JPH10332570A - Measuring apparatus for adhesion force of very small object - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an apparatus which can measure the adhesion force per unit area of a very small object which is stuck to an arbitrary face. SOLUTION: An apparatus comprises an optical microscope unit and a measuring unit. The measuring unit is provided with a probe 28 which is used to strip a cell 26 from a laboratory dish 24, with a displacement sensor 30 which is used to detect the displacement in the vertical direction of the tip of the probe and with a displacement sensor 32 which is used to detect its displacement in the horizontal direction. The optical microscope unit is provided with an objective lens 18, with an imaging element 22 and with an optical system which generates an evanescent field near the bottom face of the laboratory dish 24. The optical system is provided with a laser 301, with an optical fiber 303 which guides its radiant light, with a colimating lens 226 which changes the light into parallel light and with a prism 236 by which the parallel light is incident on the bottom face of the laboratory dish 24 at an angle exceeding a critical angle. The laboratory dish 24 in which the cell 26 is cultured is placed on the prism 236 via a coupling oil 306.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an apparatus and a method for measuring the adhesion of a microscopic object which measures the adhesion of a microscopic object adhered to a substrate or the like.

[0002]

2. Description of the Related Art In recent years, for example, in the medical field, measurement of bioaffinity of artificial bone made of titanium material or dental material made of ceramic material, and measurement of bonding strength between different materials in the industrial field have been performed. A measuring device is needed.

In particular, when measuring the adhesive force of a plurality of cells cultured on a substrate, the substrate is centrifuged, and then the mass of the cells, the centrifugal force applied to the cells, and the amount of cells detached from the substrate are measured. By applying statistical processing to the ratio of the number of cells and the number of cells remaining on the substrate, the adhesion of the cells to the substrate is measured.

[0004]

According to this prior art, however, although the total adhesion of a plurality of cells can be evaluated, the adhesion of individual cells cannot be measured independently. The present applicant has filed Japanese Patent Application No. 7-304210.
Japanese Patent Laid-Open Publication No. H11-157210 proposes an apparatus for measuring the adhesive strength of a microscopic body, which measures the adhesiveness of microscopic bodies such as individual cells adhered to a substrate or the like. This device uses a cantilever-like elastic member with displacement detection capability to apply a force to the sample cell in the lateral direction under a constant load value, measures the amount of deflection of the spring, and changes the cell over time. It measures the adhesive force per unit.

However, this device can measure the adhesive force of one cell, but cannot measure the adhesive force per unit area of a cell. SUMMARY OF THE INVENTION It is an object of the present invention to provide a microscopic body adhesion measuring device capable of measuring the adhesion per unit area of a desired microscopic body adhering to an arbitrary surface.

[0006]

SUMMARY OF THE INVENTION The present invention relates to a micro-adhesive force measuring apparatus for measuring the adhesive force of a micro-body adhering to an arbitrary surface, and a probe which is configured to be vertically and horizontally displaceable. And a first displacement sensor that optically detects the vertical displacement of the tip of the probe, and moves the probe in a horizontal direction relative to the surface to adhere to the surface. A means for pressing the tip of the probe only to a desired micro-object out of a plurality of micro-objects, and a horizontal initial displacement amount of the probe generated when the desired micro-object is pressed against the tip of the probe, And a second displacement sensor that optically detects a maximum deformation amount in the horizontal direction of the probe generated when the desired minute body is peeled off from the surface by the tip of the probe, and the initial deformation amount and the maximum Means for measuring the adhesive force of the desired minute body adhering to the surface by performing a difference operation between the shape amount, a means for generating an evanescent field near the surface, and an evanescent field. Image processing means for measuring a contact area of the minute body with the surface using fluorescence microscopy using excitation light.

