Patent Application: US-44782507-A

Abstract:
the invention relates to a sensor unit for a surface plasmon resonance unit , comprising a transparent sensor structure forming at least one wall of a cavity , the wall being defined by a concave inner surface and a convex outer surface the inner surface is provided with a layer of a conductive material capable of supporting a surface plasmon . in the cavity there is provided a flow structure in said cavity so as to form at least one compartment for sample between the flow structure and the inner wall of the cavity . also , a method for the detection of events at a surface by utilizing surface plasmon resonance is provided . it comprises placing a sample with an analyte of interest in a sensor unit as claimed in claim 1 , and measuring the reflectance from said sensor unit at a single or plurality of angle / angles .

Description:
the surface plasmon is excited by an electromagnetic wave , in the visible region denoted light , which electrical field can be described as : where e is the varying electrical field in time and space , e 0 a field strength constant , k the propagation constant , r the space vector , ω the angular frequency , and t the time . the surface plasmon is excited by the electrical field component parallel to the plane of incidence , i . e . p - polarized light , also denoted transverse magnetic ( tm ). where k o , is the free space propagation constant , n is an effective refractive index , c the speed of light in free space , and λ the wavelength in free space . the propagation constant of a plasmon can be calculated from maxwells equations , and is for the semi - infinite case given by [ 26 ]: where k sp is the ( complex ) surface plasmon propagation constant , n sp is the effective ( complex ) refractive index of the surface plasmon , ε m ( ω ) is the frequency dependent dielectric function ( complex ) of the metal carrying the plasmon , and ε a is the effective dielectric function of the ambient medium ( normally real , but can be frequency dependent and complex ). the effective dielectrical function is monitored ( weighted within the probe depth of the surface plasmon ) by the surface plasmon . the function of the weighting is exponential decaying , and given by : where z is the space coordinate out from the surface , and δ is the characteristic probe depth . the characteristic probe depth is a function of the wavelength and the polarizability of the material , and is at resonance given by : the surface plasmon will propagate along the interface and its intensity will decay exponentially : where x is the coordinate along the interface parallel to the propagation , and l sp , is a characteristic propagation length , which is given by : there are in principal three different ways to measure changes in the spr - propagation constant . first , by measuring the reflected intensity ( reflectance ) at a flank of the spr - dip at a certain wavelength and incident angle . second , by measuring the intensity of the reflected light versus the angle of the incident light ( angular interrogation ). third , by measuring the intensity of reflected light for different wavelengths at a certain incident angle ( wavelength interrogation ). for zero - dimensional spr ( measurement of a single spot ) the intensity measurement requires only a spot ( one element ) detector . a one - dimensional spr measurement ( measurement of a single line ) the intensity measurement requires a one - dimensional ( line ) detector to make an instant measurement of the position of an spr - dip . if a two dimensional ( matrix ) detector is used , a two - dimensional sensor surface can be monitored . the intensity measurement has the disadvantage that only a small part of the spr - dip is measured . this can lead to false interpretations , if , e . g . the dip is broadening or influenced by a drifting offset . if the whole dip is measured as in the angular and wavelength interrogation , these drawbacks are eliminated or at least significantly reduced . if the angular or wavelength interrogation is used , one dimension of the sensor surface is eliminated and replaced by an spr - dip , i . e . a two - dimensional detector can monitor a line , and a one line - detector can monitor a spot . in this case , one dimension is used for the length scale ( real image ) and one dimension is used for the dip ( either angle or wavelength ). however , if the spr - tube / vial is rotated , a fully two - dimensional surface can be monitored by a two - dimensional detector , measuring the whole spr - dip , and a line can be monitored by a one - dimensional detector , using this scanning technique . the preferred detection method for the spr - tube / vial is the angular interrogation . if a