Abstract:
A method and apparatus for determining a characteristic of a substance is disclosed. The apparatus comprises a light source, a prism, a sensor chip located on the prism, focusing optics located between the light source and the prism, a detector, collimating optics located between the prism and the detector, and calculation means for determining the characteristic of the substance. The sensor chip comprises a metallic film and a transparent substance. The metallic film is operatively arranged to reflect light from the light source. The transparent substance comprises a material having an index of refraction matched to an index of refraction of the prism. The sensor chip is operatively arranged to receive a sample of the substance.

Description:
FIELD OF THE INVENTION  
         [0001]    The present invention relates generally to optical instruments for measuring refractive index of a substance, and more particularly to an optical configuration and method for measuring a refractive index of a sample. The present invention is applicable to surface plasmon resonance (SPR) biosensor devices.  
         BACKGROUND OF THE INVENTION  
         [0002]    The phenomenon of surface plasmon resonance, or SPR, is well known. SPR causes a drop in the intensity of light reflected from the interface of an optically transparent substance and a metal surface at a specific wavelength and angle of incidence. The location of the intensity minimum, measured with respect to wavelength or angle of incidence, changes when differing compositions of substances are placed in a sample space on the metal surface opposite the transparent substance. By measuring the location of the intensity minimum, the identity of the substance in contact with the metal surface may be determined.  
           [0003]    Presently, the accuracy of the measurements is limited due to the spherical aberration of the light as it is incident on both the metallic film and the detector. Spherical aberration of the incident light results in multiple foci within the light beam. The multiple foci present in the beam cause the detector to measure a “hybrid” spectrum, rather than the desired intensity spectrum for an incident beam with a single focus. The hybrid spectrum is a superposition of the intensity spectra for all the foci present in the beam. This hybrid spectrum generally has a drop in the intensity level that is broader than the drop in the intensity spectrum for a single focus. The minimum of a broader drop is more difficult to compute accurately. Thus, the presence of spherical aberration decreases measurement accuracy and repeatability.  
           [0004]    Further, the SPR measurement devices presently available can only measure indices of refraction in a limited range.  
           [0005]    Clearly, then, there is a longfelt need for an SPR analysis apparatus that can reduce spherical aberration in the light beam and allow measurement of indices of refraction in the range of 1.3 to 1.4.  
         SUMMARY OF THE INVENTION  
         [0006]    The present invention comprises a method and apparatus for determining a characteristic of a substance. The apparatus comprises a light source, a prism, a sensor chip located on the prism, focusing optics located between the light source and the prism, a detector, collimating optics located between the prism and the detector, and calculation means for determining the characteristic of the substance. The sensor chip comprises a metallic film and a transparent substance. The metallic film is operatively arranged to reflect light from the light source. The transparent substance comprises a material having an index of refraction matched to an index of refraction of the prism. The sensor chip is operatively arranged to receive a sample of the substance. The focusing optics are operatively arranged to reduce spherical aberration of the light incident on the metallic film. The detector is operatively arranged to measure an intensity of light reflected by the metallic film. The collimating optics are operatively arranged to redirect the light reflected from the metallic film to substantially evenly illuminate the detector.  
           [0007]    A general object of the present invention is to provide a method and apparatus for determining a characteristic of a substance.  
           [0008]    Another object of the present invention is to determine an index of refraction of a substance.  
           [0009]    A further object of the present invention is to determine indices of refraction of substances in the range from 1.3 to 1.4.  
           [0010]    These and other objects, features and advantages of the present invention will become readily apparent to those having ordinary skill in the art upon a reading of the following detailed description of the invention in view of the drawings and claims. 
       
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0011]    The nature and mode of operation of the present invention will now be more fully described in the following detailed description of the invention taken with the accompanying drawing figures, in which:  
         [0012]    [0012]FIG. 1 is an exploded semi-schematic view of a preferred embodiment of the present invention;  
         [0013]    [0013]FIG. 2 is a top view of the prism and sensor chip of a preferred embodiment of the present invention;  
         [0014]    [0014]FIG. 2A is a side view of the prism and sensor chip of a preferred embodiment, taken at plane A-A of FIG. 2;  
         [0015]    [0015]FIG. 2B is a cross sectional view of the prism and sensor chip of a preferred embodiment, taken at plane B-B of FIG. 2;  
         [0016]    [0016]FIG. 3 is a top view of the prism and sensor chip of an alternate embodiment of the present invention;  
         [0017]    [0017]FIG. 4 is a side view of the prism and sensor chip of an alternate embodiment of the present invention;  
         [0018]    [0018]FIG. 5A is a view of light incident on a detector in a configuration wherein the incident light underfills the detector;  
         [0019]    [0019]FIG. 5B is a view of light incident on a detector in a configuration wherein the incident light overfills the detector; and,  
         [0020]    [0020]FIG. 6 is a view of the detector of the present invention wherein the incident light substantially evenly fills the detector. 
