KR20090006942A - Bio-sensor and apparatus for measuring the bio-sensor - Google Patents

Abstract

A biosensor and a biosensor measuring system are provided to measure very small signal amplitude and to monitor reaction of biomass in real time without additional label. A biosensor measuring system(20) comprises a biosensor(10) in which biomass is attached to the surface, a pumping system(30) inducing birefringence in the biosensor by applying energy beam, and an interferometer(40) measuring the variation of the birefringence induced by the pumping system so that the kind of the biomass is determined according to the birefringence variation measured by the interferometer.

Description

Bio-sensor and apparatus for measuring the bio-sensor

1 is a cross-sectional view showing a biosensor according to a preferred embodiment of the present invention.

2 is a block diagram showing the overall configuration of the biosensor measuring apparatus according to an embodiment of the present invention.

3 is a graph showing the birefringence induced according to the size of the metal nanoparticles of the biosensor and the output of the pump light source in the biosensor measuring apparatus according to the preferred embodiment of the present invention.

<Explanation of symbols for the main parts of the drawings>

10: biosensor

100: dielectric substrate

110: metal nanoparticle layer

120: biomaterial layer

20: biosensor measuring device

30: pumping system

300: pump light source

310: polarizer

320: λ / 2-phase delay plate

330: Chopper

40: interferometer

400: probe light source

410: λ / 4-phase delay plate

420: HWP

430: polarization detection unit

432 polarized beam splitter

433: first filter

434: first light detector

436: second filter

437: second photodetector

440: differential amplifier

450: lock-in amplifier

The present invention relates to a biosensor and a biosensor measuring apparatus, and more particularly, a biosensor in a SPR region using a biosensor and a homodyne interferometer having a metal nanoparticle layer formed on a glass substrate and a biomaterial attached thereto. The present invention relates to a biosensor measuring apparatus for determining a biomaterial by measuring induced birefringence.

Surface Plasmon is a collective charge density oscillation of electrons occurring on the surface of a metal thin film, and the surface plasmon waves generated are surface electromagnetic waves propagating along the interface between the metal and the dielectric. Excitation of surface plasmons is caused by the application of an electric field to two medium interfaces with different dielectric functions from outside, that is, the interface between metal and dielectric, which induces surface charges due to the discontinuity of the electric component perpendicular to the two medium interfaces, Oscillations appear as surface plasmon waves. The surface plasmon wave described above, unlike electromagnetic waves in free space, is a wave that vibrates parallel to the plane of incidence and has a polarization component of p-polarization. Therefore, excitation of surface plasmons by optical means is possible only by TM polarized electromagnetic waves.

TM polarized incident wave is totally reflected at the interface of the metal thin film and dissipated wave is exponentially reduced into the metal thin film at the interface, but at a certain angle of incidence and thickness of the thin film, the incident wave in the direction parallel to the interface and the surface plasmon wave coincide If you do, resonance will occur. At this time, the energy of the incident wave is absorbed by the metal thin film so that the reflected wave disappears and the electric field distribution in the direction perpendicular to the interface is exponentially largest and rapidly decreases toward the metal thin film. Surface Plasmon Resonance (hereinafter referred to as 'SPR') is called, and the angle at which the reflectance of the incident light decreases is called the surface plasmon resonance angle.

The resonance angle at which surface plasmon resonance occurs, that is, the angle at which the reflected light is minimized, results in an effective reactive index that changes as the mass of the metal thin film surface layer dielectric increases or the structure changes, resulting in a change in the resonance angle. Therefore, using the principle of SPR, which can measure the change of these materials by optical method, it is possible to change the resonance angle of biochemical reactions such as selective bonding or separation between various biochemicals through proper chemical modification of metal thin film surface layer. SPR sensors are often used as highly sensitive biochemical sensors.

In general, as a SPR-based biosensor, a number of prior arts have been proposed as a method of measuring a change in dielectric constant or refractive index corresponding to a change in a sample on a metal film after coating a metal film on a transparent prism about 50 nm.

The first method is mentioned in US Pat. No. 4,889,427, which is a method of measuring the resonance angle and its change while changing the incident angle θ by using the incident light of a single wavelength light source and a prism having a fixed refractive index n ₀ .

