WO2009104123A1

WO2009104123A1 - Light source for ftir biosensor

Info

Publication number: WO2009104123A1
Application number: PCT/IB2009/050617
Authority: WO
Inventors: Dominique M. Bruls; Johannes J. H. B. Schleipen
Original assignee: Koninklijke Philips Electronics N.V.
Priority date: 2008-02-22
Filing date: 2009-02-16
Publication date: 2009-08-27

Abstract

The invention provides a light source for a biosensor. The light source comprises a LED and a corrector means. The corrector means is for correcting the light emitted from the LED by filtering out most of the unwanted angles emitted by the LED. The corrector means comprises one or more holes having a large aspect ratio, i.e., having a depth that is larger than the diameter. The inside of the hole is non-reflecting and light-absorbing in order to avoid unwanted reflections of light inside the hole. The invention further provides a biosensor device comprising the light source according to the invention.

Description

LIGHT SOURCE FOR FTIR BIOSENSOR

FIELD OF THE INVENTION

The invention relates to an improved light source for biosensors and a biosensor including said light source. BACKGROUND OF THE INVENTION The demand for biosensors is increasingly growing these days. Usually, biosensors allow for the detection of a given specific molecule within an analyte, wherein the amount of said molecule is typically small. Therefore, label particles, for example magnetic actuated beads, are used which bind to a specific binding site or spot only, if the molecule to be detected is present within the analyte. One known technique to detect these label particles bound to the binding spots is FTIR. Therein, light is coupled into the sample at an angle of total internal reflection. If no particles are present close to the sample surface, the light is completely reflected. If, however, the label particles are bound to said surface, the condition of total internal reflection is violated, a portion of the light is scattered or absorbed by the beads present at the surface and thus the amount of light reflected by the surface is decreased. By measuring the (decrease of the) intensity of the reflected light with an optical detector, it is possible to estimate the amount of particles bound to the surface.

In a FTIR magnetic biosensor, a geometry may be used which is shown in Figure 1. A laser diode or a light emitting diode (LED) 10 is used as a light source to generate a collimated monochromatic beam of light through an entrance window 15 of a biosensor cartridge 16. Magnetic actuated beads 14 are detected using a total internal reflection principle sampling the surface area 11 of the biosensor cartridge 16 at the position where the light beam emitted by laser diode 10 hits the surface area 11, the light being reflected from the surface 11 is measured using a photo-detector 12. By binding or non-binding of these magnetic beads 14 to the surface in a biological assay, the presence of various substances, e.g., drugs-of abuse, can be detected in the assay, for example in saliva. A magnet 13 is used to actuate the magnetic beads 14.

The use of a laser as light source has the advantage that a narrow collimated beam can be generated to illuminate a small surface on which the bio-assay takes place, thereby illuminating the surface at one distinct angle. The laser further emits light of only one wavelength, which exactly determines the angle at which total internal reflection occurs in the biosensor cartridge. Both the parallelness and monochromaticity of the laser beam result in a better defined evanescent field intensity distribution at the surface. With a laser as light source and using a single or split photo-diode, only one or at most a few (typically 2-4) substances can be tested at the same time. However, in many applications, it is desirable to test more different substances at the same time. In a real product, for example, at least five different types of drugs-of abuse and a reference need to be tested at once, i.e. 6 tests at the same time. Furthermore, it is often required that a test is performed in duplo or triplo, which easily can add up to 20 measuring areas or more.

By printing different spots on the surface 11 of the biosensor cartridge 16, containing different anti-bodies, simultaneous testing for different substances is possible. Multiple species of magnetic beads 14 that have been coated with various anti-bodies can detect the different substances by binding or non-binding to the different regions, i.e., the printed anti-body spots. In order to determine the presence of these different substances in the assay, it is necessary to capture the full reflected FTIR beam and imaging the whole surface 11 of the biosensor, containing the different spots.