     
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT  
       [0021]    It should be appreciated that, in the detailed description of the invention which follows, like reference numbers on different drawing views are intended to identify identical structural elements of the invention in the respective views.  
         [0022]    A preferred embodiment of the present invention is shown in FIG. 1 and generally designated  10 . Apparatus  10  is an SPR analysis apparatus comprising a light source  12 , diffuser  16 , lens  18 , polarizer  19 , filter  20 , lens  21 , aperture  22 , lens  23 , lens  24 , prism  40 , sensor chip  50 , lens  32 , lens  46 , detector  60 , and processing electronics  28 . These elements define an optical axis  11 . Light is emitted by the light source and travels along beam path  14 . The light is focused by lenses  18  and  21  before it passes through aperture  22 . The light continues through lenses  23  and  24  before it enters prism  40 . The light is reflected by metallic film  54  of sensor chip  50 , shown in FIGS. 2A and 2B. The reflection of the light by the metallic film involves an interaction with the electron cloud of the metallic film. At certain wavelengths and angles of incidence, SPR results in the incident light being absorbed by the electrons in the metal, leading to a significant drop in the intensity of the light reflected. The reflected light continues down beam path  14 , through lenses  32  and  46  to detector  60 . In a preferred embodiment, detector  60  is a photodiode array, but it should be readily apparent to one having ordinary skill in the art that other apparatuses for determining the intensity of light are possible, and these modifications are within the scope of the invention as claimed. Thus, the location of the reflection minima may be found by varying the wavelength of the incident light while keeping the angle of incidence constant, or by varying the angle of incidence while keeping the wavelength constant. By comparing the location of the peak to the location of the peak for a substance of known refractive index, the refractive index of an unknown substance may be determined. Further, the presence of a substance in an unknown composition or the identity of an unknown composition may be determined by comparison of the SPR measurement results to the SPR measurement results for a known substance. These processes are well known in the art, and are detailed in U.S. Pat. No. 6,127,183, which is incorporated herein by reference.  
         [0023]    In the present application, “angle of incidence” is intended to mean the angle between the normal to the plane containing the metallic film and the light beam as it approaches the metallic film. In a preferred embodiment, light is emitted at angles of incidence in the range of 64 to 77 degrees. This allows indices of refraction in the range of 1.3 to 1.4 to be measured by the apparatus.  
         [0024]    In a preferred embodiment, diffuser  16  is a plate 20.0 millimeter (mm) in diameter and 1.50 mm thick. The surface through which light enters is ground with an abrasive material and the exit surface is planar. There is a 5.7 mm air gap between light source  12  and diffuser  16 . Lens  18  is an 8 mm in diameter lens made of Schott SK2 glass. The light entry surface is planar and the convex light exit surface has a radius of 6.05 mm. The center thickness is 3.9 mm and the effective focal length is 9.919 mm. There is a 1.0 mm air gap between diffuser  16  and lens  18 . Polarizer  19  is a 2.0 mm thick plate of HN-32 material with a diameter of 20.0 mm. There is a 0.06 mm air gap between lens  18  and polarizer  19 . Filter  20  is a 3.29 mm thick plate. The center wavelength of the passband of the filter is 780 nanometers. There is a 0.44 mm air gap between polarizer  19  and filter  20 . Lens  21  is an 8 mm in diameter lens made of Schott SK2 glass. The convex light entry surface has a radius of 6.05 mm and the light exit surface is planar. The center thickness is 3.9 mm and the effective focal length is 9.919 mm. There is a 3.22 mm air gap between filter  20  and lens  21 . Aperture  22  is a 1 mm in diameter aperture in an opaque substance. There is a 7.4 mm air gap between lens  21  and aperture  22 . Lens  23  is an 8 mm in diameter lens made of Schott SFL 56 glass. The light entry surface is planar and the convex light exit surface has a radius of 10.2 mm. The center thickness is 3.1 mm and the effective focal length is 12.882 mm. There is a 12.5 mm air gap between aperture  22  and lens  23 . Lens  24  is an 8 mm in diameter lens made of Schott SFL 56 glass. The convex light entry surface has a radius of 10.2 mm and the light exit surface is planar. The center thickness is 3.1 mm and the effective focal length is 12.882 mm. There is a 0.5 mm air gap between lens  23  and lens  24 . Prism  40  is trapezoidal in shape with entry face  40 A (See FIG. 1) at a 70.9 degree angle with respect to top face  40 B. Exit face  40 C makes a 90 degree angle with respect to top face  40 B. Top face  40 B and bottom face  40 D are parallel. Side face  40 E and side face  40 F are parallel. Prism  40  is made of Schott BK7 glass. There is a 3.0 mm air gap between prism  40  and lens  24 . Lens  32  is a 22.0 mm in diameter lens made of Schott BK7 glass. The light entry surface is planar and the concave light exit surface has a radius of 39.0 mm. The center thickness is 3.0 mm and the effective focal length is −75.188 mm. There is a 24 mm air gap between prism  40  and lens  32 . Lens  46  is a 31.0 mm in diameter lens made of Schott SK2 glass. The light entry surface is planar and the convex light exit surface has a radius of 30.0 mm. The center thickness is 8.0 mm and the effective focal length is 49.185 mm. There is a 16 mm air gap between lens  32  and lens  46 . There is a 25 mm air gap between lens  46  and detector  60 . However, it should be readily apparent to one skilled in the art that other configurations are possible and these modifications are intended to be within the scope of the invention as claimed.  
