucleotides. Attachment of acridinium ester label to primary aliphatic amine group(s) present in the synthetic DNA probe molecule is rapid and efficient. Methods have been developed for efficient separation of acridinium ester-labelled DNA from unincorporated labelling reagent and underivatized DNA. The basic hydrogen peroxide detection reaction and photon counting conditions for measurement of chemiluminescence emission from acridinium ester-labelled DNA probes have been optimized. Under optimal conditions, the observed detection limit for the labelled DNA (1:1 mole ratio) is the same as for the free acridinium ester label, which is 2 attomole sensitivity in the best case studied.     

A comparison of chemiluminescence immunoassay and radioimmunoassay for the detection and quantification of methyltestosterone residues in meat samples

A chemiluminescence immunoassay (CLIA) for methyltestosterone is compared with a radioimmunoassay (RIA) employing the same antiserum, raised against methyltestosterone-3-CMO-BSA and using N-(4-aminobutyl)-N-ethylisoluminol conjugate of MT and [1,2-H³]methyltestosterone as tracers. Muscle tissue from slaughtered animals was selected as the matrix. After enzymatic digestion and diethylether extraction only a limited sample clean-up on Lipidex-5000 and on Bond Elut C18 was required. Both methods had similar limits of detection, sensitivities, limits of quantification and speed of analysis (working-load and availability of results).