Imaging the surface of the printed spots may be done by using a CCD. However, using a CCD, which contains pixels arranged in lines and columns, i.e., in a periodic structure, the temporally and spatially coherent light beam emitted by the laser causes picture artefacts, rendering the obtained images useless. Further distortions of the image may arise due to diffraction and optical interference effects caused by edges and pinholes that are present in the optical setup. Also laser speckle distorts the image. Furthermore, in the cartridge, which generally is made of plastic, small defects may be present, such as small air bubbles or small enclosures of foreign material, giving rise to diffraction effects. Figure 2(a) shows an image of the surface of a biosensor illuminated using a laser beam in combination with a CCD as photo-detector. Many diffraction effects and optical interference effects are visible at all the edges. Furthermore, a large defect due to an imperfection in the plastic of the cartridge is visible. In order to avoid the distortions of the image which mainly relate to the coherence of the laser beam, the coherence of the laser may be destroyed, for example by adding diffusing or moving (e.g. rotating and/or vibrating) optics inside the optical setup. However, these solutions are not practical as they introduce additional parts, for example moving and/or expensive parts, which is not desirable in a product. Also modulating the laser diode at RF frequencies, typically a few hundred MHz, may be used to partially destroy the coherence of the laser. However, experiments show that this is not enough in order to overcome the problems mentioned above.

Another solution may be to replace the laser by a LED. A LED is quasi- monochromatic and therefore well suited for the application in an FTIR biosensor. However, it is not possible to obtain a well collimated beam when using LEDs, in particular when using high power LEDs which are generally used in order to generate enough light, since light from such a LED is not emitted from a point-like source but rather from a light-emitting surface which typically has a size of about 1x1 mm. Therefore, the light rays emitted by the LED will comprise a multitude of angles which typically cover the range of 10-20°. It is however favourable to illuminate the cartridge at its entrance only using one defined angle since otherwise, the intensity of the evanescent field, on which the FTIR method is based, is not well defined anymore. This, in turn, causes a decrease of signal modulation, due to the ill defined evanescent field. Furthermore, due to aberrations caused by imaging a rather large object field using non- parallel illumination, the image may become blurred. An image similar to the image shown in Figure 2(a), but using a LED as a light source, is shown in Figure 2(b). Clearly, a loss of contrast and an overall blurring of the image can be observed.

Furthermore, it is not practicable to use a lens to collimate the light of the LED since, due to the finite dimensions of the light emitting area, no real parallel beam can be obtained and, in addition, an intermediate image of the chip surface of the LED, comprising wires, electrodes etc., will be projected over and superimposed on the obtained image, which is highly undesirable.

SUMMARY OF THE INVENTION There is therefore a need for an improved light source for a biosensor which provides for a constant illumination profile over a surface of the biosensor and which may be used in combination with a CCD for imaging this surface in a two- dimensional way. In particular, the drawbacks of known light sources discussed above should be overcome. The light source for a biosensor according to the invention comprises a

LED and a corrector means for correcting the light intensity distribution of the beam emitted by the LED, and, in particular, for filtering out most of the unwanted emission angles. The corrector means may be a piece of material that is placed in front of the LED, wherein one or a plurality of holes having a large aspect ratio, that is, having a diameter that is made small with respect to the depth of the hole, is arranged in the piece of material. In this way, the piece of material acts substantially as an angle selector which filters out the unwanted angles. The inside of the holes should be opaque, i.e., non- reflecting and light-absorbing. In this way, unwanted reflections of light inside the holes is avoided and a proper angular selection is guaranteed. Using the corrector means, one or several "point-like sources" can be obtained using an LED which provides an almost monochromatic and substantially incoherent light beam.

