JPH05107479A - Scanning optical microscope - Google Patents

Abstract

PURPOSE:To provide high speed scanning equivalent to video rate and achieve theoretical resolution of a con-focal microscope in a scanning optical microscope in which an acousto-optical deflection element is used. CONSTITUTION:In a scanning optical microscope 23 in which light beam is deflected by an acousto-optical deflection device 23, an width W of ultrasonic wave surface in optical axis direction for propagate the light beam in the acousto-optical deflection element 23 is set as follows: Where an angle between an incident light beam and the ultrasonic wave surface is thetain, an acoustic velocity in the acousto-optical deflection element is V, an opening width of the acousto-optical deflection element is D, a max. deflection angle of light beam in the acousto-optical deflection element is theta max., a coefficient of frequency-time variation of high frequency sweep signal is A, and a coefficient of constant is k, W<= (k.V<2>)/(D.A.tan (theta max-thetain)).

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a scanning optical microscope which scans a sample in a predetermined direction by using an acousto-optic deflecting element.

[0002]

2. Description of the Related Art A scanning optical microscope of this type has been disclosed in Japanese Patent Laid-Open No. 61-62
It is disclosed in Japanese Patent Publication No.-80215, and its basic configuration is shown in FIG.

In this scanning optical microscope, a light beam emitted from a laser light source 1 is expanded by an expander 2 and is incident on an acousto-optic deflecting element 3, which diffracts the light beam. Is biased. Then, the light beam deflected by the acousto-optic deflecting element 3 is focused by the condenser lens 4, and the relay lens 5, the half mirror 6,
The light enters the vibrating mirror 8 via the total reflection mirror 7. The light beam reflected by the vibrating mirror 8 is focused by the objective lens 9 in the form of a minute spot and is incident on the sample 10.

In a scanning optical microscope equipped with such a scanning optical system, the minute spots formed on the sample 10 are
Due to the deflection action of the acousto-optic deflecting element 3, scanning is performed in a predetermined direction (main scanning direction), and the vibrating mirror 8 scans in a different direction (sub scanning direction).

The reflected light from the sample 10 passes through the half mirror 6 and is incident on the linear image sensor 11, and the linear image sensor 11 receives the reflected light from the sample 10 line by line in the main scanning direction. It converts optical information into electrical signals.

By the way, the acousto-optic deflecting element 3 used in the above-mentioned scanning optical microscope has an acoustic transducer 13 on the surface of the element body 12 as shown in FIG.
Is attached, and the light beam is incident from another surface orthogonal to that surface. By vibrating the acoustic transducer 13 with a high frequency sweep signal of a predetermined frequency, a traveling wave of an ultrasonic wave is generated in the element body 12 and the passing light beam is deflected with a predetermined diffraction efficiency.

In such an acousto-optical deflection element 3, the width W of the acoustic transducer 13 in the optical axis direction is set to be large in order to increase the diffraction efficiency.
The width of the ultrasonic wave front in the optical axis direction is usually about 5 mm.

[0008]

However, when high-speed optical scanning corresponding to a video rate is performed using an acousto-optic deflecting element in which the width of the ultrasonic wave front in the optical axis direction is increased in order to increase the diffraction efficiency. , Scanning that performs high-speed optical deflection equivalent to the video rate because the blur angle due to the influence of the width of the ultrasonic wave front in the acousto-optic deflector becomes larger than the blur angle due to diffraction and the laser spot on the sample surface cannot be narrowed down to the diffraction limit. The optical microscope has a problem in that it cannot realize a confocal state.

In the scanning optical microscope described above, the image forming characteristics in the direction in which light is deflected by the acousto-optic deflecting element 3 depend on the optical conjugate relationship (image forming relationship) between the linear image sensor 11 and the surface of the sample 10. is doing. Therefore, an image can be formed by the imaging action of the detection optical system even if the blur angle of the acousto-optic deflecting element 3 increases and the laser spot formed on the sample surface cannot be narrowed to the diffraction limit.