collimated ( parallel ) or slightly divergent or convergent light is used , the light beam will hit the outer wall of the tube / vial and refract into the inner surface at a continuum of incident angles due to the width of the beam . the total internal reflected beam will be further refracted at the outer surface , creating a divergent beam , where the spr - dip ( angular ) are easily monitored by a photo - detector or a photographic film . the relation between the incident angle , α , at the outer surface and the incident angle , θ , at the inner surface is given by : where r o and r i are the radii of the outer and inner surfaces respectively , and n 0 is the refractive index for the ambient medium outside the device ( normally air ) and n 1 is the refractive index of the device . from equation 8 it follows that there is only a limited range of incident angles , θ , at the inner surface , depending on the values of r o , r i , n 0 , and n 1 . for the case that the refractive index of the substrate is higher than the ratio between the outer radius and inner radius , it is not possible to obtain high incident angles at the inner surface ( it is assumed that the ambient is air ). if collimated light is used , the width of the beam will lead to a finite measuring spot , where the different positions on the curved surface is associated with a specific incident angle . this means that the position for resonance will change when the resonance condition is changed . the change is dependant on the ratio between the inner and outer radius , but can for most cases be neglected . typically , for a crown glass tube with 12 mm outer diameter and 8 mm inner radius , the movement of the measuring spot is 100 μm for every degree the resonance condition is shifted . this corresponds approximately to 10 μm for 1000 μriu , i . e . the movement is of the same order as the propagation length of the plasmon . this means that for most purposes the movement of the measuring spot is insignificant . the presence of a curved surface on the tube means that the surface plasmon will “ change direction ” along the surface , which may lead to a lower coupling between the plasmon and the incident light . the characteristic propagation length of the plasmon is approximately 2 μm and 20 μm for a wavelength of 633 and 900 nm , respectively , and for a 400 μm diameter capillary tube the deviation of the plasmon at the characteristic propagation length is 0 . 6 and 6 degrees respectively . this means that for the longer wavelength the curvature will probably interfere with the surface plasmon coupling , but for the shorter wavelength the curvature should have little effect . for an 8 mm inner diameter vial the deviation is only 0 . 03 and 0 . 3 degrees respectively , and hence the effect from the curvature is negligible . using other conducting materials with shorter surface plasmon propagation lengths , will lead to lower sensitivity due to broadening of the dip . a preferred inner radius is typically more than 1 mm , and is preferably 3 to 8 mm , and is typically less than 20 mm , preferably less than 10 mm . a useful measurement technique is to perform a ellipsomteric readout , i . e . keep track of the changes in the polarisation state . in other words , the relative phase and amplitude between the reflection coefficients of the p - polarized and the s - polarized light is measured . the s - polarized light is the electrical field component perpendicular to the plane of incidence , also denoted transverse electrical ( te ). the ellipsometric properties are often expressed as the complex ( real and imaginary numbers ) reflection coefficient : i . e . the polarization state can be expressed in the “ ellipsometric angles ” ψ and δ . the polarization state can be measured by both compensating ( null ellipsometry ) and non - compensating ( photometric ) ellipsometric systems . the compensating ellipsometer can use a polarizer - compensator - sample - analyzer ( pcsa ) setup , where the light source is placed before a rotating polarizer followed by a fixed compensator ( usually a ¼ wave plate at 45 °. between the compensator and the analyzator , is the sample placed , consisting of both spr - supporting layer and sample under test . the analyzer , is a polarizer , which is also rotating . the photo detector is placed after the analyzer . the polarizer and analyzer are independently rotated until extinction of occurs . the compensator introduces a phase shift between the p - and s - polarized light . the ellipsometric angles can be found from : where a and p are the azimuths for the analyzer and polarizer , respectively . the compensating ellipsometer is not limited to this setup . non - compensating ellipsometer can be achieved in several