Patent Description:
The surface plasmon sensing system is usually applied to detect the target molecules. When the target biomolecule interacts with the receptor molecule on the plasma sensor, the surface refractive index thereof can be changed, and the change can be measured based on the angle or frequency (wavelength) of the surface plasmon resonance mode, which is usually at the dips of intensity-angle and intensity-wavelength. By monitoring the signal in real-time, the measuring figure of the signal variations upon time can be obtained, which can be used to further analyze the affinity between the target molecule and the receptor. The surface plasmon resonance imaging method is capable of real-time, label-free and high-throughput detection for many sensing applications, so this method is usually used in the biological detection field.

With the development of the biological or pharmaceutical field, the demand for high-throughput detection is rapidly increasing, such as biomarker exploration, clinical detection and ligand display. However, the development of high-throughput surface plasmon systems is hindered because of the integration of the imaging system and the angle or wavelength analysis device is extremely difficult. For example, the frequency domain sensitive detection method uses a broadband light beam to illuminate a surface plasmon sensor, and the reflected or transmitted light is split by a prism or a grating. Then, the intensity of each frequency domain is analyzed by a one-dimensional or two-dimensional photodetector, and the surface plasmon signal response can be determined by tracking the dips or peaks on the spectrum. The angle sensitive detection method uses a single-frequency light source, which is incident to the surface plasmon sensor by different wavelengths, and the intensity of each angle is analyzed by the one-dimensional or two-dimensional photodetector. Thus, the surface plasmon signal response can be determined by tracking the dips or peaks of the light intensities in different angles. These methods can only measure the signal of a single point on the surface plasma sensor. If it is necessary to measure the signals of multiple points, an additional position scanning device must be added, which not only affects the complexity of the entire surface plasmon system, but also increases the measuring time (additional scanning time) that can limit the temporal resolution. Thus, a system and method for surface plasmon imaging is quite desired.

The first system combining surface plasmon sensing and imaging is disclosed by Yeatman in <NUM>. In this system, a linear scanning focused light beam is provided to excite surface plasmon waves. A dielectric film (with a thickness of <NUM>) is imaged through a camera by measuring the intensity of the reflected light in these scanned areas. A year later, Rothenhäusler demonstrated a surface plasmon imaging microscopy that does not require the scanning step. An expanded laser beam is used to excite surface plasma waves, which are directly imaged by the camera, and the refractive indexes can be reflected in the intensities within the image. There are many devices implemented based on this method, such as SPRimager® (GWC Technology, referred to <CIT> <CIT>). Another method of surface plasmon imaging is to add a scanning device to the system. Bengt Ivarsson et al. proposed a method for examining thin layer structures on a (sensor) surface (see <CIT>), wherein two detectors are used to detect surface plasmon signals in different areas. Lutz Hoppe et al. used a self-made mask in the surface plasmon system (see <CIT>), wherein the incident light was projected through the mask onto different positions of the plasmon sensor on the surface of prism for providing multi-point surface plasmon sensing. Carsten Thirstrup, in <CIT>, disclosed a complete surface plasmon imaging system in which a reflective diffractive optical element is used to generate linear light focusing to different areas on the surface plasmon sensor, thereby generating the surface plasmon image. Weibel proposed a surface plasmon imaging device (see <CIT>) that illuminates a surface plasma sensor with a single-frequency light source and uses double lenses to scan the light angle, thereby generating the surface plasmon image based on the angle-related surface plasmon signals. In addition to the angle-related and frequency-related methods, the surface plasmon signals can also be imaged by using the phase difference method. <CIT>, proposed a surface plasmon imaging apparatus based on Mach-Zehnder interferometry. In this patent, the s-wave is used as a reference signal, the p-wave is used to detect the refractive index, and an image with a spatial frequency can be presented by combining the s-wave and the p-wave.

