An atomic force microscope for obtaining a force spectroscopy measurement of a sample and the corresponding method

Abstract

The invention relates to an atomic force microscope (1) for obtaining a force spectroscopy measurement of a sample comprising a head (100) which is movably connected to a mounting, wherein the head (100) comprises a receiving unit (106) configured to receive a cantilever (202) comprising a tip (203) for contacting a sample of soft material. The invention further relates to a system comprising an atomic force microscope, an assembly comprising a system with an atomic force microscope as well as a method for obtaining a force spectroscopy measurement of a sample and a method to obtain a refence position of a light beam using the atomic force microscope.

Description

[0001] An atomic force microscope for obtaining a force spectroscopy measurement of a sample and the corresponding method

[0002] The invention relates to an atomic force microscope (AFM) for obtaining a force spectroscopy measurement of a sample. The invention further relates to a corresponding method for obtaining a force spectroscopy measurement of a sample.

[0003] Advantageous embodiments of the invention are given in the corresponding dependent claims and described in the following.

[0004] A first aspect of the invention relates to an atomic force microscope for obtaining a force spectroscopy measurement of a sample. The atomic force microscope comprises a head which is movably connected to a mounting, wherein the head comprises:

[0005] - a receiving unit configured to receive a cantilever comprising a tip for contacting a sample of soft material,

[0006] - a light source for generating light and focusing optics configured to focus the light and direct a first light beam along a first longitudinal axis onto the cantilever, when the cantilever is arranged at the receiving unit,

[0007] - a mirror configured to receive a second light beam reflected by the cantilever, when the cantilever is arranged at the receiving unit,

[0008] - a detector configured to detect a position of a third light beam reflected by the mirror and generate a deflection signal indicating a deflection of the cantilever along the first longitudinal axis based on the detected position of the third light beam.

[0009] The atomic force microscope further comprises an actuator configured to move the head along the first longitudinal axis, particularly wherein the head is connected to the mounting via the actuator.

[0010] In an embodiment, the atomic force microscope is configured to obtain a force spectroscopy measurement of a sample, particularly a soft material, particularly a tissue specimen.

[0011] According to an embodiment, the AFM is configured to obtain a stiffness value of a sample, particularly a soft material, particularly a tissue specimen.

[0012] According to an embodiment, the AFM is configured to determine a nanomechanical phenotype of a sample, particularly a soft material, particularly a tissue specimen.

[0013] According to an embodiment, the AFM is configured to determine a Young’s modulus of a sample, particularly a soft material, particularly a tissue specimen.

[0014] In an embodiment, the AFM is configured to determine an adhesion of a sample, particularly a soft material, particularly a tissue specimen. In an embodiment, the AFM is configured to determine a dissipation of a sample, particularly a soft material, particularly a tissue specimen.

[0015] In an embodiment, the AFM comprises an AFM head comprising a clamping mechanism for holding an optically transparent cantilever holder. Particularly, the cantilever holder can be an optically transparent cantilever chip holder with an attached cantilever chip comprising at least one cantilever.

[0016] In an embodiment, the AFM comprises a tray for arranging a specimen holder below the AFM head, the specimen holder configured to hold a sample of soft material, particularly a tissue specimen. The tray can be a sample stage. The specimen holder can be a culture dish. The specimen holder can be a petri dish. The specimen holder can be a microplate. The specimen holder can be a 6-well plate. The specimen holder can be a specimen support.

[0017] In an embodiment, the AFM head comprises a groove of the AFM head. In an embodiment, the tray comprises a slot for arranging the cantilever holder. In an embodiment, the tray comprises the slot and is configured such that the cantilever holder can be automatically inserted into the groove of the AFM head from the slot of the tray. In an embodiment, the specimen holder is a 6-well plate, wherein one of the wells may be used to store a rinsing / cleaning solution into which the cantilever can be lowered for automatic rinsing / cleaning.

[0018] In an embodiment, the AFM comprises a first linear drive for linear movement of the AFM head, particularly the complete AFM head, relative to the tray along a vertical z-direction. In the context of the present application, the vertical z-direction is also referred to as first direction. In an embodiment, the first linear drive connects the AFM head to a mounting.

[0019] In an embodiment, the AFM comprises a second linear drive for linear movement of the AFM head, particularly the complete AFM head, relative to the tray along a horizontal x-direction. In the context of the present application, the horizontal x-direction is also referred to as second direction.

[0020] In an embodiment, the AFM comprises a third linear drive for linear movement of the tray relative to the AFM head along a horizontal y-direction, particularly perpendicular to the x- direction. In the context of the present application, the horizontal y-direction is also referred to as third direction.

[0021] According to an embodiment, the first linear drive, the second linear drive and the third linear drive each comprise a coarse drive. The coarse drive can comprise or consist of a voice coil motor. The coarse drive can comprise or consist a piezo stick slip motor. According to an embodiment, the first linear drive, the second linear drive and the third linear drive each comprise at least one fine drive. The at least one fine drive can comprise or consist of a piezo motor. The at least one fine drive can comprise or consist of a piezo LEGS motor. The at least one fine drive can comprise or consist of a friction drive.

[0022] In an embodiment, the coarse drive is configured for coarse positioning of the AFM head and specimen holder.

[0023] In an embodiment, the fine drive is configured for fine positioning of the AFM head and specimen holder.

[0024] According to an embodiment, the fine drive of the first linear drive is configured for an (automated and vibration free) approach and / or leveling of the cantilever on the sample of soft material during a force spectroscopy (stiffness) measurement of the sample of soft material (combined with a fourth linear drive).

[0025] In an embodiment, the coarse drive is configured to be de-energized during fine positioning to avoid vibrations.

[0026] In an embodiment, the fine drive and the coarse drive can be magnetically coupled at a desired position of the AFM head to avoid vibrations.

[0027] In an embodiment, the fine drive and the coarse drive can be magnetically coupled at a desired position of the or tray, particularly in case of the third linear drive, to avoid vibrations.

[0028] In an embodiment, the coarse drives vibrate to a certain extent, but the fine drives are essentially vibration free. According to an embodiment, a precision of both coarse drives and fine drives is 100 nm. According to an embodiment, a drive range of the coarse drives is 85 mm in z-direction. According to an embodiment, a drive range of the coarse drives is230 mm in x-direction. In an embodiment, a drive range of the coarse drives is 230 mm in y-direction. In an embodiment, a drive range of the fine drive is 40 mm in z-direction. In an embodiment, a drive range of the fine drive is 80 mm in x-direction. In an embodiment, a drive range of the fine drive is 80 mm in y-direction.

[0029] In an embodiment, the first linear drive comprises two fine drives, positioned on either side of the coarse drive of the first linear drive along the x-axis.

[0030] In an embodiment, the AFM comprises a fourth linear drive for linear movement of the AFM head, particularly the complete AFM head, relative to the tray along the z-direction. In an embodiment, the fourth linear drive comprises a piezo-electric motor, particularly a piezo stack actuator. In an embodiment, the travel range of the fourth linear drive along the z-axis is 38 pm. In an embodiment, the resolution of the travel range of the fourth linear drive along the z- axis is less than or equal to 1 nm. In an embodiment, the fourth linear drive is configured for movement of the cantilever during a force spectroscopy measurement, particularly an automated force spectroscopy measurement.

[0031] In an embodiment, the AFM comprises at least one, particularly two constant force magnetic springs which exert an upward force along the z-direction on the complete AFM head, counteracting the weight of the head. In an embodiment, the constant force magnetic springs are arranged and configured such that the complete AFM head moves upward along the z- direction when the first linear drive, particularly the coarse drive and / or fine drive of the first linear drive, is de-energized to move the cantilever away from the sample, particularly as a safety measure because it can prevent a collision of the cantilever with the sample in case the first linear drive is de- energized.

[0032] In an embodiment, the AFM comprises a first imaging system. The first imaging system can be arranged and configured to image the sample along the z-direction from above. According to an embodiment, the second imaging system is configured to obtain a depth map, particularly a 3D reconstruction or topography deep focus image, particularly a true color image with all pixels in focus, of sample, in particular with a z-resolution (resolution in z-direction) of about 4 pm (deep focus imaging) and an xy-resolution of about 1 pm.

[0033] In an embodiment, the AFM comprises a second imaging system for imaging the sample in the specimen holder, particularly when the specimen holder is arranged on the tray. In an embodiment, the second imaging system is arranged and configured to capture an overview image of the sample in the specimen holder, particularly when the specimen holder is arranged on the tray. In an embodiment, the first imaging system is arranged and configured to read a barcode on the specimen holder on the cantilever, and / or on the cantilever holder along the z- direction from above.

[0034] In an embodiment, the AFM comprises a third imaging system, wherein the third imaging system is an optical microscope. The third imaging system can be a bottom light microscope. In an embodiment, the third imaging system is arranged and configured to for image the cantilever from below. In an embodiment, the third imaging system is arranged and configured to for image the cantilever from below to determine the dimensions of the cantilever, particularly a length, a width and / or a height of the cantilever, for obtaining a spring constant of the cantilever. In an embodiment, the third imaging system is arranged and configured to determine the position of the tip of the cantilever. In an embodiment, the third imaging system is arranged and configured for automated beam alignment in x-, y- and z-direction.

[0035] In an embodiment, the third imaging system is configured to obtain a depth map / deep focus image of the cantilever to automatically determine the exact location of the tip in x-, y- and z- direction. In an embodiment, the third imaging system is configured to obtain a depth map / deep focus image of the cantilever to automatically determine the exact location of the tip in x-, y- and z-direction to drive the optics to this position and then align the light beam optimally in x- and y-direction. In an embodiment, the third imaging system is configured to obtain a depth map / deep focus image of the cantilever to automatically determine the exact location of the tip in x-, y- and z-direction to drive the optics to this position and then align the light beam optimally in x- and y-direction and focus the light beam on the cantilever, particularly using a z-drive.

[0036] In an embodiment, the third imaging system has a fixed position with respect to the first imaging system, such that an exact tip-to-specimen offset can be determined. In an embodiment, the third imaging system has a fixed position with respect to the second imaging system.

[0037] In an embodiment, a measurement unit including the AFM head, the first, second and third linear drives and the first, second and third imaging systems are fixed to an active vibration compensation table. In an embodiment, the measurement unit is enclosed in an acoustically isolated enclosure.

[0038] According to an embodiment, the AFM head comprises a main body comprising an upper part with a rectangular shape in cross-section perpendicular to the z-direction and a lower part having a circular shape in cross-section perpendicular to the z-direction.

[0039] According to an embodiment, the AFM head comprises a bottom plate connected to the lower part comprising a circular groove for inserting a cantilever holder holding a cantilever chip comprising at least one cantilever, particularly wherein the cantilever extends along the y- direction. In an embodiment, the cantilever holder is a transparent cantilever holder chip holder. In an embodiment, the bottom plate comprises a central through-hole, wherein the central through-hole is arranged in the groove, particularly such that a light beam can enter the transparent cantilever holder from above and be reflected by the cantilever, particularly by the cantilever on the chip, if the cantilever holder comprises or is a cantilever chip holder. In an embodiment, the groove has a tapered edge for centering the cantilever holder when the cantilever holder is inserted in the groove.

[0040] In an embodiment, three ceramic hemispheres are positioned in the groove at 120° angles around the circumference of the through-hole. In an embodiment, the cantilever holder contacts the hemispheres when it is inserted into the groove.