ways . one is with fixed azimuths close to extinction and measurement of light intensity at the detector . another is to use a rotating analyzer rae , where the polarizer is fixed and the compensator is optional . by measuring both the amplitude and phase ( with respect to the azimuth of the analyzer ) at the detector , the ellipsometric angles ψ and δ can be determined . the proposed configuration of the device means that it is possible to measure changes in optical parameters with an extremely high accuracy , without using any optical coupling medium , nor using complex optics , making the system very cost effective . it would also be possible to use a divergent beam as proposed by chinowsky et al . [ 11 ], but that will require additional optics . for this case the measuring spot can be focused and it its position will be independent of the effective refractive index of the sample medium . by using a container or a tube , an integrated flow handling system can be placed inside the container , leading to a compact , precise and cost effective solution . the macroscopic shape of the container means that it has a large sensor area , ideal for multi - spot measurements . the possibility of easy rotation of the tube / vial means that the whole sensor area can be scanned extremely fast . a rotation of the tube / vial gives also the opportunity to use the centrifugal force to move samples . a typical flow cell height is preferably between 10 and 100 μm and more preferably between 20 and 50 μm . the shape of the insert makes it suitable for temperature control of the sample . there are at least three reasons for temperature control : 1 ) reaction kinetics is temperature dependent , 2 ) resemble in vivo conditions , e . g . 37 degrees c ., 3 ) baseline of spr - signal is temperature dependent . if a reaction is following the arrhenius rule every 7 ( or 10 ) degrees c . increase in temperature will double reaction rate . in case of water based buffer , spr base line will change approximately − 100 micro refraction units for every degree c . an increase in temperature will decrease the refraction unit due to thermal expansion . the configuration of the invention makes it possible to place thermal control close to the sensor surface and hence a good thermal conductivity can be obtained leading to precise thermal control . one embodiment uses a liquid , e . g . water , water / glycol , etc , to transfer heat / cold to the sample area . the sample heat unit in the insert is placed very close ( typical within 1 mm ) to the roof of the flow cell , making temperature control very fast and precise . temperature controlling liquid is flowing at the outer part of the sample temperature unit leading to very low thermal resistance to the sample . the temperature controlling liquid is preferably heated or chilled by a thermoelectric element . but , other temperature regulating devices such as compressor - condenser , or just tap water , are alternatives . if only heating is required a resistive element ( joule heating ) is preferred . another embodiment with only heating is using resistive heating close to the sample area , without using circulating liquids , and only use thermal conduction to temperate the sample . another embodiment uses two or more temperature controlling liquids , when switched into the sample temperature unit , making fast temperature steps or precise temperature ramps with respect to time . yet another embodiment uses different temperatures at different places on the sample temperature unit , which will create a well defined temperature gradient due to the thermal conductivity of the sample temperature unit . because of the rotational symmetry and macroscopic radius and a relatively short height , the tube is very easy to manufacture with high optical standards and high mechanical precision . the dimensions of the tube / vial make it easy to deposit the essential surface plasmon carrying metal film with a high precision at a low cost . the present invention provides a device and apparatus for measurement of refractive indices close to the inner surface of a tube , cup or vial or a part thereof , based on surface plasmon resonance . the sensing surface may consist of one or a plurality of sensing areas , in axial or radial or diagonal direction or a combination of these directions . the specimen under investigation may be studied in a gaseous , liquid or solid environment . the device can be used for concentration measurement of gases or analytes in a liquid . the device can also be used for analyses of interaction between molecules , e . g . biomolecules . the device can be used for measurement of changes in conformation of a specimen , e . g . coagulation of blood . the device can be used for