As mentioned above, there are three types of surface plasmon imaging technologies including the wavelength/angle/position scanning type, single-frequency type, and interference type. However, the scanning type surface plasmon imaging technology needs additional scanning time, which may cause poor temporal resolution and thus is not suitable for detecting the high affinity molecule reaction. The single-frequency type surface plasmon imaging technology is limited by the detection means of light intensity differences. Since the light intensity is easily interfered by multiple noises, including thermal disturbances and vibrations, the detection limitation of the single-frequency type surface plasmon imaging technology is worse than the angle or wavelength methods. The interference type surface plasmon imaging technology provides the best detection limitation, but the optical path system thereof is more complex. Thus, a very stable optical environment is required, and the detection dynamic region is narrower, thereby limiting the target molecular size for measurement. <NPL>, disclose a surface plasmon imaging using two light sources with different wavelengths. <NPL>, teach generation of a 1D surface plasmon signal image using a light source and a diffraction grating. <NPL>, disclose generation of a one-dimensional surface plasmon signal image using a single light source.

The present invention provides an apparatus for surface plasmon resonance imaging according to claim <NUM> or <NUM>. Further developments of the invention are defined in the dependent claims. Any embodiments and examples of the description not falling within the scope of the claims do not form part of the invention and are provided for illustrative purposes only.

This disclosure provides a method and apparatus for surface plasmon imaging without dispersive components, which are based on spectral contrast and can be applied to any surface plasmon sensors (e.g. the extraordinary transmission-type, prism-type or local-type surface plasmon resonance sensors). The images of the sensors are sensed in two different spectral bandwidths for presenting the surface plasmon signal images. The advantages of this disclosure comprise: (<NUM>) capable of measuring the extraordinary transmission-type, prism-type, and local-type surface plasmon resonance sensors for providing the surface plasmon signal images; (<NUM>) unnecessary to install the dispersive component and scanning mechanism, so that the system is simpler and the temporal resolution of the obtained data can be increased; (<NUM>) providing wider detection dynamic region of the surface plasmon signals, which is suitable for measuring biomolecules, by comparing the images of two spectral bandwidths; (<NUM>) reducing the intensity noises by comparing the images of two spectral bandwidths so as to increase the detection limit; and (<NUM>) suitable for various imaging systems such as scanner, microscope or smart phone.

The present disclosure provides an apparatus for surface plasmon resonance imaging, which comprises a surface plasmon resonance sensing chip, an imaging module and an image processing unit. The surface plasmon resonance sensing chip has a surface plasmon resonance wavelength. The imaging module is configured to receive a first bandwidth plasmon resonance light beam and a second bandwidth plasmon resonance light beam for generating a first bandwidth surface plasmon image signal and a second bandwidth surface plasmon image signal, respectively. The image processing unit is coupled to the imaging module and receives the first bandwidth surface plasmon image signal and the second bandwidth surface plasmon image signal for generating a surface plasmon signal image. The surface plasmon resonance wavelength is less than a minimum wavelength of the first bandwidth plasmon resonance light beam and greater than a maximum wavelength of the second bandwidth plasmon resonance light beam. A difference between the surface plasmon resonance wavelength and the minimum wavelength of the first bandwidth plasmon resonance light beam is less than <NUM>, and a difference between the surface plasmon resonance wavelength and the maximum wavelength of the second bandwidth plasmon resonance light beam is less than <NUM>.

In one embodiment, the first bandwidth plasmon resonance light beam and the second bandwidth plasmon resonance light beam can be both reflected light beams, or they can be both transmitted light beams.

In one embodiment, the surface plasmon resonance wavelength of the surface plasmon resonance sensing chip changes along with a surface environmental refractive index.

In one embodiment, the surface plasmon resonance sensing chip comprises a periodic metallic nanostructure.

In one embodiment, the surface plasmon resonance sensing chip is a transmission-type surface plasmon resonance sensing chip or a reflection-type surface plasmon resonance sensing chip.

In one embodiment, the surface plasmon signal image is obtained by the image processing unit according to an equation of: <MAT>.

In an example outside the scope of the appended claims, instead of a single light source module for emitting a broadband beam, the apparatus may comprise a light source module for emitting a first bandwidth incident light beam and a second bandwidth incident light beam, wherein the surface plasmon resonance sensing chip is configured to receive the first bandwidth incident light beam and the second bandwidth incident light beam for correspondingly generating the first bandwidth plasmon resonance light beam and the second bandwidth plasmon resonance light beam, respectively.

In an example outside the scope of the appended claims, instead of a single light source module for emitting a broadband beam, the light source module comprises a first bandwidth light emitting unit, a second bandwidth light emitting unit, and a switch unit. The first bandwidth light emitting unit emits the first bandwidth incident light beam, and the second bandwidth light emitting unit emits the second bandwidth incident light beam. The switch unit couples to the first bandwidth light emitting unit and the second bandwidth light emitting unit for switching between the first bandwidth light emitting unit and the second bandwidth light emitting unit.

