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Filtration and separation materials for analytical purposes
New Frontiers in Screening for Microbial Biocatalysts Edited by K. Kieslich, C.P. van der Beek, J.A.M. de Bont and W.J.J, van den Tweel © 1998 Elsevier Science B.V. All rights reserved.
77
Filtration and Separation Materials for Analytical Purposes Robert G.Hood, Managing Director, F.S.M.Technologies Ltd. Introduction The ever increasing demand for rapid and accurate screening technology could find solutions in intelligent membrane technology. The removal or attachment of target materials by filtration and separation for analytical purposes offers both rapid qualitative screening and accurate quantitative diagnostic ability with ultra small sample volumes. The Background Although transport phenomena have been studied by biologists and chemists for more than 150 years, the industrial use of membranes in separation process only dates from the introduction of dialysis in the paper industry at the end of the nineteen thirties, followed by desalination by means of electro-dialysis. Over the succeeding years steady progress was made in scientific and technological research that resulted in the development to commercial viability of many membrane-mediated separation processes for a variety of purposes. This presentation is a general introduction to membrane science and technology and the impact it is having in sample separation and process analysis. Membranes First of all let us establish the definition we are using in respect to membranes for analytical purposes. A membrane can be regarded as a phase or a group of phases that controls the transport of substances and/or energy between two essentially uniform phases, which it separates. A wide range of membrane configurations is used with each type having its own unique characteristics. Consequently the choice of the right membrane material and configuration for a specific application can be complex. The following membrane configurations are available for analytical purposes: - Flat sheets - Hollow tubes (tubular) - Hollow fibres - Candles - Spherical liposomes.
78
And The Membrane Structures Porous membranes in general terms have a rigid, pressure resistant structure and behave like a multi-layered sieve with "uniform" mesh holes, the so-called pores. The local flux of any substance in a membrane can be expressed as the product of three factors, all of which are local variables: Flux = Concentration x Mobility x Force
Mobility is the mean net molecular velocity in the force direction, induced by unit force acting on one mole of the permeating substance in the membrane. Concentrations are variable as well, being high in the pores and zero in the matrix material supporting the pores. Mathematically, the expressions for fluxes and forces are at their simplest when edge-and endeffects and the divergence of flows can be neglected. However the exact science of membrane transport is very complex and depends upon a precise understanding of molecular motions and interactions in the membrane phase and on the ways in which these are modified by the presence of the penetrating substances themselves. While a considerable body of knowledge has been built up over a period of time, understanding is still limited to simple systems. In 1981, H.Eber calculated the critical conditions for particle deposition, taking into account the dynamic forces acting on single particles. The requirements of this model have been confirmed by measurement. In 1980, Green & Belfort made use of radial migration of particles and combined the theory of standard filtration. All of these studies showed that a fluid flow parallel to the membrane, as applied in the socalled cross-flow filtration, limits the build-up of materials on the membrane surfaces (gel polarisation). Cross-flow generates shearing forces and/or turbulence across the filter medium and limits the thickness of the "filter cake". In most cases, the formation of a thin layer of retained material on the surface of the membrane cannot be completely eliminated in the steady-state-condition, where the thickness of this layer remains constant. For components which are completely retained, the mass-flow to and from the membrane is well balanced. Classifications of Filtration Techniques The classification of the aforementioned filtration techniques is based on membrane structure and driving force. Based on the actual direction of the fluid flow in relation to the membrane surface one could make a further classification of the various filtration and separation processes:
79 Dead - End Filtration The process fluid is forced through the filter membrane, usually by means of positive pressure. Materials in molecular size greater than the pore sizes of the filter will be retained and solvent together with lower molecular weight materials may be collected as filtrate. This type of filtration is normally restricted to micro-filtration only. Some people call this type of filtration flow-through (flow-thru) as opposed to the following kind of filtration. Fig 1.