         [0025]    Referring now to FIGS. 2, 2A, and  2 B, sensor chip  50  is provided with thin metallic film  54  on an upwardly facing surface thereof. In a preferred embodiment, metallic film  54  includes a layer of chromium approximately ten angstroms thick for adherence to the glass surface of chip  50 , and a gold layer approximately five hundred angstroms thick. In the present embodiment, an optical interface is defined by the contact area of sample  52  with the surface of metallic film  54 . This contact area can be established by dropping the sample  52  onto the surface of metallic film  54 , by using a flow cell designed to bring sample  52  into contact with the surface of metallic film  54 , or by otherwise applying sample  52  to the surface of metallic film  54 .  
         [0026]    Metallic film  54  is optically coupled, indirectly, to prism sample surface  40 B through transparent glass slide  56  and a thin layer of transparent oil  58  provided between the underside of glass slide  56  and sample surface  40 B. As light from illumination source  12  reaches metallic film  54  at the optical interface, certain rays will be incident at a resonance angle determined by the refractive index of sample  52  and energy associated with such rays will be absorbed, while the remainder of the rays will be reflected by metallic film  54 . Beam  13  comprises the rays reflected from the optical interface beneath sample  52 . Of course, metallic film  54  can be optically coupled to sample surface  40 B by applying the film directly to sample surface  40 B, as illustrated in FIG. 3.  
         [0027]    In an alternate embodiment, shown in FIGS. 3 and 4, gasket  70  receives sample  52  such that the optical interface is established. As light from illumination source  12  reaches metallic film  54  at the optical interface, certain rays will be incident at a resonance angle determined by the refractive index of sample  52  and energy associated with such rays will be absorbed, while the remainder of the rays will be reflected by metallic film  54 . Beam  13  comprises the rays reflected from the optical interface beneath sample  52 . Gasket  70  is made of a material such as room temperature vulcanizing (RTV) silicon. It should be readily apparent to one skilled in the art that means for receiving samples other than gaskets are possible, and these modifications are intended to be within the scope of the invention as claimed.  
         [0028]    The indices of refraction of the transparent substance  56  and the prism  40  are matched to minimize reflections at the prism/sensor chip interface and to prevent refraction of the light as it enters the sensor chip. In a preferred embodiment, transparent substance  56  of sensor chip  50  and prism  40  are both made of Schott BK7 glass. However, it should be readily apparent to one having ordinary skill in the art that the sensor chip and the prism may be made of other substances and these modifications are intended to be within the scope of the invention as claimed.  
         [0029]    Lenses  32  and  46  redirect the light reflected by metallic film  54 . There are two primary purposes for the design and configuration of these lenses. The first primary purpose is to control the size of the reflected bundle of light  13 . The size of the bundle of light is optimized to fill the entire photoelement array  27  with the available reflected bundle of light. FIGS. 5A and 5B show the reflected light incident on the detector without having traversed lenses  32  and  46 . If the size of the bundle of light is not controlled, the photoelement array  27  will be illuminated by underfilling incident light  80 , as shown on FIG. 5A, or overfilling incident light  82 , as shown on FIG. 5B. If the light incident on the detector overfills the photoelement array  27 , the measurements involving the extremes of the obtainable refractive index range will not be possible due to the projection of information outside both ends of the photoelement array. If the light incident on the detector underfills the photoelement array  27 , then the measurement resolution will be decreased due to the loss of pixels used on the array. The lens powers and position along optical axis  11  (shown on FIG. 1) determine the size of the bundle of light seen by the photoelement array. By utilizing lenses  32  and  46 , the incident light  84  is such that it fills the entire photoelement array without exceeding the photoelement array boundaries, as shown in FIG. 6. The light  84  substantially evenly illuminates the photodiode array. The precise control on the size of the bundle of light on the photoelement array contributes to the measurement accuracy and precision.  
         [0030]    The second primary purpose of the lens set is to allow for compactness of design. Lens  32  quickly expands the bundle of light to the required size in a short distance. Lens  46  substantially collimates the bundle for presentation to the photoelement array.  
         [0031]    Thus, it is seen that the objects of the present invention are efficiently obtained, although modifications and changes to the invention should be readily apparent to those having ordinary skill in the art, and these modifications are intended to be within the scope of the invention as claimed.