Further, according to another aspect of the present invention, there is provided a micro-adhesive force measuring apparatus according to the present invention, wherein the probe is configured to be vertically and horizontally displaceable, and the tip of the probe is optically displaced in a vertical direction. The first displacement sensor to be detected and the probe are moved in a horizontal direction relatively to the surface, and among the plurality of minute bodies attached to the surface,
Means for pressing the tip of the probe only to the desired microscopic object, and the initial displacement of the probe in the horizontal direction generated when the desired microscopic object is pressed against the distal end of the probe,
And a second displacement sensor that optically detects a maximum deformation amount in the horizontal direction of the probe generated when the desired minute body is peeled off from the surface by the tip of the probe, and the initial deformation amount and the maximum Means for measuring the adhesive force of the desired minute body adhered to the surface by performing a difference operation between the deformation amount and a means for generating an evanescent field near the surface; Image processing means for measuring a contact area of the minute body with the surface based on the scattered light.

[0008] A minute body adhesion measuring apparatus according to the present invention based on yet another principal object, wherein the probe is configured to be vertically and horizontally displaceable, displacement detecting means for detecting the displacement of the probe, and the surface Means for measuring the contact area of the minute body with respect to, the unit area of the minute body based on the horizontal displacement of the probe caused by the probe pressing the minute body and the contact area of the minute body It is characterized in that the contact force per contact is measured.

[0009]

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a perspective view showing an apparatus for measuring the adhesion of a microscopic object according to an embodiment of the present invention. FIG. 1 shows the configuration of a microscopic body adhesion measuring device according to the present embodiment.

As shown in FIG. 1, the micro-adhesive force measuring device includes a stage for mounting an object to be measured, an optical microscope unit disposed below the mirror 2, and an optical microscope unit disposed above the mirror 2. And a measurement unit arranged.

The stage is a coarse XY fixed to the mirror body 2.
It has a stage 14 and a fine XY stage 50 provided on the coarse XY stage 14. Coarse motion XY stage 1
4 is in the horizontal plane (XY) by operating the coarse movement XY knob 12.
The fine movement XY stage 50 is finely moved in a horizontal plane (XY plane) under the control of the stage drive circuit 64.

The fine XY stage 50 is provided with a prism 236 of an optical system for generating an evanescent field, which will be described later, on which a petri dish 24 in which cells 26 to be measured are cultured is coupled via a coupling oil 306. Put on. The petri dish 24 is formed of optically transparent glass or plastic.

The measuring unit includes an L-shaped probe 28 for pressing and peeling off desired cells 26 attached to the bottom surface (more precisely, the upper surface of the bottom) of the Petri dish 24,
8, a vertical displacement sensor 30 for optically detecting the vertical displacement of the L-shaped tip, and a horizontal displacement optical sensor for optically detecting the horizontal displacement of the probe 28 generated when the cells 26 are peeled off from the bottom of the petri dish 24. And a direction displacement sensor 32.

The probe 28 is an L-shaped cantilever made of a leaf spring made of stainless steel foil, and the pressing portion 28a has a triangular pyramid-shaped diamond probe or silicon formed integrally with the probe 28 by a semiconductor process. Probe made of silicon oxide, silicon oxide, silicon nitride, or the like is applied. The L-shaped cantilever has a length of 15 to 30 mm and a thickness of 50 to 80 μ
m and a width of 1 mm are preferably satisfied, but the values are not particularly limited as long as they are used for the same purpose as the present invention.

The probe 28, the vertical displacement sensor 30, and the horizontal displacement sensor 32 are all supported by a fine Z stage 34, and the fine Z stage 34 is a coarse Z stage 3.
6 is supported by the mirror body 2.

The coarse movement Z stage 36 moves the fine movement Z stage 34 in the vertical direction (Z direction) by operating the coarse movement Z knob 38.
The probe 28, the vertical displacement sensor 30 and the horizontal displacement sensor 32
4 in the vertical direction.

The fine movement Z stage 34 includes a piezoelectric actuator (not shown). The piezoelectric actuator is driven by a drive circuit 44, and the drive circuit 44 is controlled based on a control signal supplied from a computer 40 via a feedback circuit 42. Driven. Fine movement Z stage 34
By driving the piezoelectric actuator inside, the probe 28, the vertical displacement sensor 30, and the horizontal displacement sensor 32 are both vertically finely moved vertically with respect to the bottom surface of the petri dish 24.