In addition, the second method is a method for measuring a change in wavelength according to resonance conditions in a state in which a multi-wavelength light such as white light is used as a light source and the incident angle θ is fixed in the manner mentioned in US Pat. No. 5,359,681.

The third method focuses on the center of the medium by using a single wavelength light source expanded in the manner mentioned in US Pat. No. 4,844,613 and the like, and receives multiple channels such as a photodiode array (PDA). It is a method of measuring the resonance angle without a rotary drive by using a device.

Recently, techniques have been proposed that utilize local surface plasmon effects caused by metal nanoparticles rather than metal films. If metal nanoparticles are spread in the dielectric material, field enhancement occurs due to surface plasma resonance (SPR) phenomenon caused by the metal nanoparticles, resulting in very large optical nonlinearity. When the biomaterial to be measured is attached to the fabricated sensor substrate, the refractive index is changed by the interaction of the biomaterial, which causes a change in the surface plasma resonance phenomenon. In conclusion, the change in refractive index induced according to the type of biomaterial and the type of interaction occurs, and by measuring the change, the state and properties of the biomaterial can be determined.

Such metal nanoparticles are coated on a general glass substrate for a microscope and used as a sensor substrate, such as transmission localized surface plasmon resonance spectroscopy (T-LSPR) and propagating surface plasmon resonance spectroscopy (P-SPR). T-LSPR is a biosensor using a transparent sensor substrate coated with metal nanoparticles with a single layer or very thin thickness. In the case of P-SPR, the manufacturing of the sensor substrate is slightly different, but the basic concept is the same, so the bio-object to be measured on the sensor substrate coated with metal nanoparticles on the transparent glass is put on the SPR using UV-vis spectrometer. This is a method of measuring the interaction of the bio-object based on the change of the absorption coefficient. However, this method requires a spectrometer to measure the change of multiple light sources, which not only requires complicated alignment of the optical system but also increases the price.

An object of the present invention for solving the above problems is to generate a birefringence due to the optical Kerr effect in the SPR region with respect to the biosensor and to detect a change in the birefringence using a homodyne interferometer to determine the biomaterial of the biosensor It is to provide a sensor measuring device.

Another object of the present invention is to provide a biosensor measuring apparatus and a biosensor capable of mass production at low cost and easy alignment of optical systems in order to determine a biomaterial attached to a biosensor having a metal nanoparticle layer.

Features of the present invention for achieving the above technical problem relates to a biosensor measuring apparatus, the biosensor measuring apparatus is

A biosensor with a biomaterial attached to its surface

A pumping system which induces birefringence in the biosensor by providing an energy beam to the biosensor, and

And an interferometer for measuring the amount of change in the birefringence with respect to the birefringence induced by the biosensor by the pumping system,

The type of biomaterial attached to the biosensor is determined according to the amount of change in birefringence measured by the interferometer.

The biosensor of the biosensor measuring device having the above-mentioned characteristics

Transparent dielectric substrate,

A metal nanoparticle layer coated with metal nanoparticles on the dielectric substrate, and

A biomaterial layer formed by linking a biomaterial to the metal nanoparticles of the metal nanoparticle layer,

The metal nanoparticle layer is preferably made of a single layer or a multilayer structure of nano metal particles having a diameter of several tens to several tens of nanometers.

The pumping system of the biosensor measuring device having the characteristics described above

Pump light source,

A polarizer disposed in front of the pump light source and arranged on the same axis of the pump light source and the biosensor to polarize a beam from the pump light source, and

A λ / 2-phase delay plate (HWP) disposed between the polarizer and the biosensor and disposed on the same axis as the pump light source and the polarizer to retard a beam from the polarizer,

A λ / 4-phase delay plate (HWP) disposed between the polarizer and the biosensor and disposed on the same axis as the pump light source and the polarizer to retard a beam from the polarizer,

It is preferable to have a chopper disposed between the λ / 2-phase delay plate and the biosensor to convert the beam from the λ / 2-phase delay plate into a pulse in the form of a square wave.

