Bioluminescence- and chemiluminescence- based fiberoptic sensors.
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Animals↗
Biomedical subjects
L J Blum
Publications and source records attributed to L J Blum.
Trends and prospects.
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Biosensing Techniques↗
Design of luminescence photobiosensors.
The potential of immobilized enzyme membranes in biosensors has been explored in our group for several years. Although part of our work has been mainly devoted to electrochemical transducers and oxidases for the design of enzyme electrodes, the demand for ultrasensitive and highly selective sensors led us to consider the use of luminescent enzyme systems associated to optical transduction. When considering the need for operational and reliable biosensors in biotechnology, immobilization and stability of the sensing element still remain, in most cases, an unavoidable problem. We recently proposed a very fast and reliable procedure for preparing enzymatic membranes from Pall (Biodyne Immunoaffinity membranes) supplied in a pre-activated form. Both the firefly and bacterial systems as well as peroxidase for the chemiluminescent determination of various analytes, could be bound to such a support. Based on this approach, a fibre-optic sensor with immobilized enzymes has been designed which permits bio- or chemiluminescent analysis of ATP, NADH or H2O2 respectively. With the NADH-based system, other analytes could be detected using coupled dehydrogenases. This device appears very promising and includes the convenience of both the luminescence sensitivity as well as the handling of the biosensor design.
Adenosine Triphosphate↗
Fibre-optic sensor with co-immobilised bacterial bioluminescence enzymes.
A fibre-optic bioluminescent sensor for the microdetermination of NADH is described. Measurements can be performed in the linear range 1 x 10(-9) M-3 x 10(-6) M with a detection limit of 3 x 10(-10) M using the bacterial luciferase and NAD(P)H:FMN oxidoreductase co-immobilised on a preactivated polyamide membrane. The relative standard deviation was 4.8-5.5% at 4 x 10(-8) M NADH and the steady-state response time was 2 min. When stored at -20 degrees C with 20% glycerol, the activity of the bioactive membranes was higher than that measured prior to freezing and then remained stable for more than four months.
Bacterial Proteins↗
Optically-based chemical and biochemical sensors for the detection of some drugs and biological compounds.
The development of new methods for determining at a very low level a large spectrum of substances affecting the behaviour of living organisms is still a challenging goal. For such a purpose, chemical sensors which can be defined as the intimate combination of a sensitive and specific layer with a transducer, are undoubtedly among the more promising devices. In this field, optical sensors are expanding rapidly, mainly based on absorption, fluorescence, chemi- and bioluminescence. Beside pH and gases, drugs (anticonvulsant, antitumour, anaesthetic...) and other compounds of biological interest can be determined with specifically designed optical sensors, for instance immunosensors. Special attention will be given to optical biosensors with emphasis on chemi- and bioluminescence-based devices which are highly selective and ultrasensitive. When co-immobilizing various auxiliary enzymes in the sensing layer, the potentialities of such devices can be greatly extended as demonstrated by promising results recently obtained in our group.
Biosensing Techniques↗
Theoretical analysis of a translocation-like model with saturable kinetics.
A theoretical analysis of the initial rates of product appearance in both compartments of a specifically designed diffusion cell separated by an asymmetrical enzyme membrane is presented. Variable substrate concentrations and different substrate diffusional limitations were considered. Our analysis shows that, under specific conditions, not only a product accumulation occurs in the compartment opposite to that in which the reaction takes place, but that substrate saturable kinetics can be obtained. These product translocation-like kinetics appear similar to those observed with translocation processes reported for biological situations. For such phenomena, a key role of the diffusion layer surrounding a bioactive surface is proposed.
Biological Transport↗
Vectorial product concentration obtained with a permeable immobilized enzyme membrane. A new approach to the analysis of biological transport systems.
The theoretical analysis of the distribution on both sides of a flat porous membrane of the product generated by an enzyme covalently bound only on to one side of the membrane separating two compartments of widely different volumes is presented. Contrary to what occurs with heterogeneous symmetric systems, the diffusional limitations at the enzyme level play a prominent role, not only on the apparent enzyme activity, but also on product flux-splitting. The mathematical model developed shows that it is possible to concentrate the reaction product in the compartment opposite to that where the reaction occurs. The influence of the parameters and of the physical characteristics of an asymmetrical system on product distribution is analysed. This theoretical analysis is in excellent agreement with experimental data obtained with glucose oxidase immobilized on a porous collagen membrane.
Biological Transport↗
The histological preparation of bronchial brushings--a reliable and satisfactory method.
A method for improving the diagnostic yield of fibreoptic bronchoscopic bronchial brushings and tiny biopsies has been developed. This technique has proved reliable giving a high positive yield of malignant cells and tissue in the 232 specimens examined.
Biopsy↗
Fibre-optic biosensor based on luminescence and immobilized enzymes: microdetermination of sorbitol, ethanol and oxaloacetate.
We have investigated highly selective and ultrasensitive biosensors based on luminescent enzyme systems linked to optical transducers. A fibre-optic sensor with immobilized enzymes was designed; the solid-phase bioreagent was maintained in close contact contact with the tip of a glass fibre bundle connected to the photomultiplier tube of a luminometer. A bacterial luminescence fibre-optic sensor was used for the microdetermination of NADH. Various NAD(P)-dependent enzymes, sorbitol dehydrogenase, alcohol dehydrogenase and malate dehydrogenase, were co-immobilized on preactivated polyamide membranes with the bacterial system and used for the microdetermination of sorbitol, ethanol and oxaloacetate at the nanomolar level with a good precision.
Biosensing Techniques↗