As shown in FIG. 2, the vertical displacement sensor 30 has an emission end face of the vertical displacement measuring laser beam opposed to the L-shaped tip 28b of the probe 28. The laser light emitted from the emission end face of the vertical displacement sensor 30 is reflected by the L-shaped tip portion 28b of the probe 28, and the reflected light is again captured by the vertical displacement sensor 30, and as shown in FIG. The signal is converted into a predetermined displacement signal by the vertical displacement detection circuit 46 and output to the computer 40 via the feedback circuit 42.

On the other hand, as shown in FIG. 2, the horizontal displacement sensor 32 is disposed such that the emission end face of the laser beam for measuring horizontal displacement is opposed to the vertical extension 28c of the probe 28.
The laser light emitted from the horizontal displacement sensor 32 is reflected by the vertical extension portion 28c of the probe 28, and the reflected light is again captured by the horizontal displacement sensor 32, and then, as shown in FIG. The signal is converted into a predetermined displacement signal by 48 and output to the computer 40.

The feedback circuit 42 can perform feedback control so that the displacement signal output from the vertical displacement detection circuit 46 always has a constant value. The control value of the feedback control can be appropriately adjusted by the computer 40.

The computer 40 further includes a probe 28
The displacement amount in the vertical direction and the displacement amount in the horizontal direction of the probe 28 can be calculated and displayed on the monitor based on the respective displacement signals that change according to the displacement in the vertical and horizontal directions. I have.

The optical microscope unit includes an objective lens 18 for taking in observation light, a mirror 19 for deflecting observation light from the objective lens 18, and an image sensor 22 for taking an image with the observation light.
And a monitor 20 for displaying an image obtained by the image sensor 22
And

The optical microscope unit is provided with an optical system for epi-illumination. The epi-illumination optical system is arranged on an optical path as needed to guide illumination light to an objective lens 18 and a light source 16 for emitting illumination light. The prism 304 is provided. For fluorescence observation using this epi-illumination optical system, a filter 17 that selectively transmits light (excitation light) in a specific wavelength band from illumination light and a filter 305 that selectively transmits fluorescence are provided. I have.

Further, the optical microscope unit has an optical system for generating an evanescent field near the contact surface between the petri dish 24 and the cell 26, and this evanescent field generating optical system is driven by a laser driving circuit 302. A laser 301, an optical fiber 303 for guiding light from the laser 301, a collimating lens 226 for converting laser light emitted from the optical fiber 303 into parallel light,
The parallel light from the collimating lens 226 is
A prism 236 that is incident on the bottom surface at an angle exceeding the critical angle.

Next, regarding the measurement of the adhesive force of the cell 26,
This will be described with reference to FIGS. 1, 3, and 4. In the description that follows,
It is assumed that a predetermined culture solution (for example, water) is stored in the petri dish 24, and cells 26 (microscopic bodies) cultured by the culture solution adhere to the bottom surface of the petri dish 24.

First, the coarse movement XY knob 12 is operated to move the relative positions of the petri dish 24 and the probe 28 in the X and Y directions, and the coarse movement Z knob 38 is operated to operate the vertical displacement sensor 30. Then, the probe 28 together with the horizontal displacement sensor 32 is positioned above and in the vicinity of the desired cell 26 in the petri dish 24 (see FIG. 3A).

Thereafter, the pressing portion 28a of the probe 28 is brought into contact with or close to the bottom surface of the petri dish 24 while driving the fine movement Z stage 34 to slightly move the L-shaped tip of the probe 28. At this time, the L-shaped tip 2 of the probe 28 taken from the vertical displacement sensor 30
8b, by detecting the change in the amount of reflected light from
Pressing portion 28 of probe 28 against bottom surface of petri dish 24
The degree of contact or proximity of a is optically confirmed. In this case, the acting force (contact force or proximity force) acting between the pressing portion 28a and the bottom surface of the petri dish 24 is feedback controlled so as to be weaker than the adhesive force of the cells 26 (FIG. 3).
(B)).