Fluid Flow N
Filtrate N
Dead End Filtration
to*ftj*&&rf Cross Section Fig 1 Cross-Flow Filtration Cross-flow filtration is a pressure-driven filtration process. However the fluid flow direction is parallel to the filter membrane surface (tangential filtration) which allows, under well defined circumstances, (flow/pressure/type of membrane) to filter off a certain volume of the process fluid every time the fluid passes the membrane. By doing so, the membrane is kept cleaner for a longer period of time and consequently the filtration process will be more consistent, provided pressure is controlled at all times. Particular influences such as pressure, for example, affect the process parameters and the filtration rate (flux) through the membrane. In addition, the system and plant design and different operating modes, as well as engineering aspects such as hydrodynamics, energy requirements and the selection of construction materials are vital to the performance of cross-flow filtration and separation systems. Fig 2 an example of hollow fibre crossflowfiltration.
80
Filtrate
Fluid Flow ^
Hollow Fibre Cross Flow Filtration
Cross Section Fig 2 Cross-flow filtration can be used in the ultra and micro-filtration range. Operating pressures are normally very low, hence this type of filtration technique is ideal for applications in areas where consistent sampling can take place within especially turbid conditions. The cross-flow filtration is ideal for the separation of suspended microparticles, microorganisms, emulsion droplets and other (bio)materials. Basically there are four operating parameters which influence the performance of cross-flow filtration. a)
Transmembrane Pressure Difference The transmembrane pressure (TMP) difference influences the flux or filtrate rate. Flux increases in a linear fashion to a certain pressure and then, depending on the transport conditions and the fouling on the membrane, will become non-linear. This is one of the reasons why flux increases are not proportional to the transmembrane pressure differences.
b)
Flow-Velocity Parallel to the Membrane The flow velocity parallel to the membrane (tangential Velocity) affects the flowconditions and drag forces on the particle and the high shearing forces on the layer surfaces. A higher velocity results in a constant layer thickness and in the flux remaining constant over a long period of time. In general, the higher the tangential velocity, the higher the flux.
81 c)
Temperature In most cases, the flux increases with temperature on account of lower viscosity of the process fluid. Care must be taken not to exceed certain temperatures affecting filter membrane structures or activities of (bio)materials such as enzymes.
d)
Concentration of Retained Substances The concentration of retained substances in the bulk flow does influence the mass transport to and from the membrane and the flow conditions. Therefore, the influence of concentration in the flux may be complex.
The Cross-Flow Filtration Dynamics Dynamic filtration processes counteract the formation of a covering layer over the active surface of the membrane. This is defined as gelpolarization of fouling. In modules with tubular and capillary membranes, transmembrane pressure can be calculated as follows: Pi + Po Delta TMP= minus Pf 2 The Pf value (pressure on the filtrate side) is usually taken as zero in the case of an "open" system. For diagnostic sampling the fluid dynamics within the system can play a major part in particulate removal. SOME MEMBRANE CATEGORIES FOR SAMPLING PURPOSES Ultra-Filtration Membranes with pore sizes in the nominal range of 1-100 nm are called ultra-filtration membranes, (lnm = 0.001 urn) Most ultra-filtration membranes have an asymmetric structure with fine pores in the surface layer and coarserfinger-likepores below. The materials use are mainly organic and are shaped as hollow-fibres, spiral wound, flat sheet or tubular modules. Inorganic membranes are also available at the upper end of the ultra-filter size range (10-100 nm). Ultra-filtration membranes are normally classified not by pore size but by the largest molecular weight material, which is transmitted. This value is called the Molecular Weight Cut-OfF(M.W.C.O) or Dalton. These cut-off values are nominal values (N.MW.C.O.) and are therefore not always precise enough to be used as an exact parameter when fractioning continuously distributed polymers. However ultra-filtration from nano filtration could have a scale of 100-1 mil Dalton. Fig 3 illustrates a typical example of the membrane range .