The light emerging from the holes of the corrector means may further be collimated by using e.g. a collimator lens. In this way, the holes act as quasi-point sources. This is done in such a way that a clear image of the holes can be generated at a certain distance from the corrector means. As the total length of the light path in the biosensor device, i.e. from light source to detector plane, is typically only a fraction of the distance at which an image of the holes is generated, i.e. typically more than 1 meter, a quasi-collimated light beam is generated with an almost uniform intensity distribution, a part of which is being used for illuminating the sensor area. It is further advantageous to choose a LED which already has a light profile that is favorable, i.e. emitting a light beam with a forward propagating intensity distribution as uniform as possible. For example, by using a LED with a so-called bat- wing or Lambertian profile in combination with the corrector means, a homogeneous light distribution can be obtained.

The invention further provides a biosensor device including the light source according to the invention in combination with a CCD for detecting the light reflected from the surface of the biosensor. In such a biosensor device, which may be similar to the one shown in Figure 1 , it is preferred to place an additional diaphragm as close as possible to the entrance window 15 of the biosensor cartridge 16. The size of the diaphragm should be matched to the area which is to be illuminated, in order to suppress the generation of unwanted stray light inside the cartridge 16 and enhance the angular selection of the incoming light beam even further.

These and other aspects of the invention will be apparent from and elucidated with reference to the embodiments described hereafter.

BRIEF DESCRIPTION OF THE DRAWINGS

Fig. 1 schematically shows a FTIR biosensor device;

Fig. 2 shows images taken by a CCD in a biosensor device using (a) laser illumination and (b) LED illumination; Fig. 3 schematically shows a light source according to an embodiment of the present invention;

Fig. 4 shows an image similar to the images of Fig. 2, taken using a light source according to an embodiment of the invention; and

Fig. 5 shows illumination profiles of a light source according to an embodiment of the invention (a) without and (b) with an optimization.

DETAILED DESCRIPTION OF EMBODIMENTS

Fig. 3 schematically shows a light source comprising an LED 1 and a corrector means 2 according to an embodiment of the present invention. In the upper portion of Fig. 3, a top view on the corrector means 2 is shown. The lower portion of Fig. 3 shows a side view on the light source. In the exemplary embodiment shown in Fig. 3, the corrector means 2 includes three holes 3 in the form of elongate pinholes 3 which have a length through the corrector means 2 which is substantially longer than the diameter of the holes 3. Generally, the longer the holes 3 are, the better is the angular selection achieved with the corrector means 2. From a geometric and practical point of view, the ratio of the depth and the diameter should be at least 10 which results in an angular spread which is reduced to a few degrees only.

The inside of the holes should be absorbing and non-reflecting since otherwise, light at extreme angles may still "bounce" through the holes arranged in the corrector means. Furthermore, the diameter of the holes should not be too small since otherwise, almost no light would reach the detector means unless the LED is driven at vary high powers which should be avoided considering power consumption and heat generation. Further reasons why the holes should not have an extremely small diameter is that otherwise large diffraction effects will occur at the edges of the holes, introducing an extra unwanted angular distribution. On the other hand, the diameter of the holes should also not be too large since otherwise, the holes will not act as a quasi-point source anymore and it may happen that the structure of the LED chip is imaged upon the FTIR image leading to image artifacts, shadowing effects etc.

Typically, for the high power LEDs, the diameter of the holes preferably is around 0.3mm. This diameter yields an extra angle due to diffraction at the holes of about +0.15°, which is acceptable. However, the choice of the diameter of the holes also depends on the total length of the light path, and the available optical power, as this introduces losses. On the other hand, the maximum allowable size for the diameter of the holes is lmm, considering LEDs with a light emitting surface of around lxlmm. In a preferred embodiment of the light source, similar to what is shown in

Fig. 3, three holes are provided having a diameter of 0.3mm with a spacing of 0.6mm. This arrangement is optimized for illumination of a surface having a size of 1.5x2mm in a FTIR biosensor. The above solution results in a very compact light source which may be made into a very compact illumination module. The maximum required length of the total illumination unit can be made smaller than 20mm which is to be compared to solutions using a light source like an LED or a laser in combination with collimating optics which typically have a total length of more than 30mm. Reduction of the size of the illumination unit is especially important when integrating the illumination unit in a handheld, compact reader design.