[0041] According to an embodiment, the AFM head comprises a light source for generating a light beam comprising a laser source or a super luminescent LED, SLED (e.g., I R, 830 nm), a fiberoptic cable, and a focusing and collimating optics (particularly with a focus length of 50 mm, and / or ray lay-length of about 150 pm). In an embodiment, the focusing and collimating optics comprise a lens. In an embodiment, the light source and the focusing and collimating optics are arranged and configured to generate and direct the light beam onto a free end of the cantilever held by the cantilever holder, when the cantilever holder is arranged at the AFM, particularly when the cantilever is inserted in the groove. The free end of the cantilever can be formed resulting from a mechanical fastening of the cantilever on a cantilever holder disclosed herein, wherein the free end of the cantilever is an end section of the cantilever distant from its mechanical fastening to the cantilever holder.

[0042] In an embodiment, the focusing and collimating optics is mounted in a holder connected to a cross-table comprising a first carriage and a second carriage.

[0043] In an embodiment, the holder and the cross-table comprise a central through-hole to receive the focusing and collimating optics.

[0044] In an embodiment, the first carriage of the cross-table is movable in the y-direction, particularly with a resolution of about 10 pm, by a first light source linear drive comprising a piezo electric drive, particularly a piezo LEGS or Piezo Friction drive.

[0045] In an embodiment, the second carriage of the cross-table is movable in the x-direction, particularly with a resolution of about 10 pm, by a second light source linear drive comprising a piezo drive, particularly a piezo LEGS or a piezo stick slip motor,

[0046] In an embodiment, the holder comprises a first plate and a second plate stacked upon each other in the z-direction, wherein the holder is tiltable by three set screws connecting the first plate and the second plate.

[0047] In an embodiment, by linear movement of the cross-table and tilting of the holder, the light beam can be aligned with the cantilever, particularly with the free end of the cantilever.

[0048] According to an embodiment, the AFM head comprises a further z-drive (third carriage) configured to move the light source and focusing / collimating optics in z-direction to optimally focus the light beam on the cantilever, particularly automatically.

[0049] According to an embodiment, the AFM head comprises a third carriage and a third light source linear drive, particularly comprising a piezo-electric motor, particularly a piezo-leg motor, configured to move the third carriage along the z-direction.

[0050] In an embodiment, by linear movement of the cross-table, particularly by the first light source linear drive, the second light source linear drive and / or the third light source linear drive, the light beam can be aligned with the cantilever, particularly with the free end of the cantilever.

[0051] According to an embodiment, the AFM comprises an optical target, particularly a grid line on a plate below the AFM head next to the specimen holder. In an embodiment, the light beam of the light source is configured to scan the gridline to automatically determine and calibrate the exact position of the AFM head in X-direction and in Y-direction. In an embodiment, the AFM is configured to automatically determine and calibrate the exact position of the AFM head in X-direction and in Y-direction in the case no cantilever holder is mounted. In an embodiment, for the determination and calibration, the light beam goes through the head, hits the grid, and the reflected light goes back into the collimator, which has a beam splitter in the optical fiber that can detect if the there is a grid line or a space. In an embodiment, the AFM is configured to automatically determine and calibrate the exact position of the AFM head in X-direction and in Y-direction in the case a cantilever holder is mounted.

[0052] In an embodiment, the AFM comprises a mirror configured to reflect the light beam reflected from the cantilever, particularly with the free end of the cantilever, onto a 4-quadrant photodiode, particularly with a pivot at a place such that light reflected in air and liquid arrive at the photodiode under the same angle. In an embodiment, the mirror is glued to a mirror frame. In an embodiment, the mirror frame comprises a recess for receiving glue.

[0053] According to an embodiment, the 4-quadrant photodiode is configured to detect the deflection of the reflected light beam during a force spectroscopy measurement. In the context of the present application, the 4-quadrant photodiode is also referred to as photodiode, can also be

[0054] According to an embodiment, the photodiode is mounted on a sledge. The sledge can be moved linearly in the x-direction by a PD linear drive comprising a piezo drive, particularly a piezo LEGS motor or a friction drive. The sledge can be connected to the PD linear drive by a connecting rod extending in the z-direction. In an embodiment, the photodiode is tiltable on the sledge by three set-screws for adjustment of its position.

[0055] In an embodiment, the mirror, particularly the mirror frame, is tiltable. In an embodiment, the mirror, particularly the mirror frame, is rotatable.

[0056] In an embodiment, the AFM head comprises a push bar movable along the z-direction by a push bar linear drive, particularly wherein the push bar linear drive comprises a piezo electric drive, particularly a piezo leg motor or a piezo stick slip motor.

[0057] In an embodiment, the push bar comprises a pin configured to engage a slot of a connecting lever, particularly such that the connecting lever is pivoted in the y-z-plane when the push bar is moved along the z-direction.

[0058] In an embodiment, the mirror frame is rotatable around an axis extending in the x-direction, wherein the axis is connected to the connecting lever, such that the connecting lever pivots the mirror frame (particularly the mirror associated to the mirror frame) around the axis when the push bar is moved along the z-direction. In an embodiment, the mirror is motorized to automatically compensate the bending drift of the cantilever to keep the reflected light beam in the optimal working range on the photodiode, particularly in z-direction.

[0059] According to an embodiment, a leg spring is arranged with the spring around the axis and the legs attached to a stump and a spring fixation, respectively, to retain the mirror frame in a fixed position, particularly to avoid oscillations.

[0060] In an embodiment, the cantilever holder clamping mechanism comprises two L-shaped clamping elements for clamping the cantilever holder in the groove of the bottom plate. In an embodiment, each clamping element is connected to a respective bolt extending along the x- direction. In an embodiment, the bolts are arranged in ball bearings fixed to the bottom plate of the main body. In an embodiment, the clamping elements comprise set-screws for adjusting their position.

[0061] According to an embodiment, each bolt is connected, particularly glued, (e.g., by means of a Loctite glue) to a respective L-shaped actuation lever in the lower part of the main body.

[0062] In an embodiment, a respective leg spring is attached to each bolt, one of the legs being attached to a stump on the respective actuation lever, the leg springs hold the actuation levers in an upper position, forcing the clamping elements in a clamping position, where they exert an upward force (in the z-direction) on the cantilever holder which fixes the cantilever holder in the groove (in the context of the present application also referred to as a recess).

[0063] In an embodiment, the push bar engages the actuation levers when it is moved to a lower position along the z-direction and pushes the actuation levers into a lower position, which forces the clamping elements into an open position, where the cantilever holder can be inserted into or removed from the recess.

[0064] In an embodiment, the piezo actuator connected to the push bar has a double function: pivoting of the mirror and opening / closing of the cantilever holder clamping mechanism.

[0065] In an embodiment, the AFM comprises magnets and corresponding 3D magnetic position sensors, particularly Hall sensors, to determine the position of the respective magnets arranged on:

[0066] - the axis of the mirror frame to determine the rotary orientation of the mirror,

[0067] - the bolts connecting the clamping elements to determine the rotary orientation of the bolts, particularly indicating if the clamping elements are in the opening position or the clamping position, if a cantilever holder is inserted into the groove, and if the cantilever holder is inserted in the correct position, - the upper part of the connecting rod connecting the photodiode sledge with the PD linear drive to determine the y-position of the photodiode,

[0068] - the first carriage of the cross-table to determine its x-position, and / or

[0069] - the second carriage of the cross-table to determine its y-position.

[0070] In an embodiment, a temperature sensor is arranged at the AFM head at the bottom plate. In an embodiment, a temperature sensor is arranged at the AFM head near the groove. In an embodiment, a temperature sensor is arranged at the cantilever holder. In an embodiment, the temperature sensor is arranged at or near the cantilever holder, when the cantilever holder is inserted in the groove.

[0071] In an embodiment, the temperature sensor is configured to determine a temperature, particular the temperature of the cantilever and / or the temperature of the surrounding of the cantilever, particularly to determine the spring constant of the cantilever.

[0072] In an embodiment, the temperature sensor is arranged and configured to determine a temperature of the sample. In an embodiment, the temperature sensor is arranged and configured to determine a temperature of a surrounding of the sample, particularly of a buffer surrounding the sample.

[0073] In an embodiment, the AFM is configured to determine the temperature measurement over time. In an embodiment, the AFM is configured to determine the temperature measurement repeatedly. In an embodiment, the AFM is configured to determine the temperature measurement continuously.

[0074] In an embodiment, the AFM comprises a temperature control device configured to adapt the temperature if the measured temperature leaves a pre-defined temperature interval.

[0075] Advantageously, the viability of the sample is determined via determination of the temperature of the sample and / or the buffer surrounding the sample.

[0076] According to an embodiment, the AFM comprises an AFM controller. The AFM controller can be made from a SoM (system on module) consisting of an FPGA (field programmable gate array) for calculations, particularly for calculations on a timescale of about 5 ns, a Real Time Linux running on ARM processors for calculations, particularly for calculations on a timescale of about 50 ps, and a device controller, particularly a Linux operating system for device automation and / or user input.

[0077] In an embodiment, inputs to the controllers are from and / or outputs from the controllers are to: the photodiode (force spectroscopy photodiode), particularly via a pre-amplifier and an analog-digital-converter, - a head referencing photodiode,

[0078] - the first imaging system,

[0079] - the second imaging system,

[0080] - the third imaging system,

[0081] - the magnetic positioning sensor,

[0082] - an environment temperature sensor,

[0083] - a pressure sensor,

[0084] - a humidity sensor,

[0085] - a voltage sensor,

[0086] - a sample temperature sensor,

[0087] - the vibration compensation table,

[0088] - the coarse drive of the first linear drive,

[0089] - the coarse drive of the second linear drive,

[0090] - the coarse drive of the third linear drive,

[0091] - a position controller of the first linear drive,

[0092] - a position controller of the second linear drive,

[0093] - a position controller of the third linear drive,

[0094] - the fine drive (particularly piezo-electric actuator, particularly piezo LEG motor or friction drive) of the first linear drive,

[0095] - the fine drive (particularly piezo-electric actuator, particularly piezo LEG motor or friction drive) of the second linear drive,

[0096] - the fine drive (particularly piezo-electric actuator, particularly piezo LEG motor or friction drive) of the third linear drive,

[0097] - a position controller of the first linear drive,

[0098] - a position controller of the second linear drive,

[0099] - a position controller of the third linear drive,

[0100] - the piezo-electric actuator of the PD linear drive, particularly the piezo LEG and / or the friction drive of the PD linear drive,

[0101] - the piezo-electric actuator moving the push bar to rotate the mirror, particularly the piezo LEGS and / or friction drive moving the push bar to rotate the mirror,

[0102] - the piezo-electric actuator moving the push bar to actuate the clamping mechanism, particularly the piezo LEGS and / or friction drive moving the push bar to actuate the clamping mechanism,

[0103] - the piezo-electric actuator moving the push bar to rotate the mirror and to actuate the clamping mechanism, particularly the piezo LEGS and / or friction drive moving the push bar to rotate the mirror and to actuate the clamping mechanism, - the piezo-electric actuator, particularly the piezo LEGS actuator and / or the piezo stick slip motors actuating the cross-table to align the illuminating light beam from the light source onto the cantilever,

[0104] - a light source controller,

[0105] - a SLED controller,

[0106] - a controller configured to control the fourth linear drive, particularly a force spectroscopy actuator controller (piezo stack motor 140),

[0107] - a light curtain of a friction drive,

[0108] - a light curtain arranged and configured to monitor a state of the device doors (particularly system doors) for cantilever holder and specimen holder insertion.