monitoring of adlayer formations . for the purpose of this invention a “ flow structure ” is taken to mean any structure that cooperates with the inner surface of the sensor unit in such a way as to provide a reduced sample volume in the cavity of the unit . non - limiting examples of a such a flow structure are an insert forming an annular space in a cylindrical sensor unit , or an insert provided with grooves or recesses forming flow channels when the insert rests against the inner surface of the sensor unit . the function of such inserts is to direct the sample , to limit the extension of the sample within the unit , or to adjust the height from sensor surface of e . g . a flow cell , provided by the insert , in order to control the diffusion of sample analyte to the sensor surface , or for controlling the flow rate across the sensor surface . a “ cavity ” shall be taken to mean a space that is surrounded by something , although it need not , but can be completely surrounded . a “ compartment ” shall be taken to mean the space wherein a sample is located , i . e . the space formed in cooperation between an insert and the inner surface of the sensor unit . the invention will now be described , but not limited , with reference to the drawings . fig1 a illustrates one embodiment of the apparatus of the invention wherein a collimated input beam 300 emanates from an illumination system 400 onto a sensor unit 100 , containing a specimen under test 200 , such as a liquid sample or a gas , directed by a structure 610 , and the reflected light 330 is measured by a detection system 500 . the sensor unit 100 has a transparent substrate 103 , typically a plastic material or glass , which is rotationally symmetric or partly rotationally symmetric . the inner surface of the substrate 103 is covered with a thin , typically 50 nm , conducting layer 110 of a free electron like material , typically gold or silver , which can conduct a surface plasmon . the incident light 300 is refracted 310 at the outer surface of the substrate 103 , onto the inner surface and the conducting layer 110 , where the light is either reflected 320 or absorbed in the conducting layer 110 by a surface plasmon , or a combination of both . the reflected light 320 is then refracted at the outer surface of the substrate 103 , forming a light - beam 330 which impinges on a detector system 500 . the optical detection system 500 incorporates a photon sensitive device 510 , that may be a photographic film , but is preferably a photo - detector , e . g . a charge coupled device ( ccd ), charge injection device ( cid ), cmos - device , photo diode etc . for the case of using the photo detector 510 , the optical signal is evaluated by an electronic device 800 , preferably a computer system incorporating an analog to digital converter . the illumination system 400 incorporates at least one light source 410 , but also a plurality is possible , that may be a laser , light emitting diode ( led ), gas discharge lamp ( e . g . xenon ), tungsten halogen lamp etc ., or an array or matrix of the elements . the light source 410 may create a polarized light , e . g . some lasers , or a polarizing element 420 may be inserted anywhere within the light path . the polarization is preferably p - polarised ( transverse magnetic mode ) with regard to the inner surface of the sensor unit 100 for reflectance measurements . the polarizing element 420 can be a dichroic sheet , glan - taylor prism , glan - thompson prism , or an equivalent device . however , the light beam does not necessarily have to be polarized , but that will enhance signal to noise ratio . the light beam 300 is in this configuration expanded by an optical element 430 , e . g . lenses or diffractive optical elements ( doe &# 39 ; s ), or a combination of both , creating a line pattern , making it possible to simultaneously monitor a line ( axial ) at the sensor surface 110 . fig1 b illustrates the same configuration as fig1 a , but with a view from above . fig1 c illustrates an apparatus with imaging optics 520 . the imaging optics 520 will image the sensor surface 110 axially . the imaging optics will collect the angular spectrum 335 from each point on the sensor surface and focus the rays onto the detector 510 . the imaging optics 520 will minimize the effect from diffraction effects due to uneven reflectance distribution along the sensor surface 110 , or by obstruction of the optical wave propagation . the imaging optics 520 preferably contains at least one cylindrical lens or on equivalent doe . fig1 d , illustrates the same configuration as fig1 c , but with a view from above . fig1 e , illustrates the same configuration as fig1 c , but with a cross section of the sensor unit . fig2 , illustrates