The apparatus further comprises a spectral modulation module comprising a first bandwidth filter and a second bandwidth filter. The spectral modulation module receives a broadband light beam to generate a first bandwidth incident light beam and a second bandwidth incident light beam, and the surface plasmon resonance sensing chip is configured to receive the first bandwidth incident light beam and the second bandwidth incident light beam for correspondingly generating the first bandwidth plasmon resonance light beam and the second bandwidth plasmon resonance light beam, respectively.

In one embodiment, the spectral modulation module further comprises a switch unit coupling to the first bandwidth filter and the second bandwidth filter for switching the first bandwidth filter and the second bandwidth filter in an incident path from the broadband light source to the surface plasmon resonance sensing chip.

In one embodiment, the broadband light beam is generated by a white light source.

In one embodiment, the white light source comprises a white light LED, a halogen lamp, a tungsten lamp, or a xenon lamp.

In one embodiment, the imaging module comprises an imaging unit and a lens, the lens is located between the surface plasmon resonance sensing chip and the imaging unit, and the first bandwidth plasmon resonance light beam and the second bandwidth plasmon resonance light beam pass through the lens and are then projected on the imaging unit.

In one embodiment, the surface plasmon resonance sensing chip is configured to receive a broadband light beam for generating a plasmon resonance light beam, and the apparatus further comprises a spectral modulation module comprising a first bandwidth filter and a second bandwidth filter. The spectral modulation module receives the plasmon resonance light beam to generate the first bandwidth plasmon resonance light beam and the second bandwidth plasmon resonance light beam. The surface plasmon resonance wavelength of the surface plasmon resonance sensing chip is less than a minimum cutoff wavelength of the first bandwidth filter and greater than a maximum cutoff wavelength of the second bandwidth filter. A difference between the surface plasmon resonance wavelength and the minimum cutoff wavelength of the first bandwidth filter is less than <NUM>, and a difference between the surface plasmon resonance wavelength and the maximum cutoff wavelength of the second bandwidth filter is less than <NUM>.

In one embodiment, the spectral modulation module further comprises a switch unit coupling to the first bandwidth filter and the second bandwidth filter for switching the first bandwidth filter and the second bandwidth filter in a light path between the surface plasmon resonance sensing chip and the imaging module.

In one embodiment, the spectral modulation module further comprises a light splitting module configured to receive the plasmon resonance light beam for generating a first split light beam and a second split light beam. The first bandwidth filter is configured to receive the first split light beam for generating the first bandwidth plasmon resonance light beam, and the second bandwidth filter is configured to receive the second split light beam for generating the second bandwidth plasmon resonance light beam.

In one embodiment, the light splitting module comprises a light splitter for generating the first split light beam and the second split light beam.

In one embodiment, the light splitting module comprises a light splitter and a reflector, and the reflector is located between the light splitter and the first bandwidth filter or between the light splitter and the second bandwidth filter.

In one embodiment, the imaging module comprises an imaging unit and a lens, the lens is located between the first bandwidth filter, the second bandwidth filter and the imaging unit, and the first bandwidth plasmon resonance light beam and the second bandwidth plasmon resonance light beam pass through the lens and are then projected on the imaging unit.

In one embodiment, the imaging module comprises a first imaging submodule and a second imaging submodule. The first imaging submodule comprises a first lens and a first imaging unit, the first lens is located between the first bandwidth filter and the first imaging unit, and the first bandwidth plasmon resonance light beam passes through the first lens and is then projected on the first imaging unit. The second imaging submodule comprises a second lens and a second imaging unit, the second lens is located between the second bandwidth filter and the second imaging unit, and the second bandwidth plasmon resonance light beam passes through the second lens and is then projected on the second imaging unit.

In one embodiment, the plasmon resonance light beam is a reflected light beam or a transmitted light beam.