[0109] In an embodiment, some circuit boards of the controller are positioned in a dice-like fashion (3D, several boards perpendicular to each other) around the upper part of the AFM head main body resulting in a compact architecture. In an embodiment, a plurality of circuit boards of the controller is arranged in a 3D manner perpendicular to each other around the upper part of the AFM head main body resulting in a compact architecture.

[0110] According to an embodiment, at least one of the following methods is implemented by the controller:

[0111] - automatic loading and clamping of a cantilever holder,

[0112] - automating beam alignment on the cantilever,

[0113] - automatic beam focusing on the cantilever,

[0114] - shaking the tray by the third linear drive, wherein the first and / or the second imaging system (overview top-down camera) monitors the position of the sample in the specimen holder on the tray and the controller automatically determines if the sample is tightly attached to the specimen holder and not floating, this serves as an “intake control” to determine if the sample is complete and in good shape,

[0115] - automatic determination of a force map of the sample,

[0116] - automatic collision detection, particularly to find areas where the probe cannot measure due to geometric constraints

[0117] - automated tilt detection, particularly to find areas where the sample is too steep to perform valid force spectroscopy,

[0118] - automatic determination of the spring constant of the cantilever, particularly measuring dimensions of the cantilever by the third imaging system, temperature sensor,

[0119] - automatic determination of the system’s deflection sensitivity, wherein Deflection [nN]=Deflection signal on photodiode [V] *deflection sensitivity [nm / V]*spring constant [N / m],

[0120] - automated cantilever drift compensation, - automatic force spectroscopy measurement, particularly comprising at least one of the following steps:

[0121] • automatic selection of a measurement spot on the sample, particularly on the sample of soft material specimen, and selection of e.g., 20x20 measurement points on the measurement spot,

[0122] • automatic levelling of the cantilever tip on the sample of soft material,

[0123] • automatic ramp length optimization of the force spectroscopy measurement,

[0124] - tip-safe mechanism, particularly wherein the AFM head is automatically moved upwards in the z-direction away from the sample of soft material upon detection of vibrations or obstacles, particularly wherein the vibration and / or the obstacle is recognized by the first imaging system, the second imaging system, the vibration compensation table, and / or a geophone,

[0125] - automatic force curve quality control,

[0126] - automatic selection of a new measurement point on the sample of soft material when quality of the automatic force curve is bad, particularly below a predefined quality measure.

[0127] In an embodiment, the particular method is hardware-implemented. In an embodiment, the particular method is software-implemented.

[0128] In an embodiment, the first imaging system is repeatedly removably connected to the mounting in fixed position with respect to the head.

[0129] In an embodiment, the second imaging system is repeatedly removably connected to the mounting in fixed position with respect to the head.

[0130] In an embodiment, the third imaging system is repeatedly removably connected to the AFM in fixed position with respect to the head.

[0131] In an embodiment, the third imaging system is repeatedly removably connected to the AFM in fixed position with respect to the first imaging system.

[0132] In an embodiment, the third imaging system is repeatedly removably connected to the AFM in fixed position with respect to the second imaging system.

[0133] In an embodiment, the first imaging system is repeatedly removably connected to the mounting in fixed position with respect to the head, and the second imaging system is repeatedly removably connected to the mounting in fixed position with respect to the head. The first imaging system, the second imaging system and the head can be fixedly coupled to each other, particularly in fixed position to each other. In an embodiment, the first imaging system is repeatedly removably connected to the mounting in fixed position with respect to the head, and the second imaging system is repeatedly removably connected to the mounting in fixed position with respect to the head, and the third imaging system is repeatedly removably connected to the AFM in fixed position with respect to the first imaging system.

[0134] Advantageously, the position of each of the first imaging system, the second imaging system, the third imaging system and / or the head relative to each other can be determined and is fixed. Advantageously, a reference position of the first imaging system, the second imaging system, the third imaging system and / or the head relative to each other can be determined.

[0135] Advantageously, an external perturbation can affect each of the first imaging system, the second imaging system, the third imaging system and / or the head equally, because they are in fixed position to each other.

[0136] A label scanner for scanning specimen and / or probe holder can be arranged and configured to:

[0137] - provide traceability for specimen holder, and / or

[0138] - provide traceability for probe.

[0139] Advantageously, this avoids a double-use of consumables. It can increase the quality of the measurement.

[0140] In an embodiment, the probe is calibrated at production site and the probe characteristics are saved in a bar code, readable by the label scanner. Advantageously, the calibration (spring constant) can be removed from the measurement procedure to safe time.

[0141] In an embodiment, cantilever width and / or length can be determined in high precision at production site to improve accuracy of spring constant measurement. The width and / or length can be saved in a bar code, readable by the label scanner.

[0142] In an embodiment, the tip height is measured at production site to ensure tip is long enough for measurement. The tip height can be saved in a bar code, readable by the label scanner.

[0143] A second aspect of the invention relates to system comprising an atomic force microscope according to the first aspect of the invention, and a vibration isolation table , wherein the vibration isolation table comprises a vibration sensor.

[0144] According to an embodiment, the system comprises an interface, particularly a humanmachine interface, comprising a drawer unit configured to receive a shuttle unit for carrying a specimen holder and / or a cantilever holder, wherein the drawer unit can be moved between a loading position away from the sample stage for placing the shuttle unit onto the drawer unit and an unloading position at the sample stage, and wherein the drawer unit comprises a mechanism for unloading, particularly for transferring the shuttle unit from the drawer unit to the tray, when the shuttle unit is placed on the drawer unit and the drawer unit is moved to the unloading position.

[0145] In an embodiment, the drawer unit comprises a plurality of protrusions for placing the shuttle unit on the protrusions of the drawer unit.

[0146] In another embodiment, the drawer unit comprises slides, particularly telescopic slides, for moving the drawer unit, particularly along the third longitudinal direction, between the loading position and the unloading position.

[0147] According to an embodiment, a height-adjustable structure is slidably supported on each slide by means of at least one pin extending from the slide into a recess of the height-adjustable structure, and wherein the height-adjustable structure comprises said protrusions, and wherein the recess is shaped such that when the drawer unit is moved in the unloading position and the height-adjustable structure engages with a stop, a vertical position of the protrusions and of the shuttle unit placed on the protrusions is lowered by moving the height-adjustable structure with respect to the at least one pin of the slide.

[0148] By reversing the movements with the above mechanism, the protrusion can be lifted to pick up the shuttle unit when it is arranged on the tray, particularly on support elements of the tray.

[0149] Particularly, the recess can have two horizontal sections that extend essentially horizontally, that is, along the third longitudinal axis, wherein the two horizontal sections are arranged on different vertical positions along the first longitudinal axis, and wherein the two horizontal sections are connected by a slope portion of the recess. As such, the pin of the slide can move from one end to another end of the recess under a vertical displacement of the height- adjustable structure with respect to the slide.

[0150] In another embodiment, the sample stage comprises a plurality of support elements, particularly three supporting elements, for supporting the shuttle unit, wherein in said unloading position, the protrusions for placing the shuttle unit are arranged above said support elements, such that the shuttle unit can be unloaded from the drawer unit to the sample stage by lowering the height-adjustable structure with respect to the slide.

[0151] According to an embodiment, the system comprises a label scanner, particularly a label scanner for scanning a bar code, particularly wherein the bar code codes information regarding the sample, the specimen holder, the probe holder and / or the cantilever.

[0152] In another embodiment, the specimen holder is a culture dish, a petri dish, a microplate, particularly a 6-well plate, or a specimen support. In yet another embodiment, the system comprises a hydraulic wheel, wherein the hydraulic wheel is arranged and configured such that the system is movable by means of the hydraulic wheel, and wherein the system comprises a fixed supporting foot, wherein the fixed supporting foot is arranged and configured such that the system is stands stable and / or can be aligned by means of the supporting foot.

[0153] A third aspect of the invention relates to an assembly comprising the system according to the second aspect of the invention and a packaging, wherein the packaging is configured to receive the system, wherein the packaging comprises a ramp configured to support the release of the system from the packaging, particularly wherein the packaging comprises a lid, wherein the lid is configured as a wedge and configured to be arrangeable such that the system is releasable from the packaging via the wedge.

[0154] A fourth aspect of the invention relates to a method for obtaining a force spectroscopy measurement of a sample, particularly a sample from a soft material, more particularly a biological tissue specimen, using the atomic force microscope according to one of the claims 1 to 24 or the system according to one of the claims 25 to 34 or the assembly according to claim 34, comprising the following steps:

[0155] - contacting the sample by the tip of the cantilever at a measurement spot,

[0156] - moving the head by the actuator towards the sample parallel to the first longitudinal axis in a loading step, particularly automatically, such that the tip of the cantilever indents into the sample,

[0157] - moving the head away from the sample by the actuator parallel to the first longitudinal axis in an unloading step, particularly automatically,

[0158] - during the loading step and / or the unloading step, determining a plurality of deflection values of the cantilever from the deflection signal of the detector,

[0159] - determining the force spectroscopy measurement of the sample at the measurement spot from the determined deflection values, particularly automatically.

[0160] In an embodiment, the method comprises determining a plurality of force spectroscopy measurements indicating the force of the cantilever on the sample at a corresponding position along the first longitudinal axis, wherein the force spectroscopy measurement of the sample is determined, particularly determined automatically, from at least a subset of the force values, particularly wherein a force-position relationship and / or a force-position curve is obtained.

[0161] In another embodiment, altering a position of the sample relative to the head, particularly relative to the cantilever, is provided by moving the head along the first longitudinal direction, particularly by the first linear drive and / or the actuator and / or along the second longitudinal direction, particularly by the second linear drive, and / or by moving the sample, particularly the specimen holder holding the sample, particularly by the third linear drive, along the third longitudinal direction.

[0162] According to an embodiment, the detector is moved by the detector linear drive along the second longitudinal axis and / or the mirror frame, particularly the associated mirror, is pivoted by the mirror drive, such that the mirror and the detector are aligned such that the third light beam reflected by the mirror meets a pre-defined detection region of the detector, particularly a linear region of the detector, particularly the detector and / or the mirror frame are moved automatically keeping the third light beam in the detection region of the detector, particularly the linear region of the detector.

[0163] In another embodiment, a shape of the cantilever, a size of the cantilever, a length of the cantilever, a width of the cantilever and / or a height of the cantilever is determined, particularly determined automatically, by the third imaging system, and / or wherein the position of the tip of the cantilever is determined, particularly determined automatically, by the third imaging system.

[0164] In another embodiment, prior to contacting the sample by the tip of the cantilever, the method comprises the step of 3D profiling of the sample using information obtained by the first imaging system, and / or using information obtained by the second imaging system, particularly information related to a deep focus image and / or a depth map, particularly using information obtained by the first imaging system, and / or using information obtained by the second imaging system and calculations, particularly wherein the calculations comprise the step of a Laplacian regression.