However, in order to construct a scanning optical microscope that completely satisfies the confocal condition, it is necessary to narrow the laser spot on the sample surface to the diffraction limit, and when the laser spot cannot be narrowed to the diffraction limit. , A perfect confocal scanning optical microscope cannot be realized.

The present invention has been made in view of the above-mentioned circumstances, and it is possible to perform high-speed scanning corresponding to a video rate by using an acousto-optic deflecting element and to achieve the theoretical resolution of a confocal microscope. An object is to provide a type optical microscope.

[0012]

In order to achieve the above object, the present invention applies an ultrasonic wave generated by applying a high frequency sweep signal to an acoustic transducer to an acousto-optic deflecting element through which a light beam from a light source passes. In the scanning optical microscope, in which the light beam is deflected to scan the sample by scanning and the object light from the sample is converted into an electrical signal to obtain optical information of the sample, the light is incident in the acousto-optic deflecting element. The angle formed by the light beam and the ultrasonic wavefront is θin, the sound velocity in the acousto-optic deflector is V, the opening width of the acousto-optic deflector is D, and the maximum deflection angle of the light beam in the acousto-optic deflector is θmax. , Where the frequency temporal change rate of the high-frequency sweep signal is A and the coefficient is k, the width W of the wavefront of the ultrasonic wave in the optical axis direction is W ≦ (k · V ² ) / (D · A · tan (θmax−θin)).

[0013]

According to the present invention, by taking the above means, the blur angle due to the width W in the optical axis direction of the ultrasonic wave front propagating in the acousto-optic deflecting element at the time of high-speed scanning corresponding to the video rate is reduced by the acousto-optic deflection. It becomes smaller than the blur angle of diffraction due to the aperture of the element, and a scanning imaging optical system having a diffraction limited resolution is realized even during high-speed scanning corresponding to the video rate. Therefore, the theoretical resolution of the confocal microscope is obtained even during high-speed scanning corresponding to the video rate.

[0014]

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS An embodiment of the present invention will be described below with reference to FIG.

In the scanning optical microscope of this embodiment, the light beam emitted from the laser light source 20 is expanded by the beam expander 21 to a size that fills the aperture of the optical system, and the expanded light beam passes through the beam splitter 22. It is incident on the acousto-optic deflector 23. A cylindrical lens 24 having a refraction direction oriented in the deflection direction of the acousto-optic deflecting element 23 is arranged on the surface facing the acousto-optic deflecting element 23 from which the incident light beam from the light source side is emitted, and further a condenser lens 25 and a relay lens. A galvano-mirror 27 is connected to the acousto-optic deflecting element 23 in an optically conjugate relationship by an optical system composed of 26. The light beam reflected by the galvanometer mirror 27 is condensed on the image plane of the objective lens 29 by the pupil projection lens 28, and this condensed spot is projected on the sample 30 by the objective lens 29.

In this embodiment, a horizontal drive signal is applied from the NTSC sync signal generator 31 to the sweep oscillator 32, and the sweep oscillator 32 generates a high frequency sweep signal having a frequency sweep waveform in synchronization with the horizontal drive signal. Let The acousto-optic deflection element 23 receives a high-frequency sweep signal from the sweep oscillator 32 by an acoustic transducer (not shown), and an ultrasonic wave having a predetermined width W in the acousto-optic deflection element 23 in the optical axis direction is orthogonal to the optical axis. To cause the light beam to be deflected in a direction perpendicular to the paper surface, for example. Due to the deflecting action of the acousto-optic deflecting element 23, the minute spots projected on the sample 30 are
For example, 0 is the main scan.

Further, a vertical drive signal is applied from the NTSC sync signal generator 31 to the galvano driver 33, and the galvano driver 33 swings the galvano mirror 27 in a predetermined direction in synchronization with the vertical drive signal. There is. The minute spots projected on the sample 30 in accordance with the swing of the galvanometer mirror 27, for example, sub-scan the sample 30.