82
effective range of FSM Membranes
100
z g iz
FSM membrane zone
HI \Lil
or 0
2k
100k
1mil
Molecular Weight Cut Off (kDalton) log / linear axes Fig 3 Micro-Filtration Micro-filtration membranes are porous structures, which freely pass solvents (usually water) and molecularly dispersed solutes, including polymers. Pore sizes range from 0. lOum up to 1 Oum (1 urn = 0.001 mm). In this size range the separation is mainly a matter of inert sieving of particles, but not entirely so and important interactions occur between the particles and the membrane surface, influenced by the electrical double layers, especially in aqueous systems. These interactions are only poorly understood and yet have major practical consequences for flux decline, membrane life time and the possibility of fractioning small particles by microfiltration. Membrane Filters can be used for final filtration or prefiltration, whereas depth filters, lacking a precisely defined pore size, are used in e.g. clarifying processes, where quantitative retention is not required, or as a prefilter to prolong the life span of a downstream membrane. Most micro-filtration membranes are made from modified, natural or synthetic polymers. Macro-Filtration Macro filtration can be regarded as separation byfibrous,knitted or woven polymeric or metal structured, and ceramic materials. The scale is normally read as from 1.0 urn up.
83
Advantaees of Cross Flow Hollow Fibre Sampling Systems for Analytical Purposes •
Efficient and precise separation of solid materials from liquid carriers, and particulate materials from gaseous carriers.
•
Complete protection of product from process contamination and sensors from harsh environments.
•
Safe handling with secure systems to ensure safe and easy sampling from hazardous systems.
O High quality output offering consistent and accurate membrane separation and unrivalled results. •
A range of filters in a variety of technically advanced materials - areas from less than lsq mm to over lsq m - separation down to less than 3000 Dalton NMWCO) and less than 0.001 micron.
O Standard pumps and fluid controls for the optimisation of a continuous filtration system •
Simple one step syringe containment sampling
•
Access for monitoring and recording equipment for the integrated on-line systems
•
Some Current Uses:
In line analysis for: A) Specialised high quality chemicals and biochemicals B) R & D scientists in biotechnology, medicine or biological scientists C) Manufacturers of analytical instrumentation e.g. Luminometer - Fluorometers HPLC machines D) Process development engineers. Example of Separation and Sensory Systems for Real Time Measurement of; •
Basic Chemical and Physical Parameters
•
Medical/Biotechnology Septicaemia e.g. E.Coli, E.Coli 0157:H7, Staphylococcus Aureus, Amino acids, Carbon Dioxide, Glucose, O2, L-Glutamate, Lactose, Lactate, Oxalate, O2, Sucrose, Uric Acid, Urea etc.
•
Environmental Pesticides, SOx, toluene, benzene, water quality, Mn, Ni, P0 4 , NOx, Si, Ag, Zn etc.
84
And now a Specific Example of a Sample Separation and Analytical Membrane Based System - The Glowgrub™ : Glowgrub™ - How Does it Work? Unlike virtually all other types of membrane based sampling systems, the Glowgrub™ extracts specific materials from the sample flowing tangentially across the membrane. The molecules relative to the molecular cut-off of the membrane material pass through the membrane and are trapped within the device. Therefore the Glowgrub™ is not limited by the turbidity or volume of the sample, for example peptides or endotoxins can be separated in whole blood from 0.1M/L samples at one end of the scale, and E coli samples can be detected in river water from 1000L samples. The relevant diagnostic material can then interact with the sample which can give an accurate quantifiable reaction without blinding by other materials within the sample. Fig 4 illustrates the Glowgrub™ potential for circa 100% viral and bacterial sample entrapment.
0.005 to 0.11 //m
0.15 to 4 0 / ^
Bacteria100
z
o rz
LU \LLI
a: 0.1
1
10
100
1000
10000
Molecular Weight Cut Off (kDalton) log / linear axes Fig 4 Glowgrub™ - The Technology The patented range of optical analytical sampling devices under the Trademark title of Glowgrub™ are designed to capture suspended molecules from ultra small volumes of aqueous solutions (including whole blood). Instantaneous and consistent sample separation and exposure are possible due to the high surface area to volume ratio throughout the range of membranes available for analytical purposes.