Next, the pressing portion 2 of the probe 28
The fine movement XY stage 5 via the drive circuit 64 while performing feedback control of the fine movement Z stage 34 so that the positional relationship between the fine movement XY stage 5a and the bottom surface of the petri dish 24 is maintained constant.
0, the fine movement XY stage 5
The petri dish 24 placed at 0 is moved in the direction of the arrow S in the figure (see FIG. 3B).

It is also conceivable to perform measurement in a state where no acting force acts between the pressing portion 28a and the bottom surface of the petri dish 24.
This is related to the size of the cell 26, but is a case where the cell 26 is sufficiently higher than the pressing portion 28a in the height direction from the bottom surface of the petri dish 24.

In such a case, since the above-described feedback control cannot be performed, the bottom surface of the petri dish 24 and the pressing portion 28a
It is necessary to check the absolute position in the height direction, and perform the following operation.

After the pressing portion 28a is brought close to the position where the pressing portion 28a contacts the bottom surface of the petri dish 24, control is performed so that the pressing portion 28a is separated from the bottom surface of the petri dish 24 by an amount corresponding to the height of the cells 26 from this position. Then, measurement is performed while maintaining this positional relationship.

In the subsequent processes from this state,
Probe 28 captured from horizontal displacement sensor 32
Changes in the amount of reflected light from the vertical extension 28c of
The deformation state of the vertical extension 28c is optically detected.

First, before the cell 26 comes into contact with the pressing portion 28a of the probe 28, and when the pressing portion 28a of the probe 28 is in contact with the bottom surface of the petri dish 24, the pressing portion 28a
a has a constant frictional force from the bottom of the Petri dish 24 (FIG. 4).
, The vertical extension 28c of the probe 28 is maintained in a horizontally deformed state corresponding to the frictional force B (see the reference A in FIG. 4). In this state, the deformation state of the vertically extending portion 28c of the probe 28 is referred to as an initial deformation amount.

A probe 28 is provided on the bottom of the Petri dish 24.
No frictional force B is generated from the bottom surface of the petri dish 24 when the pressing portion 28a is out of contact (the initial deformation amount becomes zero).
However, it is conceivable that an indefinite amount of frictional force B (noise) is generated due to external factors based on the viscosity of the culture solution and the like. Such an external factor is also considered when the pressing portion 28a is in contact with the bottom surface.

Subsequently, the petri dish 24 is further moved in the direction of the arrow S in the figure, and the cell 2 is placed on the pressing portion 28a of the probe 28.
When the contact member 6 is brought into contact, a pressing force corresponding to the adhesive force of the cell 26 acts on the pressing portion 28a. At this time, probe 2
When the eight L-shaped tip portions 28b are pressed in the horizontal direction, the vertically extending portions 28c start to be further deformed from the above-mentioned initial deformation amount. At the same time, the cell 26 starts to deform due to the reaction of the probe 28 (FIG. 3 (c)).
reference).

Then, the Petri dish 24 (that is, the cell 26)
(Ie, the output signal level of the horizontal displacement sensor 32) of the vertical extension 28c of the probe 28 with the movement of the probe 28.
Increase (see the symbol C in FIG. 4).

When the petri dish 24 (that is, the cell 26) is further moved, when the reaction force of the probe 28 corresponding to the amount of deformation of the vertical extension 28c exceeds the adhesive force of the cell 26 (the vertical extension When the deformation state of 28c reaches the maximum deformation amount), the cells 26 are peeled off from the bottom surface of the petri dish 24 (see FIG. 3D).

At this time, the probe 2 having the maximum deformation amount
8 returns to the initial amount of deformation, the output signal level of the horizontal displacement sensor 32 decreases from its maximum value to the initial value (see the symbol D in FIG. 4), and the original frictional force B
It is only the value.