That is, a scanning spot which is two-dimensionally raster-scanned by the acousto-optic deflecting element 23 and the galvanometer mirror 27 is formed on the objective image plane, and this scanning spot is projected on the sample 30.

The reflected light from the sample 30 traces back the original optical path, is branched from the illumination optical path by the beam splitter 22, and enters the spot imaging lens 34. Then, the light enters the photodiode 36 through a detection optical system including the spot imaging lens 34 and the pinhole 35.

A photoelectric signal obtained by photoelectric conversion by the photodiode 36 is amplified by a video amplifier 37 and then input to a video monitor 38. This video monitor 38 is
The composite sync signal generated from the TSC sync signal generator 31 is input. As a result, the raster scan on the video monitor 38 and the raster scan of the minute spot on the sample 30 can be synchronized, and an image can be formed on the video monitor 38 by the reflected light from the sample 30.

In the acousto-optic deflecting element 23 used in the scanning optical microscope of this embodiment, the width of the acoustic transducer in the optical axis direction, that is, the width W of the ultrasonic wave front in the optical axis direction satisfies the conditional expression described later. Is set. next,
Width W of the ultrasonic wave front in the acousto-optic deflector 23 in the optical axis direction
The conditions for setting will be described.

For example, when an acousto-optic deflection element using transverse ultrasonic waves in tellurium dioxide single crystal is used at a wavelength of 458 nm, the speed of sound is 650 m / sec and the center frequency is 103.
Since the frequency is MHz and the deflection bandwidth is 50 MHz, the maximum frequency is 128 MHz, and the deflection angle θ can be obtained from the equation (1). θ = λ · f / V (1) where λ is the used wavelength, f is the frequency of the high frequency signal applied to the acoustic transducer, and V is the speed of sound in the acousto-optic deflecting element. From the above formula (1), the maximum deflection angle θmax is
It becomes 5.168 (deg).

Here, the influence of the width W of the ultrasonic wave front in the optical axis direction will be described with reference to FIG. In Figure 4,
In the acousto-optic deflecting element in which an ultrasonic wave front having a width W advances in the vertical direction in the figure, rays A, B and C forming a parallel light beam form an angle θin with respect to the ultrasonic wave front from the left side in the figure. The incident state is shown.

In the figure, both the light rays A and B are incident on the same ultrasonic wave front, but the light ray A is incident on the left end of the wave front having the width W and the light ray B is incident on the right end of the same wave front. There is. Since rays A and B are incident on the same wavefront,
Both the + 1st-order light (A) and the + 1st-order light (B), which are the second-order diffracted lights, are emitted in parallel with the ultrasonic wave front at an angle of θs. Here, the angle formed by the 0th-order light and the + 1st-order light, that is, the deflection angle θ
Can be expressed by equation (2). θ = θs + θin (2)

At this time, the + 1st-order light (A) reaches the left end of the ultrasonic wavefront having the width W in the optical axis direction at the point X. The distance h from the reflecting surface when the + 1st-order light (A) crosses the ultrasonic wave front having the width W can be geometrically obtained from the figure as h = W · tan θs (3).

Here, when the frequency time change rate of the high frequency sweep signal for generating the ultrasonic wave traveling in the acousto-optic deflecting element is A (Hz / sec), the ultrasonic wave has a sound velocity V (m).
/ Sec), the change δf (Hz) of the acoustic frequency during the period of traveling the distance h can be expressed by δf = A · (h / V) = (A / V) · Wtan θs (4).