85 The Glowgrub™ device is capable of carrying within its membrane interstices, diagnostic materials, either as a coating on the membrane or as a homogeneous external coating, separated from the sample stream by the membrane. The Glowgrub™ technology covers a number of encapsulation techniques which can totally encapsulate the treated membrane in an optically clear coating, with for example all the reactive material present. Voids between the membrane and the outer casing can be created for holding sample processing materials or surface reactive compounds in single or discrete linear sections for one-pass rapid multi-analysis. Fig 5. The environmental uses are considerable, with current samples of the Glowgrub™ being used for river analysis for fish farming, A T P . testing, and septicaemia screening for food processing and preparation.
Fig 5 The Glowgrub™ Device - in Summary: The Glowgrub™ is designed to remove from aqueous solutions, a specific molecular weight item or items within a range of molecular sizes. The design of the Glowgrub™ allows the aqueous solution to pass through a hollow fibre membrane under controlled pressure, and flow, with a specific membrane charge relating to the organism. The controlled conditions within the Glowgrub™ allow the specific molecules under consideration to have a near 100% chance of close contact to the membrane surface. The surface and interstices of the membrane are conditioned by the coating of specific chemiluminescence or bioluminescence, e.g. Fig 6 The reactive materials interact with the molecular subject, Fig 7 for example Septicaemia, E.Coli and/or Staphylococcus Aureus, produce light, and can be read with a standard luminometer or fluorometer for light output increase.
86
BIOLUMINESCENT ASSAY
This CIP assay is based upon the quantitative measurement of a stable level of light produced as a result of an enzyme reaction catalyzyed by firefly luciferase. The formula for the light producing reaction is: Luciferase ATP + Liciferin + 0 2
> Oxyluciferin + AMP + PPi + C0 2 + Light
Fig. 6
THE SAMPLING PROCESS (AN EXAMPLE)
Sample
^
Ttf
Glowgrub Membrane
Organisms
Membrane Cross Section with X ^ ^ J ^ s ^ Immobilised Reactive Material ^Capturing the Sample Organisms
GLOWGRUB1" Light Emission Determined by Organism Quantity in the Sample (Read by Luminometer or Fluorometer) World Patents in Process F.S.M.© 1996
Fig.7
77
Filtration and Separation Materials for Analytical Purposes Robert G.Hood, Managing Director, F.S.M.Technologies Ltd. Introduction The ever increasing demand for rapid and accurate screening technology could find solutions in intelligent membrane technology. The removal or attachment of target materials by filtration and separation for analytical purposes offers both rapid qualitative screening and accurate quantitative diagnostic ability with ultra small sample volumes. The Background Although transport phenomena have been studied by biologists and chemists for more than 150 years, the industrial use of membranes in separation process only dates from the introduction of dialysis in the paper industry at the end of the nineteen thirties, followed by desalination by means of electro-dialysis. Over the succeeding years steady progress was made in scientific and technological research that resulted in the development to commercial viability of many membrane-mediated separation processes for a variety of purposes. This presentation is a general introduction to membrane science and technology and the impact it is having in sample separation and process analysis. Membranes First of all let us establish the definition we are using in respect to membranes for analytical purposes. A membrane can be regarded as a phase or a group of phases that controls the transport of substances and/or energy between two essentially uniform phases, which it separates. A wide range of membrane configurations is used with each type having its own unique characteristics. Consequently the choice of the right membrane material and configuration for a specific application can be complex. The following membrane configurations are available for analytical purposes: - Flat sheets - Hollow tubes (tubular) - Hollow fibres - Candles - Spherical liposomes.