[0165] According to another embodiment, prior to contacting the sample by the tip of the cantilever, the method comprises the step of an automated spot selection based on sample-related information, based on information obtained by 3D profiling of the sample, based on information obtained by deep focus imaging, based on information obtained by the depth map, based on information regarding the shape of the cantilever, based on information regarding the size of the cantilever, and / or based on the position of the tip of the cantilever, providing a prediction of the quality of the measurement at the selected spot of measurement.

[0166] In yet another embodiment, prior to contacting the sample by the tip of the cantilever, a reference sample having a predefined stiffness is contacted by the tip of the cantilever, and the head is moved by the actuator over a moving distance while the tip contacts the reference sample without indentation of the reference sample, and wherein the deflection signal of the detector is obtained over the moving distance, and wherein a deflection sensitivity of the detector is determined, particularly automatically, from the moving distance and the deflection signal, and / or wherein the deflection sensitivity of the detector is determined in a contact-free manner particularly based on information obtained by a resonance curve, particularly the area below the resonance curve, wherein the resonance curve is taken in air or in liquid, and / or wherein the deflection sensitivity of the detector is determined in a depth stepping standards manner particularly wherein prior to contacting the sample by the tip of the cantilever, a surface of a standard sample comprising recesses with pre-defined depth is contacted by the tip of the cantilever, and the head is moved by the actuator over a moving distance, and wherein the deflection signal of the detector is obtained over the moving distance, and wherein a deflection sensitivity of the detector is determined, particularly automatically, from the moving distance and the deflection signal.

[0167] In another embodiment, the deflection sensitivity is determined, wherein the device detects when the deflection sensitivity leaves a predefined interval and initiates a recalibration, particularly automatically, by re-aligning the beam and executing an additional deflection sensitivity measurement.

[0168] In yet another embodiment, the cantilever is moved in the proximity of the sample or moved to the spot of measurement, particularly wherein the cantilever is moved in the proximity of the sample or moved to the spot of measurement in an approach-free manner, based on sample- related information, and / or based on information obtained by 3D profiling of the sample.

[0169] According to another embodiment, an adaptive optimization is performed to move the cantilever in the proximity of the sample or to the spot of measurement, wherein information considering a deviation between a detected previous spot of measurement and a proposed previous spot of measurement is used, particularly by applying the deviation on the proposed spot of measurement.

[0170] In an embodiment, a vibration signal is obtained from the vibration sensor, and wherein the head is moved away from the sample along the first longitudinal axis, particularly automatically, in case the signal indicates the presence of a vibration, particularly of a vibration having an amplitude or a frequency above a predefined threshold.

[0171] According to an embodiment, a spring constant of the cantilever is determined, particularly determined automatically, based on data provided by the third imaging system, particularly a length of the cantilever, a width of the cantilever and / or a thickness of the cantilever, based on the temperature value obtained by the temperature sensor, based on the environmental humidity obtained by the environmental sensor and / or the environmental pressure obtained by the environmental sensor, a resonance frequency of the cantilever, and / or a quality factor of the cantilever.

[0172] In another embodiment, the spring constant of the cantilever is detected repeatedly during the measurement and / or a value related to the deflection sensitivity calibration is detected repeatedly during the measurement, wherein, when the spring constant of the cantilever leaves a pre-defined range, the cantilever is exchanged and / or a re-calibration is executed, and / or wherein, when the value related to a deflection sensitivity calibration leaves a pre-defined range, the cantilever is exchanged and / or a re-calibration is executed, wherein the recalibration comprises a re-alignment of the first beam relative to the cantilever, particularly the free end of the cantilever, and an additional measurement of the value related to the deflection sensitivity, and / or wherein a quality value of the cantilever is detected repeatedly during the measurement, wherein, when the quality value of the cantilever leaves a pre-defined range, the cantilever is exchanged.

[0173] In yet another embodiment, the cantilever, particularly the cantilever holder, is picked up, particularly picked up from the sample stage, particularly from the second slot comprised in the sample stage for receiving the cantilever, when the head determines a pre-defined force value executed by the cantilever to be picked up, particularly the cantilever holder to be picked up.

[0174] In an embodiment, a correction factor related to a buffer surrounding the sample is determined, wherein prior to contacting the sample by the tip of the cantilever, the deflection signal is detected while the cantilever is moved along the first longitudinal axis towards the sample, wherein a position of the cantilever along the first longitudinal axis is determined when the deflection signal is lost.

[0175] According to an embodiment, the atomic force microscope, particularly the AFM controller, particularly the field programmable gate array, monitors at least one monitoring signal, wherein a monitoring signal is one of:

[0176] - a cantilever sum and the deflection signal,

[0177] - a vibration signal,

[0178] - a signal of the accelerometer,

[0179] - a signal the geophone,

[0180] - a signal obtained by a microphone, wherein the atomic force microscope:

[0181] - interrupts any motion in the atomic force microscope to protect the probe, when the monitoring signal leaves a pre-defined range,

[0182] - interrupts the force spectroscopy measurement, when the monitoring signal leaves a pre-defined range particularly in the case of external vibrations or external acoustic noise,

[0183] - interrupts an optical measurement by the first optical system and / or the second optical system and / or the third optical system, when the monitoring signal leaves a pre-defined range, particularly in case of external vibrations or external acoustic noise, and / or uses the monitoring signals as “gates” to only trigger measurements, when vibration and noise levels are low.

[0184] In yet another embodiment, the deflection sensitivity in a liquid, particularly the buffer, is determined based on the deflection sensitivity determined in air and positional information regarding the tip of the cantilever, particularly information regarding the position of the tip of the cantilever with respect to the first longitudinal direction, the second longitudinal direction and / or the third longitudinal direction.

[0185] A fifth aspect of the invention relates to a method to obtain a refence position of the light beam using the atomic force microscope according to one of the claims 1 to 24 or the system according to one of the claims 25 to 34, comprising the following steps: a. directing a first light beam along a first longitudinal axis at the first optical target, b. detecting the intensity of the first reference light beam reflected by the first optical target by the controller, c. detecting the position of the head perpendicular to the first longitudinal axis by the controller, d. taking an optical image of the first optical target by the first imaging system and / or the second imaging system, e. determining a position of:

[0186] - the head with respect to the first imaging system,

[0187] - the head with respect to the second imaging system,

[0188] - the first imaging system with respect to the second imaging system,

[0189] - the head with respect to the first light beam,

[0190] - the first imaging system with respect to the first light beam, and / or

[0191] - the second imaging system with respect to the first light beam.

[0192] Exemplary embodiments are described below in conjunction with the Figures. The Figures are appended to the claims and are accompanied by text explaining individual features of the shown embodiments and aspects of the present invention. Each individual feature shown in the Figures and / or mentioned in the text of the Figures may be incorporated (also in an isolated fashion) into a claim relating to the first aspect, the second aspect, the third aspect, the fourth aspect and / or the fifth aspect according to the present invention.

[0193] Fig. 1 shows an embodiment of an atomic force microscope according to the invention, wherein a cantilever holder is arranged on the head of the atomic force microscope; Fig. 2 shows a cut view through the head of an atomic force microscope according to an embodiment of the invention;

[0194] Fig. 3 shows an embodiment of an atomic force microscope according to the invention, with a tray for receiving a specimen holder arranged below the head of the atomic force microscope;

[0195] Figs. 4a, b schematically depict an optical path of light emitted from a light source and reflected from the cantilever and a mirror towards a detector (Fig. 4a), wherein a transparent body is arranged in the cantilever holder shown in Fig. 4b;

[0196] Fig. 5 shows a front view of a lower part of the head of an atomic force microscope according to an embodiment of the invention, to which a cantilever holder is connected;

[0197] Fig. 6 shows a side view of Fig. 5;

[0198] Fig. 7 shows a bottom view of the head of the atomic force microscope with a slot for inserting a cantilever holder;

[0199] Fig. 8 shows a bottom view of the head of the atomic force microscope with a cantilever holder inserted in the slot;

[0200] Fig. 9 shows an embodiment of a mechanism for operating clamps for attaching the cantilever holder to the head;

[0201] Figs. 10a, b shows an embodiment of an atomic force microscope according to the invention, with focusing and collimating optics arranged on an upper part of the head;

[0202] Fig. 11 shows an embodiment of an atomic force microscope according to the invention, with a third imaging system for imaging the cantilever;

[0203] Fig. 12 shows an embodiment of an atomic force microscope according to the invention, with focusing and collimating optics movable by a piezo stick slip motor

[0204] Fig. 13 illustrates an embodiment of a system according to the invention, with an atomic force microscope and an interface for controlling drawer units; Figs. 14a, b,c shows an embodiment of a system according to the invention, comprising two drawing units for loading and unloading sample holders and / or cantilever holders; and

[0205] Fig. 15 shows an embodiment of the atomic force microscope 1 according to the invention comprising a linear drive with an optical ruler for optically detecting a driving distance of the linear drive.

[0206] Fig. 1 shows an embodiment of an atomic force microscope 1 according to the invention. The atomic force microscope 1 comprises a head 100 which is movably connected to a mounting (not shown). Particularly, the head 100 is movable with respect to the mounting by an actuator 140 of the atomic force microscope 1. To this end, the head 100 can be connected to the mounting via the actuator 140. By means of the actuator 140, the head 100 can be moved along a first longitudinal axis L1 , which corresponds to the z-axis according to the coordinate system indicated in Fig. 1.

[0207] On top of an upper part 104a of a main body 104 of the head 100, the atomic force microscope 1 comprises a light source 105. The light source 105 is configured to generate light used to optically detect a displacement of a cantilever 202 attached to cantilever chip 201 of a cantilever chip holder 310 arranged at a receiving unit 106 of the atomic force microscope 1. Alternatively to the cantilever chip holder 310, the cantilever 202 can be attached to a cantilever holder. The main body 104 frames a mirror configured to receive light emitted from the light source and reflected from the cantilever as well as a detector configured to detect a position of a light beam reflected from the mirror toward the detector, as explained further in Fig. 2.

[0208] The light source 105 may comprise focusing and collimating optics, such as lenses, and is connected to the main body 104 via a cross-table 110 comprising a first carriage 110a, a second carriage 110b and a third carriage 110c.

[0209] The first carriage 110a of the cross-table 110 is movable in the y-direction according to the coordinate system indicated in Fig.1 , particularly with a resolution of about 10 pm, by a first light source linear drive 111a comprising a piezo electric drive, particularly a piezo LEGS or Piezo Friction drive or piezo stick slip motor.

[0210] The second carriage 110b of the cross-table 110 is movable in the x-direction according to the coordinate system indicated in Fig. 1 , particularly with a resolution of about 10 pm, by a second light source linear drive 111 b comprising a piezo drive, particularly a piezo LEGS or Piezo Friction drive or piezo stick slip motor or a piezo stick slip motor. The third carriage 110c of the cross-table is movable in the z-direction according to the coordinate system indicated in Fig. 1 , particularly with a resolution of about 10 pm, by a second light source linear drive comprising a piezo drivel 11c, particularly a piezo LEGS or Piezo Friction drive or piezo stick slip motor or a piezo stick slip motor.

[0211] By moving the light source 105 with respect to the head 100, particularly with respect to the cantilever 202, by means of said cross-table 110 and its carriages 110a, 110b, 110c, a spot position of a first light beam B1 emitted from the light source 105 along the first longitudinal axis L1 toward the cantilever 202 can be aligned with and focused on a free end of the cantilever 202, which allows for a strong signal indicative of the deflection of the cantilever 202 during a measurement and thus increases the measurement accuracy of the atomic force microscope 1.