The value obtained by calculating the frictional force B from the maximum value of the output signal, that is, the value obtained by calculating the initial deformation amount from the maximum deformation amount of the vertically extending portion 28c of the probe 28 (see reference symbol E in FIG. 4) is obtained. , Is detected as the adhesive force of the cells 26.

Since the relationship between the amount of deformation of the probe 28 and the amount of movement of the cell 26 shown in FIG. 4 is a relationship when the cell 26 is relatively hard, the output of the horizontal displacement sensor 32 The level drops at a stroke from its maximum value to its initial value (see the symbol D in FIG. 4), but may show a different relationship depending on the viscosity and elasticity of the cell 26. For example,
If the cell 26 is highly viscous, the output level of the horizontal displacement sensor 32 does not drop to its initial value until it is completely removed from the petri dish 24, but gradually drops while having a slope.

By the above operation, the adhesive force of the cells 26 to the bottom of the petri dish 24 is obtained. In order to measure the adhesive force per unit area of the cell 26, the cell 2 described above was used.
In parallel with the measurement of the adhesive force of No. 6, the contact area of the cell 26 with the bottom surface of the petri dish 24 is measured.

The measurement of the contact area of the cell 26 is performed using an evanescent field generating optical system. The laser light emitted from the laser 301 reaches the prism 236 via the optical fiber 303 and the collimating lens 226, is deflected by the prism 236, and is deflected by the coupling oil 30.
After passing through 6, the light enters the bottom of the petri dish 204 at an angle exceeding the critical angle. The prism 236 and the lower surface of the petri dish 204 are optically well coupled by the coupling oil 306, and the optical coupling is maintained without being affected by vibration applied to the apparatus.

Light incident at an angle exceeding the critical angle is totally reflected by the bottom surface of the petri dish 204. As a result, the petri dish 20
An evanescent field is generated near the bottom surface of No. 4. The evanescent field is an electromagnetic wave localized near the interface where total reflection occurs (about 10 nm region), and does not propagate in free space unlike a normal electromagnetic wave. When the evanescent field physically contacts the outside (for example, when the pressing portion 28a enters the evanescent field), the evanescent field becomes scattered light and propagates in free space.

Hereinafter, the measurement of the contact area of the cells 26 stained with the fluorescent dye will be described first, and then the measurement of the contact area of the cells 26 not stained with the fluorescent dye will be described. In the measurement of the contact area of the cell 26 stained with the fluorescent dye, the cell membrane on the surface of the cell 26 is previously dyed with the fluorescent dye. Staining can be carried out by attaching a dye directly to the cell membrane or by using GFP (Green Fluorescent Protein) on the cell membrane.
), A method of introducing a gene that can be found by a fluorescent protein. The wavelength of the laser beam is selected according to the excitation line of the fluorescent dye. For example, in the case of GFP S65T, the excitation line is 489 nm. GFP is a fluorescent substance that emits green light when excited by blue light.

Since the evanescent field is localized only in a region within about 10 nm from the bottom of the petri dish 24, the stained cells 26 emit fluorescence only at the portion substantially in contact with the bottom of the petri dish 24, The part away from the bottom of the petri dish 24 does not emit fluorescence. Since little light seeps into the sample side, background noise such as autofluorescence is also small.

For example, in the cell 26 shown in FIG. 5 (a), the portion which is in direct contact with the petri dish bottom surface 24a and which is shown by oblique lines in FIG. The portion where there is a gap with the petri dish bottom surface 24a does not emit fluorescence. Since the intensity of the fluorescence along the dashed line in FIG. 5 (b) is shown in FIG. 5 (c), the contact / non-contact of the sample 26 with the petri dish bottom surface 24a is determined by comparison with an appropriately set threshold value. it can.

In FIG. 1, the fluorescent light captured by the objective lens 18 forms an image on the light receiving surface of the image sensor 22. Since the excitation light is incident on the bottom surface of the petri dish 24 at an angle exceeding the critical angle, the reflected light does not enter the objective lens 18, and undesired light other than fluorescent light is filtered by the filter 305 disposed on the optical path. Since the light is absorbed, a good image is formed on the image sensor 22 by the fluorescence.