an apparatus utilizing a convergent beam ( fan shaped beam ), 300 and 310 , to focus on a specific spot at the sensor surface 110 . by using a convergent beam the position of the sensor spot can be small and independent of the resonance condition , i . e . all incident angles impinge on the same spot on the sensor surface 110 . fig3 a , illustrates an apparatus with a collimated or nearly collimated beam 300 from a light source 400 . the light source is preferably a laser or led . the curvature of the substrate 103 will convert the collimated beam 300 to a convergent beam 310 , with respect to the surface normal of the sensor surface 110 . the collimated beam will , due to general physics laws , have a gaussian intensity profile , which is not a disadvantage , but rather an advantage making it possible to just illuminate the angles where the surface plasmon can be excited . the specimen under test 200 is directed by a structure 610 . fig3 b , illustrates the configuration of fig3 a , but seen from a perspective view . fig4 , illustrates the optical path in the sensor unit 100 . the substrate 103 has a refractive index of n 1 and an outer and inner diameter of r o and r i respectively . the ambient medium , typically air , has a refractive index of n 0 . an incident beam 300 impinges on the substrate 103 at an incident angle , α , relative the surface normal . the incident angle , α , is controlled be the height , h , from a centerline , c l , parallel to the beam 300 , by the formula : the beam 300 is refracted into a refracted beam 310 with an angle of β relative the surface normal according to snells law : the refracted beam 310 will impinge on the inner surface of the substrate at an incident angle θ . the angle θ is denoted the resonance angle θ sp at a match of the momenta of the incident light 310 and the surface plasmon . according to optical laws the exit angle θ of the reflected beam 320 is equal to the incident angle θ of the incident beam 310 . due to symmetry , the incident and refracted angles , β and α respectively , at the substrate - ambient interface for the outgoing beam 320 and 330 will be the same as the refracted and incident angle , β and α respectively for the incident beam 300 . the substrate 103 is coated with a conducting layer 110 with refractive index n 2 . the specimen under test 200 is usually a gas or liquid with an effective refractive index of n 3 , which is directed towards the conducting layer 110 by a flow controlling structure 610 . fig5 illustrates the sensor unit 100 with an adhesion layer 105 between the substrate 103 and the conducting layer 110 . the adhesion layer is thin , typically less than 2 nm and preferably 0 . 5 nm , which means that it is thick enough to assure good adhesion and thin enough , not disturbing the surface plasmon . the adhesion layer 105 is typically titanium or chromium . fig6 illustrates the sensor unit 100 with one or a plurality of molecular recognition area / areas 130 a - c . the molecular recognition areas 130 a - c are typically specific to analytes 140 a - c respectively . the molecular recognition layer 130 is typically attached to the conductive layer 110 by the means of a linker layer 120 . the linker layer 120 is typically an alkanethiol with a carbon chain length greater than three and preferably 16 , using the thiol groups to bond to the gold surface . the linker layer 120 may incorporate a hydrogel 210 , as shown in fig7 . the linker layer 120 may incorporate avidin / streptavidin biotin molecules . when the analyte molecules 140 binds to the recognition layer 130 , the dipoles at the analyzing frequency within the probe depth of the surface will normally increase and hence increase the effective refractive index n 3 of the specimen under test 200 . the increase of n 3 will increase the propagation constant of the surface plasmon , i . e . the resonance condition will change . the specimen under test 200 is held between the substrate and the structure 610 . fig7 . illustrates a polymer layer 210 , fibrin etc . the polymer formation can be supervised in conjunction with a rheometer , i . e . the cup or vial is rotating or oscillating , e . g . free oscillating rheometer ( for ). fig8 illustrates a sensor unit 100 with a polymer layer 115 as the recognition layer . the polymer layer 115 is sensitive to the specimen under test 200 , which is either a gas or liquid . the polymer 115 will either be denser or less dense upon contact with the specimen 200 . a denser polymer will increase the effective refractive index n 3 . the polymer film can be made of polyflouroalkylsiloxane , and the gases can be halogenated hydrocarbons such as trichloroethylene , carbon tetrachloride , chloroform , methylene chloride , etc . fig9 a