[0212] Fig. 2 shows a cut view through the head 100 of an atomic force microscope 1 according to an embodiment of the invention. The cut view highlights components arranged inside the head 100, such as light and focusing optics 105a for focusing light onto the cantilever 202. From the cantilever 202, light can be reflected towards a mirror 112 mounted on a mirror frame 114 that is configured to reflect the light towards a detector 113, where the reflected light indicative for the deflection of the cantilever 202 is detected. The detector 113 in this embodiment is a 4- quadrant photodiode arranged on a detector holder 116. The head 100 comprises a detector linear drive 116a, particularly a piezo-electric motor, configured to move the detector holder 116 with the detector 113 along the x-direction, perpendicularly to the z- and the y-direction indicated in Fig. 2. To this end, the detector linear drive 116a is connected to the detector holder 116 via a connecting rod extending 1 in the z-direction. A translation movement of the connecting rod 117 caused by the detector linear drive 116 can thereby be transferred onto the detector holder 116. The detector holder 116 can be tilted by three set-screws 116b for adjusting the orientation of the detector holder 116 with respect to the incoming light beam reflected from the mirror 112. A magnet-based position sensor 126a is arranged on the photodetector holder 116, such that the position of the photodetector holder 116 with the photodetector 113 can be determined.

[0213] The mirror frame 114 is rotatable around an axis 114a extending in the x-direction, perpendicularly to the y- and the z-direction according to the coordinate system indicated in Fig. 2. A mirror drive 114b, particularly a piezo-electric motor, forms a clamping drive 1222 configured to move a means 1224 along the first longitudinal axis L1. By moving the means 1224, clamping elements 122 for clamping the cantilever chip holder 310 to the head 100 of the atomic force microscope 1 can be transferred between a receiving position in which the cantilever chip holder 310 can be arranged in the receiving unit 106 and a clamping position in which the cantilever chip holder 310 is secured at the receiving unit 106 by the clamping elements 122.

[0214] The axis 114a of the mirror frame 114 is mechanically connected to a connecting lever 121 , which in turn is mechanically connected to a push bar 119. The connecting lever 121 comprises a slot 121a and the push bar comprises a pin 120 arranged in the slot. As such, when the mirror drive 114b moves said means 1224 along the z-direction, the pin 120 moves in the slot 121 under a pivot movement of the mirror 112 on the mirror frame 114. The mirror drive 114b is thus configured to both effect the engagement of the clamping elements 122 as well as the pivoting of the mirror 112.

[0215] Fig. 3 shows an embodiment of an atomic force microscope 1 according to the invention, with a tray 300 for receiving a specimen holder 430 arranged below the head 100 of the atomic force microscope 1.

[0216] The head 100 is schematically depicted arranged on a first linear drive 101 for linear movement of the head 100 relative to the tray 300 along the vertical z-direction. The first linear drive 101 comprises a coarse drive 101a for coarse movement of the head 100 along the z-direction and a fine drive 101b for fine movement of the head 100 along the z-direction. The coarse drive 101a can comprise or consist of a voice coil motor. The fine drive 101 b can comprise or consist of a piezo motor.

[0217] The atomic force microscope according to the present embodiment comprises a constant force magnetic springs 130 which exerts an upward force along the z-direction, i.e. , away from the tray 300, on the head 100 that counteracts the weight of the head 100. The magnetic spring 130 is configured such that the head 100 moves upward along the z-direction when the first linear drive 101 , particularly the coarse drive 101a and / or the fine drive 101 b of the first linear drive 101 , is de-energized to move the cantilever 202 away from the sample. This represents a safety measure as it can prevent a collision of the cantilever 202 with the sample in case the first linear drive 101 is de-energized.

[0218] The first linear drive 101 is arranged between the head 100 and a second linear drive 102 for linear movement of the head 100 relative to the tray 300 along the x-direction, perpendicular to the y- and the z-direction. The second linear drive 102 comprises a coarse drive 102a for coarse movement of the head 100 along the x-direction and a fine drive 102b for fine movement of the head 100 along the x-direction. The coarse drive 102a can comprise or consist of a voice coil motor. The fine drive 102b can comprise or consist of a piezo motor.

[0219] As such, the head 100 can be moved with respect to the tray 300 along two orthogonal directions by means of the first and second linear drive 101 ,102. Moreover, the atomic force microscope 1 according to the present embodiment comprises a third linear drive 103 for linear movement of the tray 300 relative to the head 100 along the y- direction. The third linear drive 103 comprises a coarse drive 103a for coarse movement of the tray 300 along the y-direction and a fine drive 103b for fine movement of the tray 300 along the y-direction. The coarse drive 103a can comprise or consist of a voice coil motor. The fine drive 103b can comprise or consist of a piezo motor.

[0220] Thus, the head 100 and the tray 300 can be moved with respect to each other along three orthogonal directions using the first, second and third linear drive 101 ,102,103, which allows for precise and flexible positioning of the sample with respect to the cantilever 202 arranged on the head 100.

[0221] The atomic force microscope 1 according to the present embodiment further comprises a first imaging system 450 arranged on the first linear drive 101. The first imaging system 450 is configured to image a sample arranged on the specimen holder 430 along the z-direction from above. The first imaging system 450 is particularly configured to obtain a depth map, particularly a 3D reconstruction or topography deep focus image, particularly a true color image with all pixels in focus, of the sample. For example, a z-resolution, i.e., the optical resolution in the z-direction, can be about 4 pm and an xy-resolution, i.e., an optical resolution along the x- and the y-direction, can be about 1 pm.

[0222] As can further be seen in Fig. 3, a second imaging system 460 is arranged on the head 100. The second imaging system 460 is configured to capture an overview image of a sample arranged in the specimen holder 430. Moreover, the first imaging system 450 can be arranged and configured to read a barcode on the specimen holder, and / or on the cantilever chip holder 310 along the z-direction from above, i.e., from the perspective of the head 100.

[0223] Fig. 3 also shows two shuttle units 650 arranged on the tray 300. The shuttle units 650 are transferable between the tray 300 and a drawer unit 620 explained in the context of Fig. 14a, b,c. The shuttle units 650 are configured to receive said specimen holder 430 with a sample and / or a cantilever chip holder 310.

[0224] A third imaging system 440 is arranged on the tray 300. The third imaging system 440 is an optical microscope, for example a bottom light microscope, configured to image the cantilever 202 from below, i.e., from the perspective of the tray 300. The third imaging system 440 can be configured to determine the dimensions of the cantilever 202, particularly a length, a width and / or a height of the cantilever 202, in order to determine a spring constant of the cantilever 202. Additionally, the position of the tip of the cantilever 202 can be determined by means of the third imaging system 440. As such, the third imaging system 440 can be used for automated alignment of the light beam on the tip of the cantilever 202 in x-, y- and z-direction. The third imaging system 440 can be configured to obtain a depth map and / or deep focus image of the cantilever 202 to automatically determine the location of the tip in x-, y- and z- direction.

[0225] The atomic force microscope 1 according to the present embodiment also comprises a controller 410. The controller 410 can be made from a SoM (system on module) consisting of an FPGA (field programmable gate array) for calculations, particularly for calculations on a timescale of about 5 ns, a Real Time Linux running on ARM processors for calculations, particularly for calculations on a timescale of about 50 ps, and a device controller, particularly a Linux operating system for device automation and / or user input.

[0226] Inputs to the controller 410 can be from and / or outputs from the controller 410 can be to one or more of the following: the detector 113, particularly via a pre-amplifier and an analog-digital- converter, the first, second and third imaging system 450,460,440, the magnetic positioning sensor 126a, an environment temperature sensor, an ambient air pressure sensor, an ambient air humidity sensor, voltage sensors, a sample temperature sensor, a vibration compensation table on which the atomic force microscope 1 is arranged, the first, second and third linear drive 101 ,102,103, particularly their coarse drives 101 a, 102a, 103a and / or their fine drives 101b, 102b, 103c, a position controller for operating the first linear drive, second and / or third linear drive 101 ,102,103, the detector linear drive 116a, the mirror drive 114b, for pivoting the mirror 112 and for engaging the clamping elements 122, a light source controller for controlling the light source 105, a SLED controller.

[0227] As can further be seen in Fig. 3, an optical target 402 is arranged on the tray 300 below the head 100. The atomic force microscope 1 is configured to automatically determine and calibrate the position of the head 100 in x-direction and in y-direction when no cantilever chip holder 310 is mounted on the head 100. For the determination and calibration, the light beam emitted from the light source 105 passes through the light and focusing optics 105a of the head 100, and hits the optical target 402. The light reflected from the optical target 402 goes back towards the light and focusing optics 105a, which has a beam splitter connected to an optical fiber that is configured to calibrate the position of the head 100 with respect to the tray 300 along the x- and the y-direction based on the light reflected from the optical target 402.

[0228] Fig. 4a schematically depicts an optical path of light emitted from said light source 105 and detected by said detector 113. The light is emitted from the light source 105 as a first light beam B1 towards the cantilever 202 held by the cantilever holder 310. From the cantilever 202, particularly from a top surface of the cantilever 202 facing the light source 105, the light is reflected as a second light beam B2 towards the mirror 112, which can be rotated by means of said mirror drive 114b, cf. Fig. 2. From the mirror 112, the light is finally reflected as a third light beam B3 towards the detector 113, which may be a 4-quadrant photodiode. The optical path can be changed by moving the light source 105 with respect to the cantilever 202 using the cross table 110 with its first, second and third light source linear drives 111 a, 111b, 111c along the x-y- and the z-direction, respectively. Additionally, the photodetector 113 can be moved along the x-direction, i.e., outside the drawing plane shown in Fig. 4a, by means of said detector linear drive 116a. The deflection of the cantilever 202 is indicative of the distance between the tip 203 of the cantilever and the sample 3 arranged on the sample stage 300.

[0229] Fig. 4a also schematically indicates a first linear drive 101 with a coarse linear motor 101a and a fine linear motor 101 b for moving the cantilever 202 held by the cantilever holder 310 along the first longitudinal axis L1 as well as a second linear drive 102 with a coarse linear motor 102a and a fine linear motor 102b for moving the cantilever 202 held by the cantilever holder 310 along the second longitudinal axis L2. Additionally, the sample stage 300 can be moved by a third linear drive 103 comprising a coarse linear motor 103a and a fine linear motor 103b along the third longitudinal axis L3.

[0230] Fig. 4b shows an embodiment, wherein the cantilever holder 310 is configured to receive a transparent body 320. The transparent body 320 is formed from an optically clear material, such as a glass material. In particular, the material of the transparent body comprises a refractive index which is greater than the refractive index of air. In certain embodiments, the material of the transparent body comprises a refractive index which is greater than the refractive index of the material of the holder. For example, the transparent body comprises a refractive index of 1.6 to 2.2, more particularly about 1.9.