Outside the scope of the appended claims, the present disclosure also provides a method for surface plasmon resonance imaging, comprising steps of: receiving a first bandwidth plasmon resonance light beam and a second bandwidth plasmon resonance light beam from a surface plasmon resonance sensing chip by an imaging module for generating a first bandwidth surface plasmon image signal and a second bandwidth surface plasmon image signal, respectively; wherein the surface plasmon resonance sensing chip has a surface plasmon resonance wavelength, the surface plasmon resonance wavelength is less than a minimum wavelength of the first bandwidth plasmon resonance light beam and greater than a maximum wavelength of the second bandwidth plasmon resonance light beam, a difference between the surface plasmon resonance wavelength and the minimum wavelength of the first bandwidth plasmon resonance light beam is less than <NUM>, and a difference between the surface plasmon resonance wavelength and the maximum wavelength of the second bandwidth plasmon resonance light beam is less than <NUM>; and receiving the first bandwidth surface plasmon image signal and the second bandwidth surface plasmon image signal by an image processing unit for generating a surface plasmon signal image, wherein the image processing unit is coupled to the imaging module.

A difference between the surface plasmon resonance wavelength and the minimum wavelength of the first bandwidth plasmon resonance light beam is less than <NUM>.

A difference between the surface plasmon resonance wavelength and the maximum wavelength of the second bandwidth plasmon resonance light beam is less than <NUM>.

This disclosure provides a new method and apparatus for surface plasmon imaging. <FIG> introduces the working theory of the method of this disclosure. On the metal surface with surface plasmon resonance, the transmitted or reflected spectrum thereof has a resonance peak or dip, and the center position of the peak or dip is λSPR (see <FIG>). The above-mentioned surface plasmon sensor is irradiated by a light beam, and the light beam can pass through two designed bandpass filters. The two filters can divide the transmitted peak of the surface plasmon sensor into a short-wavelength portion and a long-wavelength portion. In order to determine the two-dimensional surface plasmon signal, the light beam passing the long bandwidth filter is imaged to form a long bandwidth surface plasmon image (image Aij), wherein i and j are the corresponding pixels in the x and y directions. In addition, the light beam passing the short bandwidth filter is imaged to form a short bandwidth surface plasmon image (image Bij). The images can be calculated to provide the surface plasmon signal images. Through the custom equation of (Aij-Bij)/(Aij+Bij), a self-reference image can be obtained, wherein Aij and Bij represent the light intensities of the image pixel ij. The custom equation is usually applied to double sensors or quaternary position sensors for sensing the position of light beam. If the light beam contains noises, the equation can be modified as: <MAT>.

Since A~B»dI, the affection of the noise to the signal can be sufficiently decreased.

In this disclosure, the dynamic regions are evaluated by numerical method. <FIG> indicate that the positions of transmitted peaks can be shifted as the change of refractive index during the sensing procedure. <FIG> shows the blue shift surface plasmon spectrum signal, which represents the molecular dissociation event detected by the surface plasmon sensor. The contrast signal is defined as γ= (A - B) / (A + B). Compared with the wavelength shift, it is well known that the wavelength shift method can provide extremely wide dynamic region. <FIG> show the relationship of the contrast signal and the wavelength shift. The examples of red shift and blue shift both have linear relationship within the wavelength shift range from -<NUM> to +<NUM>. The dynamic region corresponds to the refractive index range from -<NUM> to +<NUM>. In other words, this method is suitable for the interaction of most biomolecules.

Referring to <FIG>, to implement the above method, an optical setup includes an illumination system <NUM>, two narrow-band filters <NUM> and <NUM>, a sensing unit <NUM> (including a surface plasmon resonance sensing chip), and a monochrome imaging device <NUM>. The sensing unit <NUM> is composed of a fluidic device <NUM> and a surface plasmon sensor <NUM>. The surface plasmon sensor <NUM> can be an abnormal transmission-type surface plasmon sensor or a localized surface plasmon resonance sensor. In this embodiment, the surface plasmon sensor <NUM> is an abnormal transmission-type surface plasmon sensor for example. That is, although this embodiment utilizes the abnormal transmission-type surface plasmon sensor for measuring the transmitted light as an example, the objective can also be achieved by measuring the reflected light, the setup of which will be described later. The fluidic device <NUM> is used to inject a test sample onto the surface plasmon sensor <NUM>. The abnormal transmission-type surface plasmon sensor <NUM> is composed of gold-capped nanowire arrays. The above-mentioned two narrow-band filters <NUM> and <NUM> all have a narrow bandwidth (< <NUM>), wherein the long-band boundary of one of the narrow-band filters is close to the surface plasmon resonance wavelength of the surface plasmon sensor (λlong-boundary - λSPR≦<NUM>~<NUM>), and the short-band boundary of the other narrow-band filters is close to the surface plasmon resonance wavelength of the surface plasmon sensor (λSPR - λshort-boundary≦<NUM>~<NUM>). A drive device <NUM> is used to switch the positions of these filters. The illumination system <NUM> is a halogen bulb, a light emitting diode, or a white light source having a continuous bandwidth. The imaging module <NUM> described above is a monochrome imaging device <NUM>. In the image captured by the imaging device <NUM>, the image generated by the light beam passing through the long-band filter <NUM> is called a long-band surface plasmon image, and the image generated by the light beam passing through the short-band filter <NUM> is called a short-band surface plasmon image. The surface plasmon signal image can be obtained by the following equation of: <MAT>.