78
And The Membrane Structures Porous membranes in general terms have a rigid, pressure resistant structure and behave like a multi-layered sieve with "uniform" mesh holes, the so-called pores. The local flux of any substance in a membrane can be expressed as the product of three factors, all of which are local variables: Flux = Concentration x Mobility x Force
Mobility is the mean net molecular velocity in the force direction, induced by unit force acting on one mole of the permeating substance in the membrane. Concentrations are variable as well, being high in the pores and zero in the matrix material supporting the pores. Mathematically, the expressions for fluxes and forces are at their simplest when edge-and endeffects and the divergence of flows can be neglected. However the exact science of membrane transport is very complex and depends upon a precise understanding of molecular motions and interactions in the membrane phase and on the ways in which these are modified by the presence of the penetrating substances themselves. While a considerable body of knowledge has been built up over a period of time, understanding is still limited to simple systems. In 1981, H.Eber calculated the critical conditions for particle deposition, taking into account the dynamic forces acting on single particles. The requirements of this model have been confirmed by measurement. In 1980, Green & Belfort made use of radial migration of particles and combined the theory of standard filtration. All of these studies showed that a fluid flow parallel to the membrane, as applied in the socalled cross-flow filtration, limits the build-up of materials on the membrane surfaces (gel polarisation). Cross-flow generates shearing forces and/or turbulence across the filter medium and limits the thickness of the "filter cake". In most cases, the formation of a thin layer of retained material on the surface of the membrane cannot be completely eliminated in the steady-state-condition, where the thickness of this layer remains constant. For components which are completely retained, the mass-flow to and from the membrane is well balanced. Classifications of Filtration Techniques The classification of the aforementioned filtration techniques is based on membrane structure and driving force. Based on the actual direction of the fluid flow in relation to the membrane surface one could make a further classification of the various filtration and separation processes:
79 Dead - End Filtration The process fluid is forced through the filter membrane, usually by means of positive pressure. Materials in molecular size greater than the pore sizes of the filter will be retained and solvent together with lower molecular weight materials may be collected as filtrate. This type of filtration is normally restricted to micro-filtration only. Some people call this type of filtration flow-through (flow-thru) as opposed to the following kind of filtration. Fig 1.
Fluid Flow N
Filtrate N
Dead End Filtration
to*ftj*&&rf Cross Section Fig 1 Cross-Flow Filtration Cross-flow filtration is a pressure-driven filtration process. However the fluid flow direction is parallel to the filter membrane surface (tangential filtration) which allows, under well defined circumstances, (flow/pressure/type of membrane) to filter off a certain volume of the process fluid every time the fluid passes the membrane. By doing so, the membrane is kept cleaner for a longer period of time and consequently the filtration process will be more consistent, provided pressure is controlled at all times. Particular influences such as pressure, for example, affect the process parameters and the filtration rate (flux) through the membrane. In addition, the system and plant design and different operating modes, as well as engineering aspects such as hydrodynamics, energy requirements and the selection of construction materials are vital to the performance of cross-flow filtration and separation systems. Fig 2 an example of hollow fibre crossflowfiltration.
80
Filtrate
Fluid Flow ^
Hollow Fibre Cross Flow Filtration
Cross Section Fig 2 Cross-flow filtration can be used in the ultra and micro-filtration range. Operating pressures are normally very low, hence this type of filtration technique is ideal for applications in areas where consistent sampling can take place within especially turbid conditions. The cross-flow filtration is ideal for the separation of suspended microparticles, microorganisms, emulsion droplets and other (bio)materials. Basically there are four operating parameters which influence the performance of cross-flow filtration. a)
Transmembrane Pressure Difference The transmembrane pressure (TMP) difference influences the flux or filtrate rate. Flux increases in a linear fashion to a certain pressure and then, depending on the transport conditions and the fouling on the membrane, will become non-linear. This is one of the reasons why flux increases are not proportional to the transmembrane pressure differences.
b)
Flow-Velocity Parallel to the Membrane The flow velocity parallel to the membrane (tangential Velocity) affects the flowconditions and drag forces on the particle and the high shearing forces on the layer surfaces. A higher velocity results in a constant layer thickness and in the flux remaining constant over a long period of time. In general, the higher the tangential velocity, the higher the flux.
81 c)
Temperature In most cases, the flux increases with temperature on account of lower viscosity of the process fluid. Care must be taken not to exceed certain temperatures affecting filter membrane structures or activities of (bio)materials such as enzymes.
d)
Concentration of Retained Substances The concentration of retained substances in the bulk flow does influence the mass transport to and from the membrane and the flow conditions. Therefore, the influence of concentration in the flux may be complex.