The interferometer of the biosensor measuring device having the above-mentioned characteristics

A probe light source for outputting a linearly polarized beam,

A λ / 4-phase delay plate disposed on the front surface of the probe light source and arranged on the same axis of the probe light source and the biosensor to retard the linearly polarized beam from the probe light source and convert it into a circularly polarized beam;

A λ / 2-phase delay plate disposed in front of the biosensor to retard the elliptical polarization beam output from the biosensor,

A polarization detector for dividing the beam passing through the λ / 2-phase delay plate into a vertically polarized beam and a horizontally polarized beam;

A differential amplifier receiving a vertical polarization beam and a horizontal polarization beam output from the polarization sensing unit as input signals and amplifying the difference values of the input signals;

A lock-in amplifier configured to amplify the output signal of the differential amplifier at a reference frequency,

The biosensor is disposed between the λ / 4-phase delay plate and the λ / 2-phase delay plate, and the probe light source preferably uses a He-Ne Laser.

The polarization detection unit of the interferometer having the above-described characteristics is to detect and separately separate the vertical beam and the horizontal polarization beam of light output from the biosensor,

A polarization beam splitter (PBS) for reflecting vertical polarization components of the input beam and passing horizontal polarization components,

A first filter disposed on the same axis as the reflective surface of the PBS and allowing the vertical polarization component to pass therethrough;

A first photo detector for detecting a vertically polarized beam that has passed through the first filter,

A second filter disposed on the same axis as the front surface of the PBS and passing the horizontal polarization component;

Preferably, a second photo detector for detecting the horizontally polarized beam passing through the second filter.

Hereinafter, the structure and operation of a biosensor measuring apparatus and a biosensor according to a preferred embodiment of the present invention will be described in detail with reference to the accompanying drawings.

Bio sensor

1 is a cross-sectional view showing a biosensor according to a preferred embodiment of the present invention. Referring to FIG. 1, the biosensor 10 according to the present invention includes a dielectric substrate 100, a metal nanoparticle layer 110 formed on the substrate 100, and a biomaterial layer 120 attached to the metal nanoparticle layer. It is provided.

The dielectric substrate 100 may be a transparent glass substrate. The metal nanoparticle layer 110 is a layer in which metal nanoparticles are coated very thinly on the dielectric substrate 100 as a single layer or a multilayer structure. The metal nanoparticles preferably use precious metals such as Au, Ag, Pt, and the diameter of the particles is preferably in the range of 0.5 to 5.0 nm. The biomaterial layer 120 is a layer to which biomaterials or biomaterials such as DNA and protein are attached to the metal nanoparticles of the metal nanoparticle layer 110.

The biosensor having the above-described configuration is made of very thin metal nanoparticles on a transparent glass substrate, thereby enabling mass production at low cost.

Biosensor measuring device

On the other hand, the biosensor measuring device according to the present invention is optically induced in the biosensor with a homodyne interferometer of the pump-probe type using a local surface plasmon resonance phenomenon in the biosensor having the above-described configuration By monitoring the change in the birefringence degree, it is possible to determine the biomaterial attached to the biosensor.

2 is a block diagram showing the overall configuration of the biosensor measuring apparatus according to an embodiment of the present invention. 2, the configuration and operation of the biosensor measuring apparatus according to the present invention will be described in detail.

2, the biosensor measuring apparatus 20 according to the present invention is the biosensor 10, the pumping system 30 to induce birefringence by the optical kerr effect in the biosensor and the birefringence diagram in the biosensor An interferometer 40 is provided for measuring the fine change of.

The pumping system 30 includes a pump light source 300, a polarizer 310, a λ / 2-phase delay plate (HWP) 320, and a chopper 330. Equipped.

The pump light source 300 is preferably Nd; YAG laser. The HWP 320 adjusts the angle of the linear polarization beam by delaying the phase of the input polarization beam. By sequentially placing the polarizer 310 and the HWP 320 in front of the pump light source 300, the biosensor 10 after the beam from the pump light source 300 passes through the polarizer 310 and the HWP 320. Is supplied. The biosensor 10 generates an optical kerr effect in the SPR region by a beam provided from a light source, and as a result, ordinary rays and extraordinary rays are generated by birefringence. At this time, the phase of the birefringent two light beams is generated according to the type of biomaterial attached to the biosensor 10. The birefringence degree Δn is determined by the difference between the refractive index n _x of the normal light and the refractive index n _y of the abnormal light.

On the other hand, the chopper 330 cuts the beam at a predetermined time to convert the beam provided from the light source into a pulse wave in the form of a square wave.