By substituting the equation (4) into the equation (1), the + 1st-order light (C) and the + 1st-order light (A) which are diffracted and deflected light rays of the light ray C incident on the point X at the left end of the ultrasonic wavefront. ) Angle δ
The value of θ is obtained. Therefore, the above δθ is obtained from the equation (5) by substituting the equation (4) into the equation (1). δθ = (λA / V ² ) · Wtan θs (5) Here, when the aperture of the acousto-optic deflector is D (m),
The blurring angle δθd of the diffraction due to the aperture is δθd = λ / D (6)

The blur angle δθd obtained from the above determines the sweep resolution of the acousto-optic deflector due to the diffraction limit.
Therefore, in order not to impair the diffraction limit resolution, the blur angle δθ at the point X in FIG. 4 must be smaller than δθd. That is, the conditional expression represented by δθ ≦ k · δθd (7) is obtained. Note that k is a constant coefficient of 1. By substituting the equations (5) and (6) into the equation (7), the condition of the width W of the ultrasonic wave front in the optical axis direction is obtained. W ≦ (k · V ² ) / (D · Atan θs) (8) Note that k ≦ 1 is a constant coefficient.

Here, θs is θs = θ−θ from the equation (2).
Since it is in and the condition becomes the worst when the deflection angle θ is the maximum, the maximum deflection angle can be set to θmax, and the equation (8) can be converted into the equation (9). W ≦ (k · V ² ) / (D · Atan (θmax−θin)) (9)

That is, the conditional expression of the width W in the optical axis direction of the ultrasonic wave front in the acousto-optic deflector is obtained so as not to impair the diffraction limit resolution in the scanning optical system using the acousto-optic deflector. Become.

Therefore, in this embodiment, the acousto-optic deflection element 2 is used.
3 is configured similarly to the acousto-optic deflecting element shown in FIG. 3, the width W of the acoustic transducer 13 is set so as to satisfy the above expression (9).

In the equation (9), a constant coefficient of k ≦ 1 is used because in a diffraction-limited imaging optical system, if the wavefront aberration is suppressed to λ / 4 or less, the theoretical diffraction limit is set. It is known that resolution is not impaired. From this point of view, it suffices to set the constant coefficient k in the equation (7) to 1/4. However, in a microscope including a large number of lenses, it is general to set the surface aberration of each optical element to λ / 10 or less. From this viewpoint, the constant coefficient k is 1/10.
Will be set to. However, if the width of the acoustic transducer is reduced, the diffraction efficiency of the acousto-optic deflecting element is reduced as described above, so the constant coefficient k is 1/10 to 1
It is desirable to set it appropriately between the intervals.

In the present embodiment configured as described above, the sweep oscillator 32 outputs a frequency sweep waveform to the acousto-optic deflection element 23 in synchronization with the horizontal drive signal. As a result, the light beam passing through the acousto-optic deflecting element 23 is deflected. Further, the galvano driver 33 swings the galvano mirror 27 in synchronization with the vertical synchronizing signal. As a result, the light beam is deflected by the galvanometer mirror 27. In this way, the sample surface is raster-scanned by the minute spots formed on the sample 30, and the sample reflected light is converted into a photoelectric signal by the photodiode 36 and input to the video monitor 38. On the video monitor 38, a scanning microscope image is formed by imaging the photoelectric signal based on the composite synchronizing signal.

The cylindrical lens 24 arranged so as to face the acousto-optic deflecting element 23 functions to correct the cylindrical effect of the acousto-optic deflecting element 23 by inputting the sweep frequency.

In the present embodiment, the width of the acoustic transducer for generating ultrasonic waves is kept small so that the width W of the ultrasonic wave front in the acousto-optic deflecting element 23 in the optical axis direction satisfies the expression (9). Therefore, a minute spot on the sample surface can be projected as small as the diffraction limit determined by the NA of the objective lens 29, and at the same time, the pinhole 3 of the detection optical system.
5 is a spot projected onto the spot projection lens 34.
It is possible to project as small as the diffraction limit determined by the NA.

This is described in the document "CONFOCAL MICROSCOPY" T.
As described in Wilson Academic Press, 1990 P95, make the pinhole diameter about 1 / of the spot diameter on the pinhole.
By setting it to 8 or less, it means that a perfect confocal state can be realized and the theoretical resolution of the confocal microscope can be realized. In addition, regarding the acousto-optic deflecting element which has been conventionally used, the result of trial calculation by the equation (9) is shown below.