The Cross-Flow Filtration Dynamics Dynamic filtration processes counteract the formation of a covering layer over the active surface of the membrane. This is defined as gelpolarization of fouling. In modules with tubular and capillary membranes, transmembrane pressure can be calculated as follows: Pi + Po Delta TMP= minus Pf 2 The Pf value (pressure on the filtrate side) is usually taken as zero in the case of an "open" system. For diagnostic sampling the fluid dynamics within the system can play a major part in particulate removal. SOME MEMBRANE CATEGORIES FOR SAMPLING PURPOSES Ultra-Filtration Membranes with pore sizes in the nominal range of 1-100 nm are called ultra-filtration membranes, (lnm = 0.001 urn) Most ultra-filtration membranes have an asymmetric structure with fine pores in the surface layer and coarserfinger-likepores below. The materials use are mainly organic and are shaped as hollow-fibres, spiral wound, flat sheet or tubular modules. Inorganic membranes are also available at the upper end of the ultra-filter size range (10-100 nm). Ultra-filtration membranes are normally classified not by pore size but by the largest molecular weight material, which is transmitted. This value is called the Molecular Weight Cut-OfF(M.W.C.O) or Dalton. These cut-off values are nominal values (N.MW.C.O.) and are therefore not always precise enough to be used as an exact parameter when fractioning continuously distributed polymers. However ultra-filtration from nano filtration could have a scale of 100-1 mil Dalton. Fig 3 illustrates a typical example of the membrane range .
82
effective range of FSM Membranes
100
z g iz
FSM membrane zone
HI \Lil
or 0
2k
100k
1mil
Molecular Weight Cut Off (kDalton) log / linear axes Fig 3 Micro-Filtration Micro-filtration membranes are porous structures, which freely pass solvents (usually water) and molecularly dispersed solutes, including polymers. Pore sizes range from 0. lOum up to 1 Oum (1 urn = 0.001 mm). In this size range the separation is mainly a matter of inert sieving of particles, but not entirely so and important interactions occur between the particles and the membrane surface, influenced by the electrical double layers, especially in aqueous systems. These interactions are only poorly understood and yet have major practical consequences for flux decline, membrane life time and the possibility of fractioning small particles by microfiltration. Membrane Filters can be used for final filtration or prefiltration, whereas depth filters, lacking a precisely defined pore size, are used in e.g. clarifying processes, where quantitative retention is not required, or as a prefilter to prolong the life span of a downstream membrane. Most micro-filtration membranes are made from modified, natural or synthetic polymers. Macro-Filtration Macro filtration can be regarded as separation byfibrous,knitted or woven polymeric or metal structured, and ceramic materials. The scale is normally read as from 1.0 urn up.
83
Advantaees of Cross Flow Hollow Fibre Sampling Systems for Analytical Purposes •
Efficient and precise separation of solid materials from liquid carriers, and particulate materials from gaseous carriers.
•
Complete protection of product from process contamination and sensors from harsh environments.
•
Safe handling with secure systems to ensure safe and easy sampling from hazardous systems.
O High quality output offering consistent and accurate membrane separation and unrivalled results. •
A range of filters in a variety of technically advanced materials - areas from less than lsq mm to over lsq m - separation down to less than 3000 Dalton NMWCO) and less than 0.001 micron.
O Standard pumps and fluid controls for the optimisation of a continuous filtration system •
Simple one step syringe containment sampling
•
Access for monitoring and recording equipment for the integrated on-line systems
•
Some Current Uses:
In line analysis for: A) Specialised high quality chemicals and biochemicals B) R & D scientists in biotechnology, medicine or biological scientists C) Manufacturers of analytical instrumentation e.g. Luminometer - Fluorometers HPLC machines D) Process development engineers. Example of Separation and Sensory Systems for Real Time Measurement of; •
Basic Chemical and Physical Parameters
•
Medical/Biotechnology Septicaemia e.g. E.Coli, E.Coli 0157:H7, Staphylococcus Aureus, Amino acids, Carbon Dioxide, Glucose, O2, L-Glutamate, Lactose, Lactate, Oxalate, O2, Sucrose, Uric Acid, Urea etc.