An image of a biosensor surface, similar to the images shown in Fig. 2, but taken using a light source according to the above embodiment of the invention, is shown in Fig. 4.

The number and size of the holes may be varied in order to optimize the illumination profile of the light source. The illumination profile is influenced by using multiple holes but also depends on the spacing between the holes and the diameter of the holes. An example of different illumination profiles achieved and optimized in this way is shown in Fig. 5(a) wherein no optimization has been performed. On the other hand, by tweaking the sizes and the position of the holes, a more homogeneous illumination of the surface of a biosensor can be achieved, as can be seen in Fig. 5(b). In Figs. 5(a) and 5(b), brightness of the illuminated surface is presented by the height of the plot. In order to couple in as much light as possible an additional collimator lens can be used collecting the light emitted by the holes array and making an image of the holes array at large distance, thereby obtaining a quasi-collimated light beam at the position of the sensor area. The collection NA of this lens is governed by the geometry of the holes. If each hole has length L and diameter D, the NAc₀₁₁ of the collimator lens should be typically equal to ~D/L in order to collect as much light as possible from each single hole. The focal length of the collimator lens is governed by the outer diameter of the complete holes array. Let d be the distance from the optical axis to the outermost hole, then the maximum allowed FTIR sensor acceptance angle CC_FTIR imposes the focal length to be F_con~d/cCFτiR. A thorough raytrace analysis is required in order to fully optimize the geometry of the holes array in combination with a specific light intensity distribution from a given LED.

The invention is described with regard to FTIR for optical detection, other optical detection technologies are applicable.

While the invention has been illustrated and described in detail in the drawings and foregoing description, such illustration and description are to be considered illustrative or exemplary and non-restrictive; the invention is thus not limited to the disclosed embodiments. Variations to the disclosed embodiments can be understood and effected by those skilled in the art and practicing the claimed invention, from a study of the drawings, the disclosure, and the appended claims. In the claims, the word "comprising" does not exclude other elements or steps, and the indefinite article "a" or "an" does not exclude a plurality. A single processor or other unit may fulfill the functions of several items recited in the claims. The mere fact that certain measures are recited in mutually different dependent claims does not indicate that a combination of these measures can not be used to advantage. Any reference signs in the claims should not be considered as limiting the scope.

Claims

CLAIMS:

1. A light source for a biosensor comprising

(a) an LED (1); and

(b) a corrector means (2) for correcting light emitted from the LED (1), the corrector means (2) comprising at least one hole (3) having a length that is larger than the diameter of the hole (3), the inside of the hole (3) being non-reflecting and light- absorbing.

2. The light source of claim 1 , wherein the LED (1) is adapted to emit substantially monochromatic light.

3. The light source according to claim 1, wherein the ratio of the length to the diameter of the hole (3) is at least 10.

4. The light source according to claim 1, wherein the hole (3) has a diameter of 0.1 to lmm, preferably 0.3mm.

5. The light source according to claim 1, wherein the corrector means (2) includes at least two holes (3) which are parallel to each other.

6. The light source according to claim 5, wherein the holes (3) are arranged in a plane.

7. The light source according to claim 5, wherein the distance between the holes (3) is larger than their diameter.

8. The light source according to claim 1, further comprising a collimator lens.

9. The light source according to claim 1, wherein the LED (1) emits light having a bat-wing or Lambertian profile.

10. A biosensor device comprising a light source (10) according to claim 1 and a photo-detector (12) for detecting light reflected from a sensor surface (11) of the bio sensor device .

11. The biosensor device according to claim 10, wherein the photo-detector (12) is a CCD.

12. The biosensor device according to claim 10, further comprising a pinhole arranged between the light source (10) and the sensor surface (11).