illustrates the sensor unit 100 incorporating slices 100 a - j containing different sensing layers 130 a - j attached preferably to a linker layer 120 , however the linkerlayer can be omitted . fig9 b illustrates the sensor unit 100 incorporating slices 100 a - j incorporating different polymer sensing layers 115 a - j . fig9 c illustrates the sensor unit 100 incorporating slices 100 a - j incorporating different polymer sensing layers 115 a - j , inserted into a tube . fig1 a illustrates a sensor unit 100 , which rotates . the rotation of the sensor unit 100 means that the whole sensing surface 110 can be scanned . a rotating sensor unit 100 means that the surface concentration at the inside of the device is monitored fast for many sensing spots or e . g . gradients . fig1 b illustrates the same set up shown in fig1 a , but with a rotable bob 600 inserted into the cavity of the sensor unit . the rotating bob introduces shear rates in the specimen under test 200 . the bob will also reduce the sample volume of the specimen under test 200 . the bob 600 can also be fixed to reduce the sample volume 200 . fig1 c illustrates the same set up shown in fig1 b , but with a rotating or fixed bob 600 and a rotating or fixed sensor unit 100 . fig1 illustrates a similar setup to fig1 b , but with a conical sensor unit 100 and a conical bob 600 . the conical set up will introduce different shear rates along the axis , which will e . g . affect a polymerization process monitored by surface plasmon resonance . this is particularly interesting for studies of the hemostasis system . the conical sensor unit 100 will obtain a larger dynamic range of share rates if conical shape of the substrate 103 and the bob are different , as illustrated in fig1 . fig1 a illustrates a sensor unit 100 with a fluidic insert 610 to be inserted into the substrate 103 . fig1 b illustrates the sensor unit 100 with the fluidic insert 610 . the fluidic insert 610 has radial channels 615 , which can be used for both immobilized molecules for the recognition layer , or for analyte molecules . the substrate 103 may be conical to assure good sealing to a conical insert 610 , but the substrate 103 can also be cylindrical , utilizing a cylindrical insert 610 . the substrate could be made of an elastic material e . g . silicon rubber , or made of a rigid material with elastic sealings 118 , as illustrated in fig1 c . fig1 c illustrates the fluidic insert 610 with elastic seals 618 , e . g . o - rings . the substrate 103 and the insert 610 can either have a conical or cylindrical shape . fig1 a - b illustrates the sensor unit 100 with a fluidic insert 620 incorporating axial flow channels 625 . the insert 620 may create a cavity 107 which can contain the waste 210 after performed analysis . the waste may be drained by a channel 627 . the flow channels 625 may be fed by individual tubes 626 , that may be filled , e . g . by an external tube , needle or syringe tip . the tube 626 may be sealed by a membrane or an ordinary elastic seal . rotation of either the units 103 and 610 or external tubes may act as valves , changing flow cells . fig1 illustrates calculation of the reflectance , i . e . the ratio between the incident light 310 at the sensing surface 110 and the reflected light 320 from the sensing surface 110 versus the incident angle at the inner surface of the substrate 103 , for three different wavelengths , 633 nm , 733 nm , and 833 nm respectively , and two different effective refractive index , n 3 , ( n 3 is equal to 1 . 330 and 1 . 333 respectively ) of the specimen under test 200 . the conducting layer 110 is in this case gold with a thickness of 50 nm . the substrate 103 is in this case bk7 glass , with a refractive index n 1 of approximately 1 . 5 . the position of the spr - curves is a function of the effective refractive index n 3 seen by the surface plasmon . fig1 illustrates simulations of the reflected intensity at a detector 510 shown in fig3 , at a wavelength of 633 nm , where the substrate 103 is made of bk7 glass , and a 50 nm thick conduction layer 110 made of gold is applied onto the inside of the substrate 103 . the refractive indices n 3 of the specimen under test 200 are 1 . 33 and 1 . 34 respectively . the fall off of the intensity at the border of the detector 510 is an effect of the gaussian shape of the input beam 300 . the irregular shape of the curves emanates from photon shot noise . the incident angle , θ , at the inner surface of the substrate 103 depends on the incident angle , α , at the outer surface , the refractive indices of the ambient environment and the substrate , n 0 and n 1 respectively , and the ratio between the radii of the inner and outer surfaces , r i and r o respectively , according to equation 8 . fig1 illustrates the incident angle , α , at the outer surface of the substrate 103 as a function of the incident angle , θ , at the inner surface , at different inner and outer radii ratios , 1 . 