[0231] In case a transparent body 320 is provided, the light beam used to determine the deflection of the free end of the cantilever 202 travels through the transparent body 320 and onto the cantilever 202 and the light beam reflected from the cantilever 202 travels towards the mirror 112 through the transparent body 320. At the interfaces of the transparent body 320, the light is refracted. The transparent body 320 improves the stability of an assembly formed by the transparent body 320 and the cantilever holder 310, such that the cantilever holder 310 can be made longer in the z-direction, which simplifies the handling of the cantilever holder 310. In addition, in case the material of the transparent body comprises a higher refractive index than air, this allows the optics of the atomic force microscope 1 to be built more compactly, since the refraction of the light beam at the transparent body 320 allows to refract the light beam towards the mirror 112, whereas in the case without the transparent body 320 the light has to be guided from the cantilever 202 to the mirror entirely due to the reflection of the cantilever 202, which requires a rather bulky shape of the cantilever holder 310, as can be understood from comparing the cantilever holders 310 of Fig. 4a and 4b.

[0232] In case a transparent body 320 is provided, the light beam used to determine the deflection of the tip of the cantilever 202 travels through the transparent body 320 and onto the cantilever 202 and the light beam reflected from the cantilever 202 travels towards the mirror 112 through the transparent body 320. At the interfaces of the transparent body 320, the light is refracted. The transparent body 320 improves the stability of an assembly formed by the transparent body 320 and the cantilever chip holder 310, such that the cantilever chip holder 310 can be made longer in the z-direction, which simplifies the handling of the cantilever chip holder 310. In addition, in case the material of the transparent body comprises a higher refractive index than air, this allows the optics of the atomic force microscope 1 to be built more compactly, since the refraction of the light beam at the transparent body 320 allows to refract the light beam towards the mirror 112, whereas in the case without the transparent body 320 the light has to be guided from the cantilever 202 to the mirror entirely due to the reflection of the cantilever 202, which requires a rather bulky shape of the cantilever chip holder 310, as can be understood from comparing the cantilever chip holders 310 of Fig. 4a and 4b.

[0233] Fig 5 shows a front view (corresponding to the xz-plane) of the cantilever chip holder 310 arranged in said receiving unit 106 of the lower part 104b of the head 100 in the area of a longitudinal axis L extending through the head 100, according to an embodiment of the invention.

[0234] Fig. 6 shows a side view (corresponding to the yz-plane) of the cantilever chip holder 310 arranged at said receiving unit 106 of the lower part 104b of the head 100, according to an embodiment of the invention. Particularly, Fig. 6 depicts said clamping elements 122 for clamping the cantilever chip holder 310 to the head 100. Each clamping element 122 is connected to a respective bolt 123 extending along the x-direction, perpendicularly to the drawing plane of Fig. 6, which is further explained in Fig. 9.

[0235] Fig. 7 shows a bottom view of the head 100 of the atomic force microscope 1 according to an embodiment of the invention. The view corresponds to the perspective of the tray 300 below the head 100, facing towards the head 100, such that said receiving unit 106 for insertion of the cantilever chip holder 310 is visible. In this embodiment, the receiving unit 106 forms a recess framing a circular through-hole 107 arranged centrally in the receiving unit 106. Three aligning elements 108, particularly formed by ceramic hemispheres, are arranged at the recess around the circumference of the through-hole 107 for aligning and particularly centering the cantilever chip holder 310, when the cantilever chip holder 310 is inserted in the recess. The edge around the recess may additionally be tapered along the z-direction to simplify the insertion of the cantilever chip holder 310.

[0236] Fig. 8 shows the cantilever chip holder 310 inserted in the receiving unit 106 of the head 100 of the atomic force microscope 1 , according to an embodiment of the invention. The cantilever chip holder 310 is held in a clamping position by two clamping elements 122 arranged on opposite sides of the recess, which engage with connecting surfaces 211 of the cantilever chip holder 310 to fix the cantilever chip holder 310 in the recess 106. Furthermore, the clamping elements 122 are connected to bolts 123 (e.g., rods) which are pivotably mounted in bearings

[0237] 124. One end of each bolt 123 is further connected to a lever 125. The levers 125 each comprise a stump 125a configured to attach a spring, such as a leaf spring, to bias the clamping elements 122 into the clamping position. The levers 125 extend from the outside of the head 100 shown in Fig. 8 to the inside of the head 100 shown in Fig. 9, such that the levers 125 can be operated by said push bar 119 arranged inside the head 100. The levers 125 are shown from the inside of the head 100 in Fig. 9. As illustrated in Fig. 2, the head 100 further comprises a push bar 119 which is configured to be moved along a longitudinal axis L, particularly by said mirror drive 114b. When the push bar 119 is moved down from the position displayed in Fig. 2 to the cantilever chip holder 310, the push bar 119 engages with the levers

[0238] 125, as indicated in Fig. 9, and pushes the levers 125 against the force of the springs into a downward position. Consequently, the bolt 123 is pivoted and thereby forces the clamping elements 122 towards an opening position (not shown), where the cantilever chip holder 310 can be inserted into the recess or removed from the recess, particularly automatically. Fig. 9 also shows a magnet 126 and a position sensor 126a for determining a position of the clamping elements 122, which allows to control and confirm if the clamping elements 122 and thus the cantilever chip holder 310 is arranged in the clamping or the opening position.

[0239] Fig. 10a shows an embodiment of an atomic force microscope 1 according to the invention, with a light source 105 arranged on an upper part 104a of the head 100. For example, the light source 105 comprises or is a laser source, a super luminescent LED. In particular, the light source 105 can comprise a fiber optic cable, wherein light generated by a laser source or a super luminescent LED is guided from the laser source or the super luminescent LED through the fiber optic cable into the head 100 toward the cantilever 202. The atomic force microscope 1 according to the present embodiment can comprise focusing and collimating optics 105a, such as lenses, arranged between the light source 105 and the cantilever 202, for example as depicted in Fig. 2. The cantilever 202 (not shown in Fig. 10a) is fixedly attached to a lower part 104b of the head 100, adjacent to the upper part 104a indicated in Fig. 10a. To control the optical path of the light beam between the light source 105 and the cantilever 202, the light source 105 is movably mounted on the upper part 104a of the head 100, such that the light beam emitted from the light source 105 can be moved with respect to the cantilever 202. The light source 105 is mounted to the upper part 104a of the head 100 via a cross-table 110 comprising a first carriage 110a, a second carriage 110b and a third carriage 110c.

[0240] The first carriage 110a of the cross-table 110 is movable in the y-direction, particularly with a resolution of about 10 pm, by a first light source linear drive comprising a piezo electric drive, particularly a piezo LEGS or Piezo Friction drive (111a). The second carriage 110b of the cross-table 110 is movable in the x-direction, particularly with a resolution of about 10 pm, by a second light source linear drive comprising a piezo electric drive, particularly a piezo LEGS or Piezo Friction drive (111 b). The third carriage 110c of the cross-table 110 is movable in the z-direction, by a third light source linear drive, particularly comprising a piezo-electric motor, particularly a piezo-leg motor, configured to move the third carriage 110c along the z-direction. As such, the light source 105 can be displaced with respect to the cantilever independently along the x-, y- and the z-direction, which allows for simplified alignment and focusing of the light beam on the cantilever 202.

[0241] To record a position of the light source 105 with respect to the cantilever 202, each the first, second and third carriage 110a,110b, 110c comprises a magnetic position sensor 126a. In particular, the displacement of the light source 105 with respect to the cantilever 202 can be controlled automatically using said controller 410 based on positional information of the position of the light source 105 with respect to the cantilever 202, such that the light beam can be automatically focused on the cantilever 202 and kept in focus during a measurement.

[0242] Optionally, as indicated in Fig. 10b, the first carriage 110a can be formed as a first plate 109a and the second carriage 110b can be formed as a second plate 109b, wherein the first plate 109a and the second plate 109b circumferentially enclose the body 150 with the light source 105. The first plate 109a and the second plate 109b are stacked on each other along the z- direction and connected by three set screws 114b. By screwing and by unscrewing the set screws 109c, the body 150 with the light source 105 can be tilted with respect to the head 100 and the cantilever 202, which allows to adjust the focus of the light beam emitted by the light source 105 on the cantilever 202 arranged on the head 100.

[0243] Fig. 11 shows an embodiment of the atomic force microscope 1 according to the invention, comprising a third imaging system 440 arranged and configured to image the cantilever 202. In particular, the third imaging system 440 can be arranged on the tray 300, for example as shown in Fig. 3, such that the third imaging system 440 is configured to image the cantilever 202 arranged on the head 100 from below, i.e. , from the perspective of the tray 300.

[0244] According to the present embodiment, the third imaging system 440 comprises a light source 444 arranged and configured for illumination of the cantilever 202, particularly the tip of the cantilever 202. Light emitted from the light source 444 can be reflected by a mirror comprised by the third imaging system 440 toward an objective of the third imaging system 440, wherein the objective faces the cantilever 202. In particular, the third imaging system 440 is an optical microscope 441.

[0245] Fig. 12 shows an embodiment of the atomic force microscope 1 according to the invention with focusing and collimating optics 105a for focusing and collimating light emitted from the light source 105 to the cantilever 202. The focusing and collimating optics 105a comprises a collimator 151 for collimating light coming from the light source 105 and a focusing lens 152 for focusing light collimated by the collimator 151 onto the cantilever 202. Between the collimator 151 and the focusing lens 152, the light propagates through a body 150 in which lenses of the focusing and collimating optics 105a are arranged. The focusing and collimating optics 105a further comprises an optical fiber connector 153 for connecting an optical fiber to the focusing and collimating optics 105a, such that light generated by a light source 105 can be guided via the optical fiber into the focusing and collimating optics 105a along the z-direction indicated in Fig. 12 and toward the cantilever 202. For example, the light source 105 comprises or is a laser source or a super luminescent LED. The focusing and collimating optics 105a according to the present embodiment comprises a third light source linear drive 111c for moving the body 150 together with the collimator 151 , the focusing lens 152 and the optical fiber connector 153 up and down along the z-direction with respect to the upper part 104a of the head 100. As such, the focus position on the cantilever 202 of the light emitted by the light source 105 can be changed by moving the focusing and collimating optics 105 with respect to the cantilever 202 along the z-direction. Fig. 13 shows an embodiment of a system 600 according to the second aspect of the invention. The system 600 comprises an atomic force microscope 1 according to the first aspect of the invention comprising a head 100. The system 600 according to the present embodiment comprises an interface 610, particularly a humanmachine interface, comprising two drawer units 620. The functions of the drawer units 620...

[0246] The system 600 further comprises a label scanner 612 for scanning a label arranged or arrangeable on the sample to be measured, the specimen holder 430, the cantilever chip holder 310 and / or the cantilever 202. In particular, the label comprises or is a bar code. As such, the system is configured to register the sample, the specimen holder 430, the cantilever chip holder 310 and / or the cantilever 202 used in measurements.

[0247] Fig. 14a shows an embodiment of an interface 610 with two drawer units 620 of a system 600 according to an embodiment of the second aspect of the invention.

[0248] The interface 610 can form a human-machine interface for transferring a specimen holder 430 with or without a sample to be measured to or from the sample stage 300 via the interface 610. In the same fashion, a cantilever holder or cantilever chip holder 310 can be transferred to or from the sample stage 300 via the interface 610. The cantilever chip holder 310 and / or the specimen holder 430 can be placed on and carried by a shuttle unit 650 which can be placed on protrusions 634 of each drawer unit 620. The interface 610 can be part of a housing 700 of the system 600 that frames the atomic force microscope 1 with the sample stage 300 and the head 100 movably connected to the mounting. The two drawer units 620 can be moved along the third longitudinal axis L3, between a loading position away from the sample stage 300, which is indicated in Fig. 14a, and an unloading position at the sample stage 300, as further shown in Fig. 14c below. To this end, the interface 610 comprises motors for driving the drawer units 620 between the loading position and the unloading position.