The image obtained by the image sensor 22 is fetched into the image memory 307, and the image arithmetic circuit 309 calculates the number of pixels having an intensity exceeding the threshold value for the image fetched into the image memory 307. And sends it to the host computer 40. In the host computer 40, the number of pixels sent from the image operation circuit
The contact area of the sample 26 with the petri dish bottom surface 24a is determined by multiplying the actual area per pixel determined by the relationship between the magnification of 8, the intermediate magnification, and the area of one pixel of the image sensor 22.

On the other hand, for unstained cells (in other words, cells that cannot be stained) 26, the measurement of the contact area is performed based on the scattered light generated as a result of the evanescent field being scattered by the cells 26. The evanescent field is greatly scattered by the contour of the region where the bottom surface of the petri dish 24 and the cell 26 are in contact (the boundary between contact and non-contact). The evanescent field is near the bottom of the Petri dish 24 (10
(a region of about nm), the cell portion existing near the bottom surface of the petri dish 24 physically contacts the evanescent field and becomes scattered light. That is, the scattered light becomes stronger at the outline of the contact area between the bottom surface of the petri dish and the cells.

In the contact area between the bottom of the petri dish and the cell, the evanescent field permeates (propagates) inside the cell,
It is thought that it is localized on the surface of the cell on the probe 28 side.
Hereinafter, in order to simplify the description of the present embodiment, it is assumed that scattered light is generated only at the contour of the contact area between the bottom surface of the petri dish and the cells.

For example, in the cell 26 shown in FIG. 6A, the evanescent field is scattered at the edge of the portion directly in contact with the petri dish bottom 24a. In this case, scattered light is generated only on the contour of the Petri dish bottom 24a and the cell 26. The intensity of the scattered light along the alternate long and short dash line in FIG. 6B is shown in FIG. 6C.
The edge of the contact area between the sample 4a and the sample 26 can be determined.

In FIG. 1, the scattered light captured by the objective lens 18 forms an image on the light receiving surface of the image sensor 22. Since the excitation light enters the bottom surface of the petri dish 24 at an angle exceeding the critical angle, the reflected light does not enter the objective lens 18, so that a good image is formed on the image pickup device 22 by the fluorescent light.

The image obtained by the image sensor 22 is fetched into the image memory 307, and the image calculation circuit 309 sets the image fetched into the image memory 307 within the contour of the closed curve showing the intensity exceeding the threshold value. The number of pixels included is calculated and sent to the host computer 40. The host computer 40 multiplies the number of pixels sent from the image operation circuit 309 by the actual area per pixel determined by the relationship between the magnification of the objective lens 18, the intermediate magnification, and the area of one pixel of the image sensor 22. As a result, the petri dish bottom 24
The contact area of the sample 26 with a is obtained.

In this way, the adhesive force in the shear elastic direction acting between the bottom surface of the Petri dish 24 and the cell 26 and the interlocking force change in both the stainable sample and the unstainable sample. The contact area between the bottom surface of the Petri dish 24 and the cell 26 can be measured simultaneously. As a result, important clues can be obtained in analyzing the mechanism of the attachment of the minute object to the substrate or the like.

The present invention is not limited to the above embodiment. Although various changes and improvements can be made to the above-described embodiment, all the embodiments performed without departing from the gist of the invention are included in the present invention.

For example, in the measurement of stained cells, an optical mechanism, generally called a chopper, for alternately blocking and transmitting light is provided on the optical path of the excitation light for generating an evanescent field, and measures are taken to delay the fading of the cells. Can also.

[0057]

According to the present invention, the adhesive force in the shear elastic direction acting between the substrate and the cell and the contact area between the substrate and the sample are simultaneously measured, whereby the amount of the adhesive force per unit area is measured. Can be regarded as a time change. In addition to the shear elasticity measurement, it can also be used for evaluation of the bonding force between the substrate and the sample prior to the measurement.

[Brief description of the drawings]

FIG. 1 shows a configuration of an apparatus for measuring the adhesion of a microscopic object according to an embodiment of the present invention.