<FIG> shows a modification of the structure of <FIG>, and the modification is mostly the same as the structure of <FIG>. Different from the structure of <FIG>, the spectral modulation module <NUM> of <FIG>, which comprises a linear polarizer <NUM>, a long-band filter <NUM> and a short-band filter <NUM>, is disposed between the surface plasmon sensor <NUM> and the imaging device <NUM> (camera).

<FIG> shows another substitute structure of this disclosure, which is mostly the same as the structure of <FIG>. Different from the structure of <FIG>, in the optical setup of <FIG>, the long-band surface plasmon image and the short-band surface plasmon image are captured by two monochrome imaging devices <NUM> and <NUM>' (cameras), respectively. To implement this method, a <NUM>/<NUM> light splitter (<NUM>, Chroma) is provided to split the light beam, and two bandpass filters <NUM> and <NUM> (ET640/<NUM> and ZET660/20x, Chroma) are disposed in front of the two imaging devices <NUM> and <NUM>' (cameras), respectively. According to this structure, the long-band surface plasmon image and the short-band surface plasmon image can be captured simultaneously, so the additional time for switching the two bandpass filters is unnecessary, thereby improving the temporal resolution.

<FIG> shows a modification of the structure of <FIG>, and the modification is mostly the same as the structure of <FIG>. Different from the structure of <FIG>, in the structure of <FIG>, the prism-type surface plasmon sensor <NUM>' is used for replacing the abnormal transmission-type surface plasmon sensor <NUM> of <FIG>.

<FIG> shows another substitute structure of this disclosure, which is mostly the same as the structure of <FIG>. Different from the structure of <FIG>, in the embodiment of <FIG>, the long-band surface plasmon image and the short-band surface plasmon image are captured by one monochrome imaging device <NUM> (camera), respectively. To implement this method, a built-in <NUM>/<NUM> light splitter (<NUM>, Chroma) is provided to split the light beam, and an image dividing device <NUM> containing two bandpass filters <NUM> and <NUM> (ET640/<NUM> and ZET660/20x, Chroma) is provided to divide the image. Then, the imaging device <NUM> (camera) can form the images. According to this structure, the long-band surface plasmon image and the short-band surface plasmon image can be captured simultaneously by the same camera in different pixel regions, so the additional time for switching the two bandpass filters <NUM> and <NUM> is unnecessary, thereby improving the temporal resolution.

<FIG> is a schematic diagram showing an apparatus for surface plasmon imaging, which comprises a prism-type surface plasmon sensor <NUM>', according to an embodiment of this disclosure. In this embodiment, the light beam is emitted from the light source to the prism-type surface plasmon sensor <NUM>' and is then reflected to the imaging device <NUM> (camera). The spectral modulation module <NUM>, which comprises a linear polarizer <NUM>, a long-band filter <NUM> and a short-band filter <NUM>, is disposed between the broadband light source <NUM> and the prism-type surface plasmon sensor <NUM>'. The drive device <NUM> of the spectral modulation module <NUM> is configured to make the light beam to pass through the long-band filter <NUM> for capturing the long-band surface plasmon image. Afterwards, the drive device <NUM> further makes the light beam to pass through the short-band filter <NUM> for capturing the short-band surface plasmon image. Finally, the surface plasmon signal image can be calculated.