2 , 1 . 33 , 1 . 5 , 1 . 71 , and 2 , for a substrate 103 made of schott duran glass in air . if the ratios are greater than the ratio between the refractive indices of the substrate 103 and the ambient , it is not possible to obtain all incident angles , θ , at the inner surface . for a collimated input beam 300 with a physical width , there are a diversity of incident angles , θ , along the inner surface of the substrate 103 , which means that the resonance angle θ sp will occur at different places along the inner surface . the movement of the measuring spot as a function of the incident angle ( e . g . resonance angle ) at the inner surface of the substrate 103 is given in fig1 for different values of the inner diameter , r i , ( 6 , 7 , 8 , 9 , and 10 mm ) at an outer diameter of 12 mm of the substrate made of schott duran glass . using the approximation of a change of 10 − 6 refractive index units corresponds to 10 − 4 degrees in resonance shift of θ sp , leads to a sensing spot movement as illustrated in fig1 . the conditions are the same as for fig1 . fig1 illustrates the mismatch between the surface plasmon wave vector , k sp , and the parallel component of the exciting light at the start k sp ( 0 ) and at the propagation length k sp ( l sp ) of the surface plasmon , due to the curvature of the substrate 103 . the angular mismatch ( in radians ) at the characteristic propagation length is given by ξ . where ξ is given by the formula : fig2 a - c illustrates how the sensor unit 100 can be closed in one or both ends by means of a cap 190 . fig2 illustrates a sensor unit 110 placed in a holder 900 ensuring exact positions . fig2 a , illustrates the signal from the detector 500 , seen on the computer 800 from one embodiment of the surface plasmon resonance apparatus of the invention as illustrated in fig3 a - b . the embodiment utilizes a monochromatic light source comprising a 5 mw hene laser ( melles - griot ) 400 , emitting coherent and p - polarized ( with respect to the sensor surface ) light 300 at a wavelength of 633 nm . the light 300 impinges on the outer surface of the substrate 103 made of duran ( schott ) glass with a refractive index of 1 . 5 , and with outer and inner diameters of 15 mm and 10 mm respectively . the glass substrate is sputtered with approximately 0 . 5 nm chromium acting as the adhesion layer 105 for the surface plasmon supporting metal layer 110 . the surface plasmon supporting metal layer 110 is sputtered to a thickness of 50 nm . both metal layers 105 and 110 are deposited using physical vapor deposition techniques ( pvd ). sample 200 consists of sucrose solutions ranging from zero to 5 % w / w surcrose in deionized water ( millipore ) in steps of 1 %. the sucrose solutions 200 get the following refractive indices : 1 . 3330 , 1 . 3337 , 1 . 3359 , 1 . 3379 , 1 . 3388 , and 1 . 3403 . the reflected light pattern 330 from the sensor surface impinges on the ccd camera 500 ( orbis 2 , spectra source inc .). the computer 800 is an ordinary pc . fig2 b , illustrates the signal from the detector 500 , with noise reduction by the computer 800 for two refractive indices ( 1 . 33 and 1 . 34 ) of the sample 200 . fig2 a , illustrates an exploded view of the sensor unit 100 as an “ angular cut out ” of the substrate 103 , here as a half pipe . at least one , but not limited to one , insert 610 is placed in the cavity , either in direct contact with the surface using flow channels in the insert 610 , or in the vicinity of the surface with or without a structure to control the flow of gas or fluid used . the use of an “ angular cut ” substrate means that formation of the adhesion layer 105 as shown in fig5 and conductive layer 110 is easily formed by traditional physical vapor deposition ( pvd ) techniques , such as evaporation , electronic beam , and sputtering . moreover , angular break up of the structure means that there will be an open large cavity which will help formation of the molecular recognition layers 130 as shown in fig6 , with a plurality of techniques , as soft lithography stamps etc . fig2 b , illustrates an exploded view of the sensor unit 100 as assembled of two , but not limited to that number , substrates 103 and 104 forming a large cavity , in which a flow controlling part 610 is inserted , either in contact with the inner surface or in the vicinity . the flow controlling part has preferably integrated flow channels when in contact with the curved surface . the setup is not limited to that for the case when the insert 610 is in contact with the surface . for the non contacting case the insert 610 will preferably rotate or oscillate in the large cavity formed by the substrates 103 and 104 , or vice versa when the substrates 103 and 104 together are moving , or both when both insert 610 and both substrates 103 and 104 is moving . fig2 , illustrates a sensor unit 100 with a double curved surface 107 , e . g . the bottom of a vial , with a surface plasmon supporting conductive layer 110 , where the surface plasmon can be excited in many directions . fig2 , illustrates a sensor unit 100 with an integrated flow structure 611 into the optical substrate 103 . the cavity that is formed between the substrate 103 and the flow structure 611 is filled with a sample 200 . the surface plasmon supporting conductive layer 110 is present at the concave surface of the wall . fig2 a , illustrates a sensor unit 100 with a flow structure 610 inserted in the optical substrate 103 . the cavity that is formed between the substrate 103 and the flow structure 610 is filled with a sample 200 . the molecular recognition sites 130 are placed on the surface plasmon supporting conductive layer 110 which is present at the concave surface of the wall of the substrate 103 . the flow structure incorporates a temperature control unit 700 the purpose of which is to keep sample at sensor surface at a controlled temperature . temperature can either be held constant or ramped . the thermo control unit consists basically of two parts , one sample temperature unit 710 which performs the local heat exchange at the sensor surface , and a heat / cold supplying unit 720 , which supplies the sample temperature unit with heat or cold . the sample temperature unit 710 is in a simple embodiment , typically a piece of metal , and in a more advanced embodiment an insert made of e . g . polymers , ceramics , metal or a combination of these materials with channels for liquids carrying the heat flux . the heat / cold supplying unit is typical a thermoelectrical device ( e . g . peltier unit ), but can also be a conventional compressor with heater / condenser or for the case only heating is necessary , a resistive element . using a resistive element , the unit can be integrated with the sample temperature unit 710 , forming a single unit . to obtain fast temperature ramps , different temperatures can be provided simultaneously at the heat / cold supplying unit 720 , and these temperature stores can be switched into the sample control unit 710 . fig2 b , illustrates a temperature control unit 700 , where the sample temperature unit 710 and heat / cold supply unit 720 is separated by tubes , inlet 733 and outlet 736 carrying the liquid for thermal control . fig2 c , illustrates a temperature control unit 700 , where the sample temperature unit 710 has two ( or more ) separated temperature zones 723 and 726 , creating a temperature gradient between them . fig2 a , shows the sample temperature unit 710 inserted into the substrate 103 , with the temperature controlling liquid 732 close to the sample 200 , therefore obtaining fast and precise temperature control of the sample . fig2 , shows an ellipsomeric setup , i . e . measurement of the polarisation state . the light source 420 , which can be monochromatic or multispectral , provides both p - and s - polarized light . this basic setup can be used for compensated as well as uncompensated ellipsometry . the uncompensated ellipsometer is shown as a pcsa - setup . for this case the compensator 460 , which is a quarter wave plate , is set to 45 ° to introduce a phase shift between the p - and s - polarized light . both the polarizer 450 and analyzer 550 are rotated until the light at the detector 510 vanishes . the setup can also be used off - null , where both the analyzer 550 and polariser 450 are fixed at azimuths close to zero light at the detector 510 . a rotating analyser ellipsometer , rae , is obtained for a fixed polarizer 450 , usually at 45 °, and a rotating analyzer 550 . for this case the compensator 460 is optional . the light source 410 and detector 510 can be used in imaging . 1 . nylander , c ., b . liedberg , and t . lind , gas detection by means of surface plasmon resonance , sensors and actuators , 1983 . p . 79 - 88 . 2 . abdelghani , a ., et al ., surface plasmon resonance fiber - optic sensor for gas detection . sensors and actuators b - chemical , 1997 . b38 - 39 : p . 407 - 410 . 3 . tengvall , p . and i . lundström , determination of polymerization / coagulation in a fluid . 1998 , global hemostasis inst mgr ab ( se ); tengvall pentti ( se ); lundstroem ingemar ( se ). 4 . vikinge , t . p ., et al . blood plasma coagulation studied by surface plasmon resonance . in bios &# 39 ; 98 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