FIG. 2 shows a positional relationship between a vertical displacement sensor, a horizontal displacement sensor, and a probe in the apparatus shown in FIG.

3A and 3B are explanatory diagrams of an operation of measuring an adhesive force by the apparatus of FIG. 1, wherein FIG. 3A shows a state in which a probe is positioned near a cell, and FIG. (C) shows a state where the cells are brought into contact with the pressing portion of the probe, and (d) shows a state where the cells are peeled off from the bottom surface of the petri dish.

FIG. 4 shows a relationship between the amount of probe deformation and the amount of cell movement in the state shown in FIG.

5A and 5B are explanatory diagrams of measurement of a contact area of a stained cell with a petri dish bottom, where FIG. 5A shows a cross section of a cell to be measured, FIG. 5B shows a state of the cell viewed from below, and FIG. )
(B) shows the intensity distribution of the fluorescence along the one-dot chain line.

6A and 6B are explanatory diagrams of measurement of a contact area of unstained cells with a petri dish bottom, where FIG. 6A shows a cross section of a cell to be measured, FIG. 6B shows a state of the cell viewed from below,
(C) shows the fluorescence intensity distribution along the dashed line in (b).

[Explanation of symbols]

 Reference Signs List 18 Objective lens 22 Image sensor 28 Probe 28a Pressing part 30 Vertical displacement sensor 32 Horizontal displacement sensor 40 Host computer 48 Horizontal displacement detecting circuit 50 Fine XY stage 226 Collimating lens 236 Prism 301 Laser 303 Optical fiber 309 Image processing circuit

Claims (3)

[Claims] An apparatus for measuring the adhesion of a micro-body adhering to an arbitrary surface, comprising: a probe configured to be vertically and horizontally displaceable; and a vertical tip of the probe. A first displacement sensor that optically detects a displacement in a direction, and moving the probe in a horizontal direction relative to the surface to obtain a desired one of the plurality of minute bodies attached to the surface. Means for pressing the tip of the probe only to the microscopic object, and the initial displacement of the probe in the horizontal direction that occurs when the desired microscopic object is pressed against the distal end of the probe, and
A second displacement sensor that optically detects a maximum horizontal deformation amount of the probe generated when the desired minute body is peeled off from the surface by the tip of the probe; and an initial deformation amount and the maximum deformation amount. A means for measuring the adhesive force of the desired minute body adhering to the surface by performing a difference operation between: a means for generating an evanescent field near the surface; and an excitation light for exciting the evanescent field. And an image processing means for measuring the contact area of the minute body with the surface using fluorescence microscopy. 2. A micro-adhesive force measuring device for measuring an adhesive force of a micro-body adhering to an arbitrary surface, comprising: a probe configured to be vertically and horizontally displaceable; A first displacement sensor that optically detects a displacement in a direction, and moving the probe in a horizontal direction relative to the surface to obtain a desired one of the plurality of minute bodies attached to the surface. Means for pressing the tip of the probe only to the microscopic object, and the initial displacement of the probe in the horizontal direction that occurs when the desired microscopic object is pressed against the distal end of the probe, and
A second displacement sensor that optically detects a maximum horizontal deformation amount of the probe generated when the desired minute body is peeled off from the surface by the tip of the probe; and an initial deformation amount and the maximum deformation amount. A means for measuring the adhesive force of the desired minute body adhering to the surface by performing a difference operation between: a means for generating an evanescent field near the surface; and a scattered light of the evanescent field. And an image processing means for measuring a contact area of the minute object with the surface based on the image data. 3. A micro-adhesive force measuring device for measuring an adhering force of a micro-body adhering to an arbitrary surface, comprising: a probe configured to be vertically and horizontally displaceable; and detecting a displacement of the probe. And a means for measuring a contact area of the minute body with the surface, based on a horizontal displacement of the probe caused by the probe pressing the minute body and a contact area of the minute body. And measuring the adhesive force per unit area of the minute body.