To be noted, the position of the spectral modulation module <NUM>, which comprises the linear polarizer <NUM>, the long-band filter <NUM> and the short-band filter <NUM>, is not limited to that shown in <FIG>. For example, the spectral modulation module <NUM> can be disposed between the surface plasmon sensor <NUM> and the imaging device <NUM> (camera). <FIG> shows a modification of the structure of <FIG>, and the modification is mostly the same as the structure of <FIG>. Different from the structure of <FIG>, the spectral modulation module <NUM>, which comprises the linear polarizer <NUM>, the long-band filter <NUM> and the short-band filter <NUM>, is disposed between the prism-type surface plasmon sensor <NUM>' and the imaging device <NUM> (camera).

In addition, and outside the scope of the appended claims, this disclosure also provides an apparatus for surface plasmon imaging. Referring to <FIG>, the apparatus comprises a light source module <NUM>, a sensing unit <NUM> (including a surface plasmon resonance sensing chip), an imaging device <NUM>, and an image processing unit. The structure of the apparatus is mostly the same as the structure of <FIG>. Different from the structure of <FIG>, the example of <FIG> comprises a light source module <NUM> for emitting incident light beams of two different bandwidths to substitute the broadband light source and spectral modulation module <NUM> of <FIG>. As shown in <FIG>, the light source module <NUM> comprises a first bandwidth light-emitting submodule <NUM> and a second bandwidth light-emitting submodule <NUM>. The first bandwidth light-emitting submodule <NUM> comprises a first bandwidth light-emitting unit <NUM>, a first lens <NUM> and a first linear polarizer <NUM>. The first bandwidth light-emitting unit <NUM> emits a first bandwidth incident light beam, which is emitted to the surface plasmon sensor (the surface plasmon resonance sensing chip) <NUM> through the first lens <NUM> and the first linear polarizer <NUM> in order. Similarly, the second bandwidth light-emitting submodule <NUM> comprises a second bandwidth light-emitting unit <NUM>, a second lens <NUM> and a second linear polarizer <NUM>. The second bandwidth light-emitting unit <NUM> emits a second bandwidth incident light beam, which is emitted to the surface plasmon resonance sensing chip <NUM> through the second lens <NUM> and the second linear polarizer <NUM> in order. The surface plasmon sensor <NUM> is an abnormal transmission-type surface plasmon sensor for example. That is, although this example utilizes the abnormal transmission-type surface plasmon sensor for measuring the transmitted light as an example, the objective can also be achieved by measuring the reflected light, the setup of which will be described later. The fluidic device <NUM> is used to inject a test sample onto the surface plasmon sensor <NUM>. The abnormal transmission-type surface plasmon sensor <NUM> is composed of gold-capped nanowire arrays. The above-mentioned two incident light beams (the first bandwidth incident light beam and the second bandwidth incident light beam) both have a narrow bandwidth (< <NUM>), wherein the long-band boundary of the second bandwidth incident light beam is close to the resonance wavelength of the surface plasmon sensor (λlong-boundary - λSPR≦<NUM>~<NUM>), and the short-band boundary of the first bandwidth incident light beam is close to the surface plasmon resonance wavelength of the surface plasmon sensor (λSPR - λshort-boundary≦<NUM>∼<NUM>). A drive device <NUM> is used to switch the positions of the first bandwidth light-emitting submodule <NUM> and the second bandwidth light-emitting submodule <NUM>. The imaging system described above is a monochrome imaging device <NUM>. In the image captured by the imaging device <NUM>, the image generated by the long-band light beam (the first bandwidth incident light beam) is called a long-band surface plasmon image, and the image generated by the short-band light beam (the second bandwidth incident light beam) is called a short-band surface plasmon image. The surface plasmon signal image can be obtained by the following equation of: <MAT>.

<FIG> shows a scanning device according to an embodiment of this disclosure and an image of a nanostructure plasmon sub-biochip; and <FIG> show red, green and blue segment analysis method applied to a transmission-type plasmon sub-chip with narrow resonance peak and narrow resonance dip. The white light LED lamp of the transmission-type scanner can be divided into red, green and blue segments, and the resonance peak or dip of the metal nanostructure can be adjusted to be presented on the overlapping region of red and green segments or blue and green segments. Regarding the resonance peak, when the peak increases according to the environmental refractive index so as to generate the red shift, the shift can cause the increase of the transmitted light intensity of red segment (AR) and the decrease of the transmitted light intensity of green segment (BG). On the contrary, regarding the resonance dip, the shift of dip can cause the decrease of the transmitted light intensity of red segment and the increase of the transmitted light intensity of green segment. The custom equation of the red and green segments at the resonance peak and resonance dip can be represented as follow: γ=(AG-BR)/(AG+BR) and γ=(AR-BG)/(AR+BG). This equation can remove the common noise and increase the sensing ability. In <FIG>, it indicates that the metal nanostructure can be metal-capped nanowire multi-array structure.