In another embodiment of the pumping system of the present invention, a λ / 4-phase delay plate (hereinafter referred to as 'QWP'; not shown) is further provided between the pump light source and the biosensor. The linearly polarized beam from the pump light source may be converted into a circularly polarized beam.

On the other hand, the interferometer 40 measures the degree of minute change of the birefringence generated in the biosensor by the pumping system, and determines the type of biomaterial attached to the biosensor according to the degree of change. The interferometer 40 includes a probe light source 400, a λ / 4-phase delay plate (QWP) 410, a HWP 420, a polarization detector 430, and a differential amplifier. 440, a lock-in amplifier 450.

The probe light source 400 preferably uses a He-Ne Laser that generates a linearly polarized beam.

The QWP 410 generates circularly polarized light by rotating the linearly polarized beam starting from the light source 400 by 45 ° from the main axis. The circularly polarized light from the QWP 410 is incident on the biosensor, and the phase difference is generated by the biomaterial of the biosensor 10, and as a result, the circularly polarized light is converted into elliptical polarized light.

The HWP 420 is a λ / 2-phase retardation plate, and the elliptical polarization beam output from the biosensor has a phase difference of π between the vertical component and the horizontal component after passing through the HWP 420.

The polarization detector 430 separates and detects vertical and horizontal polarization components of the beam output from the biosensor, and is referred to as a polarization beam splitter (PBS) 432; a first filter 433 and a first photodetector 434 for detecting the vertically polarized beam I ₁ passing through the first filter, a second filter 436 and a horizontal polarized beam I ₂ passing through the second filter. The second photodetector 437 which detects this is provided. The polarization beam splitter 432 reflects the polarization vertical component I ₁ of the beam passing through the HWP 420 to the first filter 433, and reflects the polarization horizontal component I ₂ to the second filter 436. Pass it through. The polarized vertical component passing through the first filter 433 is detected by the first photo detector 434 and provided to the first input signal I ₁ of the differential amplifier 440, and passes through the second filter 436. One polarized horizontal component is detected by the second photo detector 437 and provided to the second input signal I ₂ of the differential amplifier 440.

The first photo detector 434 and the second photo detector 437 may use photodiodes for receiving light.

The differential amplifier 440 receives a first input signal and a second input signal from the polarization detector 430, respectively, and the difference between the received first input signal and the second input signal (I _diff = I ₁ -I ₂ ) to amplify and output.

The lock-in amplifier 450 receives a reference frequency signal from the chopper 330, fixes the output signal of the differential amplifier 440 to the reference frequency signal, and amplifies and outputs the output signal.

The interferometer 40 having the above-described configuration becomes a circularly polarized beam and enters the biosensor 10 while the linearly polarized beam starting from the light source 400 passes through the QWP 410 aligned with the main axis at 45 degrees. The phase difference optically induced by the biomaterial of the sensor converts the incident circularly polarized beam into an elliptical polarized beam, which is divided by the rear end PBS to amplify the difference in the signal measured by the photodetectors 1 and 2 By detecting it, a very small signal can be detected. The output signal output from the differential amplifier 440 is a signal obtained by amplifying an amount of change in birefringence induced by the biosensor. Therefore, the biosignal attached to the biosensor is determined according to the output signal of the differential amplifier 440.

Although the present invention has been described above with reference to preferred embodiments thereof, this is merely an example and is not intended to limit the present invention, and those skilled in the art do not depart from the essential characteristics of the present invention. It will be appreciated that various modifications and applications which are not illustrated above in the scope are possible. For example, in the embodiment of the present invention, the type of light source used for the pumping system and the interferometer may be modified in various ways to improve the performance of the entire system. And differences relating to such modifications and applications should be construed as being included in the scope of the invention defined in the appended claims.

According to the present invention, the biosensor can be produced in large quantities at low cost.

In addition, the biosensor measuring apparatus according to the present invention is very simple in the configuration of the optical system can measure a very small signal size, and also provides a label-free real-time interaction of the biomaterials You can monitor.

In addition, while conventional SPR sensors using absorption spectrum and resonant angle variation measurement have to be difficult to align and expensive optical components such as spectrometers and rotational optical systems, the biosensor measuring apparatus according to the present invention is configured of the optical system This simple and more precise signal detection is not only possible, but also easy to miniaturize.