A horizontal synchronizing signal (T = 63.5) at NTSC video rate is used by using a transverse ultrasonic wave in tellurium dioxide single crystal as an acousto-optic deflecting element which has been conventionally used.
When the width W of the ultrasonic wave front in the optical axis direction when the light is deflected in synchronization with 6 μsec) is calculated from the equation (9), if k = 1 and θin = θmax / 2, then W ≦ 650 ² /3.5×10 ^-3 / (50 × 10 ⁶ /63.56×10 ⁻⁶ ) / tan (5.168 / 2) = 0.0034 (m). That is, the width W of the ultrasonic wave front must be 3.4 mm or less.

When an element with an opening width of 10 mm using transverse ultrasonic waves in tellurium dioxide single crystal is used as the acousto-optic deflecting element, the opening width D is 10 mm, so the width W
Should be even smaller, specifically 1.2 mm or less. That is, the width W of the ultrasonic wave front in the optical axis direction increases as the size of the high-resolution acousto-optical deflector increases.
Needs to be small.

As described above, the width W of the ultrasonic wave front in the conventional acousto-optic deflector in the direction of the optical axis is usually about 5 mm, and the condition of the width W obtained from the equation (9) is N as a trial calculation.
Assuming scanning at the TSC video rate, W ≦ 3.4 mm for an element with the maximum resolution point 250 of the specifications and the aperture width of 3.5 mm, and W ≦ 3.4 mm with the maximum resolution point of 750 of the specifications. In the element, W ≦ 1.2 mm.
That is, it is considered that the conventional acousto-optic deflector does not satisfy the expression (9).

As described above, according to this embodiment, as the acousto-optic deflecting element 23 for deflecting the light beam from the light source, the one satisfying the above expression (9) is used, which corresponds to the video rate. A scanning optical microscope having the theoretical resolution of a confocal microscope can be realized even during high-speed scanning.

[0041]

As described above in detail, according to the present invention, it is possible to perform high-speed scanning corresponding to a video rate by using an acousto-optic deflecting element, and to achieve the scanning resolution of a confocal microscope. A microscope can be provided.

[Brief description of drawings]

FIG. 1 is a configuration diagram of a scanning optical microscope according to an embodiment of the present invention.

FIG. 2 is a configuration diagram of a scanning optical microscope using a conventional acousto-optic deflector.

FIG. 3 is an external view of an acousto-optic deflector.

FIG. 4 is a diagram showing a deflection state of a light beam by an acousto-optic deflection element.

[Explanation of symbols]

20 ... Laser light source, 23 ... Acousto-optic deflection element, 27 ... Galvano mirror, 29 ... Objective lens, 30 ... Sample, 34 ...
Spot projection lens, 35 ... pinhole, 36 ... photodiode, 38 ... video monitor.

Claims (1)

[Claims] 1. A sample is scanned by causing an ultrasonic wave generated by applying a high-frequency sweep signal to an acoustic transducer to propagate to an acousto-optic deflecting element through which a light beam from a light source passes, thereby scanning the sample. Then, in the scanning optical microscope to obtain the optical information of the sample by converting the object light from the sample into an electric signal, the angle formed by the incident light beam and the ultrasonic wave front in the acousto-optic deflecting element is θin, The velocity of sound in the optical deflecting element is V, the opening width of the acousto-optical deflecting element is D, the maximum deflection angle of the light beam in the acousto-optical deflecting element is θ max, the frequency time change rate of the high frequency sweep signal is A, and the coefficient is Assuming that the ultrasonic wave has a width W in the direction of the optical axis of the ultrasonic wave, W ≦ (k · V ² ) / (DA * tan ((theta) max- (theta) in)), It is set so that it may be set, The scanning optical microscope characterized by the above-mentioned.