The apparatus of <FIG> and <FIG> is used in this example, wherein the images of different spectra can be captured by the same camera, and the surface plasmon signal image can also be calculated. In this structure, a stable intensity halogen lamp is used to generate a broadband light source, and two bandpass filters (ET640/<NUM> and ZET660/20x, Chroma) are mounted on a motorized filter wheel for light switching. The metal-capped nanowire array structure is used as the surface plasmon sensor, wherein the period of the nanowire array is designed to be <NUM> so that the surface plasmon resonance wavelength in water is <NUM>. This surface plasmon sensor is placed on the microscope's observation platform for imaging. A complementary monochromatic metal oxide semiconductor camera (C11440, Hamamatsu) is used to capture images of the surface plasmon sensor. The upper/lower rows of images of <FIG> represent the transmitted images of the surface plasmon sensors before and after the injection of the high refractive index solution. Comparing the intensities of the long-band surface plasmon images and the short-band surface plasmon images before and after the injection of the high refractive index solution, it is found that there is no obvious change, but the surface plasmon signal images clearly show a clear signal in the flow channel region.

The detection limit is an important indicator of the surface plasmon sensor. In this embodiment, the detection limit of this disclosure will be tested by using the apparatus of <FIG> and <FIG> and compared with the standard wavelength shift analysis method. In the test using the apparatus and method of the present disclosure, a structure composed of gold-capped nanowire arrays is used as a surface plasmon sensor, and a period of the nanowire array structure in the surface plasmon sensor is designed as <NUM>. This surface plasmon sensor is placed on the microscope's observation platform for imaging, and a complementary monochrome metal oxide semiconductor (CMOS) camera (C11440, Hamamatsu) is used to capture the image of the surface plasmon sensor. The glucose/water with different proportions and known refractive indexes is sequentially feed by using the micro-fluidic device, and the variation and stability of the sensing curves are analyzed so as to obtain the sensitivity and sensing limit. The standard wavelength shift analysis method uses a single-point measurement method to illuminate the same surface plasmon sensor with a focused spot, and the period of the nanowire array structure is designed to be <NUM>. The micro-fluidic device sequentially injects the glucose/water with different proportions and known refractive indexes (the same as that used in the above test utilizing the apparatus and method of this disclosure). The transmitted light is collected by the optical fiber, and transmitted to the spectrometer for obtaining the spectrum signal, thereby analyzing the sensitivity and detection limit by observing the wavelength shifts of the spectral peak. The sensitivity is defined as the amount of change of the surface plasmon signal caused by the change of refractive index per unit, and the detection limit is defined as the minimum amount of refractive index change that can be resolved by the system. The up-left graph of <FIG> shows the sensing curve measured by the standard wavelength shift analysis device and method. The up-right graph of <FIG> shows the sensing curve measured by the apparatus and method of the present disclosure. The two graphs can clearly indicate that when the refractive index of the test sample increases from n<NUM> = <NUM> to n<NUM> = <NUM>, the wavelength and surface plasmon change signal also increase, and when the refractive index returns from n<NUM> = <NUM> to n<NUM> = <NUM>, the wavelength and surface plasmon change signal also return to the baseline. Comparing the two signals, it can be seen that this disclosure can not only form the surface plasmon resonance image, but also reduce the influence of noise because of the self-reference signal. Compared to the commonly used resonance peak changes, this disclosure has better performance in signal to noise ratio. Further, the linear regression method is used to analyze the sensing curve as shown in the down-left figure of <FIG>, and the relationship between the peak wavelength and the refractive index is λ=<NUM>+<NUM>×RIU (RIU: refractive index unit). Wherein, <NUM> is the detection sensitivity of the standard wavelength shift analysis method, and the system stability can be obtained by continuously analyzing the peak wavelength signal of the glucose/water with the same refractive index. In this embodiment, the standard deviation of the peak wavelength signal obtained by analyzing the n<NUM> measurement region is <NUM>. Therefore, the detection limit of the method can be found to be <NUM> × <NUM>-<NUM> RIU. On the other hand, the relationship between the change of the surface plasmon signal and the refractive index is γ=-<NUM>+<NUM>×RIU, wherein <NUM> is the detection sensitivity of this disclosure. In this embodiment, the standard deviation of the surface plasmon signal obtained by analyzing the n<NUM> measurement region is <NUM> × <NUM>-<NUM>. Therefore, the detection limit of the method can be found to be <NUM> × <NUM>-<NUM> RIU. This value fully shows that this disclosure can achieve lower detection limit than the standard wavelength shift analysis method by the influence of self-reference signal noise. More importantly, the standard wavelength shift analysis method is only suitable for single point measurement, but the method and apparatus proposed in the present disclosure are suitable for full-field imaging, and can analyze multi-point signals at the same time, which has obvious benefits in measuring throughput.