[0249] According to the present embodiment, each drawer unit 620 comprises slides 630, particularly telescopic slides, for moving the respective drawer unit 620 between the loading position away and the unloading position.

[0250] Each drawer unit 620 comprises a mechanism for unloading the shuttle unit 650 from the drawer unit 620 to the sample stage 300. To this end, as can be seen in Fig. 14a and, in more detail, in Fig. 14b and Fig. 14c, a height-adjustable structure 626 is slidably supported on each slide 630 by means of at least one pin 628 that extends from the slide 630 through a recess 632 of the height-adjustable structure 626. As can be understood from comparing Fig. 14b and Fig. 14c, moving the drawer unit 650 from the loading position to the unloading position causes the height-adjustable structure 626 to engage with a stop 627 of the drawer unit 620, which in turn causes the height-adjustable structure 626 to move relative to the slide 630, wherein the pins 628 of the slot 630 move from one end of the associated recess 632 to another. Because of the shape of the recess 632, which comprises two horizontal portions that each extend essentially along the third longitudinal direction L3 at two different vertical positions along the first longitudinal axis L1 , which are connected by a slope portion of the recess 632, the height- adjustable structure 626 and thus the shuttle unit 650 placed on the protrusions 634 of the shuttle unit 650 are lowered as the height-adjustable structure 626 engages with said stop 627.

[0251] As can further be seen in Fig. 14b and Fig. 15b, the sample stage 300 is arranged below the drawer unit 620. The sample stage 300 comprises a plurality, particularly three support elements 303 for supporting the shuttle unit 650 when the shuttle unit 650 is lowered by the above-described mechanism. In Fig. 14c, the shuttle unit 650 is shown placed on support elements 303 of the sample stage 300, wherein the height-adjustable structure 626 is lowered with respect to the slide 630 compared to Fig. 14b.

[0252] The drawer unit 620 can comprise elastic means such as springs that are biased once the height-adjustable structure 626 is moved by the motor from the position depicted in Fig. 14a to the position depicted in Fig. 14b. The height-adjustable structure 626 can be kept in the position depicted in Fig. 14b against the spring force of the biased spring by a resistance exerted by the motor for driving the drawer units 620. The shuttle unit 650 can be picked up again from the support elements 303 of the sample stage 300 by the protrusions 634 of the height-adjustable structure 626 under relaxation of the elastic means, which causes the heightadjusting structure 626 to move up the first longitudinal axis L1 to the position indicated in Fig. 14a. The above mechanism can be used to mechanically decouple the interface 610 and its drawer units 620 from the atomic force microscope 1 , particularly from the sample stage 300 and the head 100. In particular, the atomic force microscope 1 with the sample stage 300 and the head 100 are arranged on a vibration isolation table, and are not in mechanical contact with the interface 610 and the drawer units 620, particularly the housing 700 of the system 600. As such, operations of users at the interface 610 do not affect measurements of the atomic force microscope 1 , while allowing a transfer of sample holders 430 and / or cantilever chip holders 310 to and from the sample stage 300.

[0253] Fig. 15 shows an embodiment of the atomic force microscope 1 according to the invention. The present embodiment shows a linear drive, particularly a fine linear drive such as a piezo stick-slip drive with an optical ruler for optically detecting a driving distance of the linear drive. A linear drive according to the present embodiment can form in particular at least one of the carriages 110a, 110b, 110c of said cross-table 110 for moving the light source 105 with respect to the cantilever 202, as described for example in Fig. 1.

Claims

Claims1. An atomic force microscope (1) for obtaining a force spectroscopy measurement of a sample comprising a head (100) which is movably connected to a mounting, wherein the head (100) comprises:- a receiving unit (106) configured to receive a cantilever (202) comprising a tip (203) for contacting a sample of soft material,- a light source (105) for generating light and focusing optics (105a) configured to focus the light and direct a first light beam (B1) along a first longitudinal axis (L1) onto the cantilever (202), when the cantilever (202) is arranged at the receiving unit (106),- a mirror (112) configured to receive a second light beam (B2) reflected by the cantilever (202), when the cantilever (202) is arranged at the receiving unit (106),- a detector (113) configured to detect a position of a third light beam (B3) reflected by the mirror (112) and generate a deflection signal indicating a deflection of the cantilever (202) along the first longitudinal axis (L1) based on the detected position of the third light beam (B3), wherein the atomic force microscope (1) comprises an actuator (140) configured to move the head (100) along the first longitudinal axis (L1), particularly wherein the head (100) is connected to the mounting via the actuator (140).

2. The atomic force microscope (1) according to claim 1 , wherein the atomic force microscope (1) comprises a first imaging system (450), wherein the first imaging system (450) is movably connected to the mounting in fixed position with respect to the head (100), and / or wherein the atomic force microscope (1) comprises a second imaging system (460), wherein the second imaging system (460) is movably connected to the mounting in fixed position with respect to the head (100), and / or wherein the atomic force microscope (1) comprises a third imaging system (440), wherein the third imaging system (440) comprises a fixed orientation with respect to the head (100), with respect to the first imaging system (450) and / or with respect to the second imaging system (460).

3. The atomic force microscope (1) according to one of the claims 1 or 2, wherein the atomic force microscope (1) comprises a referencing assembly wherein the referencing assembly comprises a first optical target (402) and a controller (410), wherein the first optical target (402) comprises a high-reflection section and a low- refl ection section, wherein the controller (410) is configured to detect an intensity of a first reference lightbeam reflected by the first optical target (402), wherein the controller (410) is further configured to detect the position of the head (100) perpendicular to the first longitudinal axis (L1), and wherein the first imaging system (450) is arranged and configured to take an optical image of the first optical target (402) and / or wherein the second imaging system (460) is arranged and configured to take an optical image of the first optical target (402), wherein the referencing assembly is configured to determine: a position of the head (100) with respect to the first imaging system (450), a position of the head (100) with respect to the second imaging system (460), a position of the first imaging system (450) with respect to the second imaging system (460), a position of the head (100) with respect to the first light beam (B1), a position of the first imaging system (450) with respect to the first light beam (B1), and / or a position of the second imaging system (460) with respect to the first light beam (B1).

4. The atomic force microscope (1) according to one of the claims 1 to 3, wherein the atomic force microscope (1) comprises a cantilever (202) comprising a tip (203) for contacting a sample of soft material, wherein the cantilever (202) is arranged at the receiving unit (106), particularly wherein the cantilever (202) is arranged at the receiving unit (106) via a cantilever holder (310), wherein the light source (105) is configured and arranged such that the first light beam (B1) is directed along the first longitudinal axis (L1) onto the cantilever (202).

5. The atomic force microscope (1) according to one of the claims 1 to 4, wherein the atomic force microscope (1) comprises at least one linear drive (101 , 102, 103) configured to position the cantilever (202) relative to a sample stage (300) along the first longitudinal axis (L1) and / or along a second longitudinal axis (L2) which is perpendicular to the first longitudinal axis (L1), and / or along a third longitudinal axis (L3) which is perpendicular to the first longitudinal axis (L1) and the second longitudinal axis (L2), wherein the at least one linear drive (101 , 102, 103) comprises a coarse linear motor (101a, 102a, 103a), particularly a voice coil motor, for coarse positioning of the cantilever (202) relative to the sample stage (300), and a fine linear motor (101b, 102b, 103b), particularly a piezo-electric motor, particularly a piezo-stack motor, for fine positioning of the cantilever (202) relative to the sample, wherein a first linear drive (101) is configured to move the cantilever (202) along the first longitudinal axis (L1) and / or a second linear drive (102) is configured to move the cantilever (202) along thesecond longitudinal axis (L2) and / or a third linear drive (103) is configured to move the sample stage (300) along the third longitudinal axis (L3).

6. The atomic force microscope (1) according to one of the claims 1 to 5, wherein the atomic force microscope (1) comprises at least one linear drive (101 , 102) configured to position the head (100) relative to the mounting, wherein the at least one linear drive (101 , 102) comprises a coarse linear motor (101 a, 102a), particularly a voice coil motor, for coarse positioning of the head (100) and a fine linear motor (101 b, 102b), particularly a piezo-electric motor, particularly a piezo-stack motor, for fine positioning of the head (100), wherein the head (100) is connected to the mounting via the at least one linear drive (101 , 102) and the actuator (140), wherein more particularly the actuator (140) is directly connected to the mounting and the at least one linear drive (101 , 102) connects the actuator (140) and the head (100).

7. The atomic force microscope (1) according to one of the claims 1 to 6, wherein the atomic force microscope (1) comprises at least one spring (130) connecting the head (100) to the mounting, wherein the at least one spring (130) is configured to exert a force on the head (100) along the first longitudinal axis (L1), particularly wherein the force remains constant when the spring (130) is compressed or elongated along the first longitudinal axis (L1).

8. The atomic force microscope (1) according to one of the claims 1 to 7, wherein the head (100) comprises an alignment mechanism for aligning the first light beam (B1) comprising a support (109a) for holding the light source (105) and the focusing optics (105a), a first carriage (110a) and a first light source linear drive (111a), particularly comprising a piezo-electric motor, particularly a piezo-leg motor, configured to move the first carriage (110a) along the second longitudinal axis (L2) perpendicular to the first longitudinal axis (L1) and a second carriage (110b) and a second light source linear drive (111 b), particularly comprising a piezo-electric motor, particularly a piezo leg motor, configured to move the second carriage (110b) along the third longitudinal axis (L3) perpendicular to the first longitudinal axis (L1) and the second longitudinal axis (L2).

9. The atomic force microscope (1) according to one of the claims 1 to 8, wherein the head (100), particularly the support (109a), further comprises a first plate (109b) configured to hold the light source (105) and the focusing optics (105a), a second plate (109c), and at least one set-screw (109d) connecting the first plate (109b) and the second plate (109c), wherein the first plate (109b) is tiltable with respect to the second plate (109c) by adjusting the set-screw (109d) to align the first light beam (B1) orwherein the head (100) further comprises a third carriage (110c) and a third light source linear drive (111c), particularly comprising a piezo-electric motor, particularly a piezoleg motor, configured to move the third carriage (110c) along the first longitudinal axis (L1).

10. The atomic force microscope (1) according to one of the claims 1 to 9, wherein the head (100) comprises a mirror frame (114), wherein the mirror (112) is connected to the mirror frame (114), and wherein the mirror frame (114) is pivotably connected to a mirror pivot axis (114a), wherein the head (100) further comprises a mirror drive (114b) configured to pivot the mirror frame (114) about the mirror pivot axis (114a), wherein particularly the mirror drive (114b) is a linear drive, particularly a piezo-electric motor, particularly a piezo-leg motor, and wherein the head (100) comprises a push bar (119) comprising a pin (120) and a connecting lever (121) comprising a slot (121a), wherein the mirror drive (114b) is configured to move the push bar (119) along the first longitudinal axis (L1), wherein the connecting lever (121) is connected to the mirror pivot axis (114a) and the pin (120) of the push bar (119) is arranged in the slot (121a) of the connecting lever (121), such that the mirror frame (114) is pivoted around the mirror pivot axis (114a) by means of the connecting lever (121) when the push bar (119) is moved along the first longitudinal axis (L1) by the mirror drive (114b).