On the other hand, Figure 3 is a graph showing the change in the birefringence according to the change in the birefringence according to the change in the diameter of the metal nanoparticles of the biosensor according to the present invention and the output of the light source of the pumping system. Referring to FIG. 3, it can be seen that the birefringence induced by the biosensor increases as the diameter size of the metal nanoparticles of the biosensor increases and the output of the pump light source of the pumping system increases. On the other hand, when the biosensor is composed of only a glass layer (Corning glass) without the metal nanoparticle layer, it can be seen that birefringence does not occur at all even if the output of the pump light source is increased.

Claims (9)

Biosensors having biomaterials attached to surfaces; A pumping system providing an energy beam to the biosensor to induce birefringence in the biosensor; And Interferometer for measuring the amount of change in birefringence with respect to the birefringence induced by the biosensor by the pumping system And a type of biomaterial attached to the biosensor according to the amount of change in the birefringence measured by the interferometer. The method of claim 1, wherein the biosensor Transparent dielectric substrate, A metal nanoparticle layer coated with metal nanoparticles on the dielectric substrate, and And a biomaterial layer formed by linking a biomaterial to the metal nanoparticles of the metal nanoparticle layer. The biosensor measurement according to claim 2, wherein the metal nanoparticle layer is made of metal nanoparticles having a diameter of several tens to several tens of nanometers, and the metal nanoparticle layer is formed of a single layer or a multilayer structure of nano metal particles. Device. The method of claim 1, wherein the pumping system Pump light source; A polarizer disposed in front of the pump light source and arranged on the same axis of the pump light source and the biosensor to polarize the beam from the pump light source; And A λ / 2-phase delay plate (HWP) disposed between the polarizer and the biosensor and disposed on the same axis as the pump light source and the polarizer to retard a beam from the polarizer; A λ / 4-phase delay plate (HWP) disposed between the polarizer and the biosensor and disposed on the same axis as the pump light source and the polarizer to retard a beam from the polarizer; Biosensor measuring apparatus consisting of. The pumping system of claim 4, wherein the pumping system further comprises a chopper disposed between the λ / 2-phase delay plate and the biosensor, wherein the chopper comprises a beam from the λ / 2-phase delay plate. The biosensor measuring apparatus characterized by converting the pulse into a square wave form and outputting. The method of claim 1, wherein the interferometer A probe light source for outputting a linearly polarized beam; A λ / 4-phase delay plate disposed on a front surface of the probe light source and disposed on the same axis of the probe light source and the biosensor to retard the linearly polarized beam from the probe light source and convert the linearly polarized beam into a circularly polarized beam; A λ / 2-phase delay plate disposed in front of the biosensor to retard an elliptical polarization beam output from the biosensor; A polarization detector for dividing the beam passing through the λ / 2-phase delay plate into a vertical polarization beam and a horizontal polarization beam; A differential amplifier receiving a vertical polarization beam and a horizontal polarization beam output from the polarization sensing unit as input signals and amplifying a difference value between the input signals; And Lock-In amplifier for amplifying the output signal of the differential amplifier to a reference frequency And a biosensor disposed between the λ / 4-phase delay plate and the λ / 2-phase delay plate. The method of claim 6, wherein the polarization detector detects the vertical beam and the horizontal polarization beam of light output from the biosensor separately. A polarization beam splitter (PBS) for reflecting vertical polarization components of the input beam and passing horizontal polarization components, A first filter disposed on the same axis as the reflective surface of the PBS and allowing the vertical polarization component to pass therethrough; A first photo detector for detecting a vertically polarized beam that has passed through the first filter, A second filter disposed on the same axis as the front surface of the PBS and passing the horizontal polarization component; A second photo detector for detecting a horizontally polarized beam that has passed through the second filter Biosensor measuring apparatus comprising a. The biosensor measuring apparatus of claim 6, wherein the lock-in amplifier fixes and amplifies the output signal of the differential amplifier according to a reference frequency signal input from the chopper of the pumping system. Transparent dielectric substrates; A metal nanoparticle layer coated with metal nanoparticles on the dielectric substrate; And A biomaterial layer formed by linking a biomaterial to metal nanoparticles of the metal nanoparticle layer; And a metal nanoparticle layer, wherein the metal nanoparticles have a single layer or a multilayer structure.

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