The change of surface plasmon signal over time is called the sensing curve, which is an important application in the field of surface plasmon detection. The sensing curve can provide quite a lot of information, such as the affinity, the dissociation and binding rate of the labeled and test samples, the concentration of the target molecule, and the specificity of the interactive molecules. <FIG> illustrates an embodiment of a high throughput measurement sensing curve based on this disclosure. In this embodiment, the long-band surface plasmon image and the short-band surface plasmon image are sequentially recorded, and the time-series surface plasmon signal image is also calculated. Since all surface plasmon signal images are recorded at all times, the user can extract the sensing map in a particular area. The right graph of <FIG> includes three sensing curves labeled by exp, ref and non, which correspond to the surface plasmon signals extracted from the regions exp, ref, and non of the left graph, respectively. The region exp is located in the flow channel and is on the metal-capped nanowire array structure, so the surface plasmon signal can be obtained. Conversely, the region ref is not located on the nanowire array structure and the region non is outside the flow channel, so there is no surface plasma signal change.

To further confirm the feasibility of applying the scanner and the red, green and blue segment analysis method to the plasmon sensing chip. <FIG> shows the detection experiments of different proportions of glycerin/water by using a commercial transmission-type scanner and a double-layer aluminum nano-slit chip (left), and the right side shows the transmission images of the plasmon sub-chips scanned by the scanner in different proportions of glycerin/water, wherein the refractive index of glycerin water is between <NUM> and <NUM>. In this example, in order to allow the resonance peak of the structure to appear in the red and green segments, the period length of the double-layer aluminum nano-slit chip is adjusted to <NUM>.

Claim 1:
An apparatus for surface plasmon resonance imaging, comprising:
a single light source module (<NUM>) for emitting a broadband light beam;
a spectral modulation module (<NUM>) comprising a first bandwidth filter (<NUM>) and a second bandwidth filter (<NUM>), wherein the spectral modulation module (<NUM>) is arranged to receive the broadband light beam to generate a first bandwidth incident light beam and a second bandwidth incident light beam, respectively;
a surface plasmon resonance sensing chip (<NUM>) having a surface plasmon resonance wavelength, wherein the surface plasmon resonance sensing chip (<NUM>) is arranged to receive the first bandwidth incident light beam and the second bandwidth incident light beam for correspondingly generating a first bandwidth plasmon resonance light beam and a second bandwidth plasmon resonance light beam;
an imaging module (<NUM>) configured to receive the first bandwidth plasmon resonance light beam and the second bandwidth plasmon resonance light beam for generating a first bandwidth surface plasmon image signal and a second bandwidth surface plasmon image signal, respectively; and
an image processing unit coupled to the imaging module (<NUM>) and receiving the first bandwidth surface plasmon image signal and the second bandwidth surface plasmon image signal for generating a surface plasmon signal image;
wherein the surface plasmon resonance wavelength of the surface plasmon resonance sensing chip (<NUM>) is less than a minimum wavelength of the first bandwidth plasmon resonance light beam and greater than a maximum wavelength of the second bandwidth plasmon resonance light beam, a difference between the surface plasmon resonance wavelength and the minimum wavelength of the first bandwidth plasmon resonance light beam is less than <NUM>, and a difference between the surface plasmon resonance wavelength and the maximum wavelength of the second bandwidth plasmon resonance light beam is less than <NUM>.