•
Environmental Pesticides, SOx, toluene, benzene, water quality, Mn, Ni, P0 4 , NOx, Si, Ag, Zn etc.
84
And now a Specific Example of a Sample Separation and Analytical Membrane Based System - The Glowgrub™ : Glowgrub™ - How Does it Work? Unlike virtually all other types of membrane based sampling systems, the Glowgrub™ extracts specific materials from the sample flowing tangentially across the membrane. The molecules relative to the molecular cut-off of the membrane material pass through the membrane and are trapped within the device. Therefore the Glowgrub™ is not limited by the turbidity or volume of the sample, for example peptides or endotoxins can be separated in whole blood from 0.1M/L samples at one end of the scale, and E coli samples can be detected in river water from 1000L samples. The relevant diagnostic material can then interact with the sample which can give an accurate quantifiable reaction without blinding by other materials within the sample. Fig 4 illustrates the Glowgrub™ potential for circa 100% viral and bacterial sample entrapment.
0.005 to 0.11 //m
0.15 to 4 0 / ^
Bacteria100
z
o rz
LU \LLI
a: 0.1
1
10
100
1000
10000
Molecular Weight Cut Off (kDalton) log / linear axes Fig 4 Glowgrub™ - The Technology The patented range of optical analytical sampling devices under the Trademark title of Glowgrub™ are designed to capture suspended molecules from ultra small volumes of aqueous solutions (including whole blood). Instantaneous and consistent sample separation and exposure are possible due to the high surface area to volume ratio throughout the range of membranes available for analytical purposes.
85 The Glowgrub™ device is capable of carrying within its membrane interstices, diagnostic materials, either as a coating on the membrane or as a homogeneous external coating, separated from the sample stream by the membrane. The Glowgrub™ technology covers a number of encapsulation techniques which can totally encapsulate the treated membrane in an optically clear coating, with for example all the reactive material present. Voids between the membrane and the outer casing can be created for holding sample processing materials or surface reactive compounds in single or discrete linear sections for one-pass rapid multi-analysis. Fig 5. The environmental uses are considerable, with current samples of the Glowgrub™ being used for river analysis for fish farming, A T P . testing, and septicaemia screening for food processing and preparation.
Fig 5 The Glowgrub™ Device - in Summary: The Glowgrub™ is designed to remove from aqueous solutions, a specific molecular weight item or items within a range of molecular sizes. The design of the Glowgrub™ allows the aqueous solution to pass through a hollow fibre membrane under controlled pressure, and flow, with a specific membrane charge relating to the organism. The controlled conditions within the Glowgrub™ allow the specific molecules under consideration to have a near 100% chance of close contact to the membrane surface. The surface and interstices of the membrane are conditioned by the coating of specific chemiluminescence or bioluminescence, e.g. Fig 6 The reactive materials interact with the molecular subject, Fig 7 for example Septicaemia, E.Coli and/or Staphylococcus Aureus, produce light, and can be read with a standard luminometer or fluorometer for light output increase.
86
BIOLUMINESCENT ASSAY
This CIP assay is based upon the quantitative measurement of a stable level of light produced as a result of an enzyme reaction catalyzyed by firefly luciferase. The formula for the light producing reaction is: Luciferase ATP + Liciferin + 0 2
> Oxyluciferin + AMP + PPi + C0 2 + Light
Fig. 6
THE SAMPLING PROCESS (AN EXAMPLE)
Sample
^
Ttf
Glowgrub Membrane
Organisms
Membrane Cross Section with X ^ ^ J ^ s ^ Immobilised Reactive Material ^Capturing the Sample Organisms
GLOWGRUB1" Light Emission Determined by Organism Quantity in the Sample (Read by Luminometer or Fluorometer) World Patents in Process F.S.M.© 1996
Fig.7