11. The atomic force microscope (1) according to one of the claims 1 to 10, wherein the head (100) comprises a detector holder (116), wherein the detector (113) is connected to the detector holder (116), and wherein the head (100) comprises a detector linear drive (116a), particularly a piezo-electric motor, particularly a piezo-leg motor, configured to move the detector holder (116) along the second longitudinal axis (L2) perpendicular to the first longitudinal axis (L1) to adjust the position of the detector (113).

12. The atomic force microscope (1) according to one of the claims 1 to 11 , wherein the head (100) comprises at least one position sensor (126a), particularly a Hall sensor, configured to determine:- a rotational orientation of the mirror frame (114) on the mirror pivot axis (114a),- a position of the detector holder (116) along the second longitudinal axis (L2),- a position of the first carriage (110a) of the alignment mechanism along the second longitudinal axis (L2),- a position of the second carriage (110b) of the alignment mechanism along the third longitudinal axis (L3), and / ora position of the third carriage (110c) of the alignment mechanism along the first longitudinal axis (L1).

13. The atomic force microscope (1) according to one of the claims 1 to 12, wherein the head (100) comprises a temperature sensor, particularly wherein the temperature sensor is positioned at or close to the cantilever (202), such that a temperature of the cantilever (202) can be determined from a temperature value measured by the temperature sensor and / or such that a temperature of the sample can be determined from a temperature value measured by the temperature sensor, particularly wherein the temperature sensor is arranged and configured to determine the temperature value repeatedly, particularly wherein the temperature sensor is arranged and configured to determine the temperature value continuously over time.

14. The atomic force microscope (1) according to one of the claims 1 to 13, wherein the atomic force microscope (1), particularly the head (100), comprises an environmental sensor configured to detect an environmental humidity and / or an environmental pressure at or close to the cantilever (202).

15. The atomic force microscope (1) according to one of the claims 1 to 14, wherein the atomic force microscope (1) further comprises the sample stage (300) for receiving the sample, particularly a biological tissue specimen, particularly wherein the sample stage (300) is configured to receive a specimen holder (430) configured to hold the sample, particularly wherein the specimen holder (430) is one of a culture dish, a petri dish, a microplate, particularly a 6-well plate, or a specimen support, wherein more particularly the sample stage (300) comprises a first slot for receiving the specimen holder (430), wherein even more particularly the sample stage (300) comprises a second slot for receiving a cantilever holder (310) comprising a cantilever (202), wherein the atomic force microscope (1) is configured such that the cantilever holder (310) can be automatically mounted to the head (100).

16. The atomic force microscope (1) according to one of the claims 1 to 15, wherein the third imaging system (440) is arranged and configured to detect the cantilever (202), particularly the tip (203) of the cantilever (202), particularly wherein the third imaging system (440) comprises a light source configured for coaxial illumination and / or particularly wherein the atomic force microscope (1) comprises a light source (444) arranged and configured for illumination of the cantilever (202), particularly the tip (203) of the cantilever (202), particularly wherein the third imaging system (440) is an optical microscope (441).

17. The atomic force microscope (1) according to one of the claims 1 to 16, wherein the atomic force microscope (1) comprises a vibration sensor, particularly wherein the vibration sensor is a vibrometer, an accelerometer or a geophone, and / or wherein the atomic force microscope (1) comprises an AFM controller, particularly wherein the AFM controller comprises a field programmable gate array and / or wherein the AFM controller comprises a device controller for automation of the atomic force microscope (1).

18. The atomic force microscope (1) according to one of the claims 1 to 17, wherein the atomic force microscope (1) comprises a clamping element (122), wherein the clamping element (122) is configured to be in a pre-loaded position, in a clamping position, or in a receiving position, wherein in the receiving position, the clamping element (122) is configured to receive a cantilever holder (310), wherein in the clamping position, the clamping element (122) is configured to clamp the cantilever holder (310) to the atomic force microscope (1), wherein the atomic force microscope (1) comprises at least one spring configured to bias the clamping element (122) into the clamping position.

19. The atomic force microscope (1) according to one of the claims 1 to 18, wherein the atomic force microscope (1) comprises a magnetic sensor configured so sense the position of the clamping element (122), particularly to sense that the clamping element (122) is in the pre-loaded position, in the clamping position, or in the receiving position.

20. The atomic force microscope (1) according to claim one of the claims 1 to 19, wherein the atomic force microscope (1) comprises a clamping drive (1222), particularly wherein the clamping drive (1222) is a linear drive, particularly a piezo-electric motor, particularly a piezo-leg motor, wherein the clamping drive (1222) is configured to move a means (1224) along the first longitudinal axis (L1), wherein the means (1224) is configured to transfer the clamping element (122) from the pre-loaded position to the receiving position and / or from the clamping position to the receiving position, when the means (1224) is moved along the first longitudinal axis (L1), particularly wherein the clamping drive (1222) is the mirror drive (114b).

21. The atomic force microscope (1) according to one of the claims 1 to 20, wherein the atomic force microscope (1) comprises a control device configured to control the actuator (140), such that the head (100) is moved along the first longitudinal axis (L1), particularly such that the tip (203) of the cantilever (202) contacts the sample, whereinparticularly the control device is configured to receive the deflection signal from the detector (113).

22. The atomic force microscope (1) according to claims 2 to 21 , wherein the atomic force microscope (1) comprises the first imaging system (450) and the second imaging system (460), wherein the first imaging system (450) is configured to provide a deep focus image and / or a depth map, particularly wherein the first imaging system (450) is configured to record a plurality of tiles, particularly tiles extending perpendicular to the first longitudinal direction (L1), stacked along the first longitudinal direction (L1) to provide a deep focus image and / or a depth map.

23. The atomic force microscope (1) according to one of the claims 1 to 22, wherein the light source (105) is an optical fiber.

24. The atomic force microscope (1) according to one of the claims 1 to 23, wherein the atomic force microscope (1) comprises a source of electric power, particularly a battery.

25. A system (600) comprising the atomic force microscope (1) according to one of the claims 1 to 24, and a vibration isolation table, wherein the vibration isolation table comprises a vibration sensor.

26. The system (600) according to claim 25, wherein the system (600) comprises an interface (610), particularly a human-machine interface, comprising a drawer unit (620) configured to receive a shuttle unit (650) for carrying a specimen holder (430) and / or a cantilever holder (310), wherein the drawer unit (620) can be moved between a loading position away from the sample stage (300) for placing the shuttle unit (650) onto the drawer unit (620) and an unloading position at the sample stage (300), and wherein the drawer unit (620) comprises a mechanism for unloading the shuttle unit (650) from the drawer unit (650) to the tray (300), when the shuttle unit (650) is placed on the drawer unit (650) and the drawer unit (650) is moved to the unloading position.

27. The system (600) according to claim 26, wherein the drawer unit (620) comprises a plurality of protrusions (634) for placing the shuttle unit (650) on the protrusions (634) of the drawer unit (620).

28. The system (600) according to claim 26 or 27, wherein the drawer unit (620) comprises slides (630), particularly telescopic slides, for moving the drawer unit (620), particularly along the third longitudinal direction (L3), between the loading position and the unloading position.

29. The system (600) according to claim 28, wherein a height-adjustable structure (626) is slidably supported on each slide (630) by means of at least one pin (628) extending from the slide (630) into a recess (632) of the height-adjustable structure (626), and wherein the height-adjustable structure (626) comprises said protrusions (634), and wherein the recess (632) is shaped such that when the drawer unit (650) is moved in the unloading position and the height-adjustable structure (626) engages with a stop (627), a vertical position of the protrusions (634) and of the shuttle unit (650) placed on the protrusions (634) is lowered by moving the height-adjustable structure (626) with respect to the at least one pin (628) of the slide (630).

30. The system (600) according to claim 29, wherein the sample stage (300) comprises a plurality of support elements (303), particularly three supporting elements (303), for supporting the shuttle unit (650), wherein in said unloading position, the protrusions (634) for placing the shuttle unit (650) are arranged above said support elements (303), such that the shuttle unit (650) can be unloaded from the drawer unit (620) to the sample stage (300) by lowering the height-adjustable structure (626) with respect to the slide (630).31 . The system (600) according to one of the claims 25 to 30, wherein the system (600) comprises a label scanner, particularly a label scanner for scanning a bar code, particularly wherein the bar code codes information regarding the sample, the specimen holder, the probe holder and / or the cantilever.

32. The system (600) according to one of the claims 25 to 31 , wherein the specimen holder (430) is a culture dish, a petri dish, a microplate, particularly a 6-well plate, or a specimen support.

33. The system (600) according to one of the claims 25 to 32, wherein the system (600) comprises a hydraulic wheel, wherein the hydraulic wheel is arranged and configured such that the system (600) is movable by means of the hydraulic wheel, and wherein the system (600) comprises a fixed supporting foot, wherein the fixed supporting foot is arranged and configured such that the system is stands stable and / or is alignable by means of the supporting foot.

34. An assembly comprising the system (600) according to one of the claims 25 to 33 and a packaging, wherein the packaging is configured to receive the system (600), wherein the packaging comprises a ramp configured to support the release of the system fromthe packaging, particularly wherein the packaging comprises a lid, wherein the lid is configured as a wedge and configured to be arrangeable such that the system is releasable from the packaging via the wedge.

35. A method for obtaining a force spectroscopy measurement of a sample, particularly a sample from a soft material, more particularly a biological tissue specimen, using the atomic force microscope (1) according to one of the claims 1 to 24 or the system (600) according to one of the claims 25 to 34 or the assembly according to claim 34, comprising the following steps:- contacting the sample by the tip (203) of the cantilever (202) at a measurement spot,- moving the head (100) by the actuator (140) towards the sample parallel to the first longitudinal axis (L1) in a loading step, particularly automatically, such that the tip (203) of the cantilever (202) indents into the sample,- moving the head (100) away from the sample by the actuator (140) parallel to the first longitudinal axis (L1) in an unloading step, particularly automatically,- during the loading step and / or the unloading step, determining a plurality of deflection values of the cantilever (202) from the deflection signal of the detector (113),- determining the force spectroscopy measurement of the sample at the measurement spot from the determined deflection values, particularly automatically.

36. The method according to claim 35, wherein the method comprises determining a plurality of force spectroscopy measurements indicating the force of the cantilever (202) on the sample at a corresponding position along the first longitudinal axis (L1), wherein the force spectroscopy measurement of the sample is determined, particularly determined automatically, from at least a subset of the force values, particularly wherein a force-position relationship and / or a force-position curve is obtained.

37. The method according to one of the claims 35 or 36, wherein altering a position of the sample relative to the head (100), particularly relative to the cantilever (202), is provided by moving the head (100) along the first longitudinal direction (L1), particularly by the first linear drive (101) and / or the actuator (140) and / or along the second longitudinal direction (L2), particularly by the second linear drive (102), and / or by moving the sample, particularly the specimen holder (430) holding the sample, particularly by the third linear drive (103), along the third longitudinal direction (L3).

38. The method according to one of the claims 35 to 37, wherein the detector (113) is moved by the detector linear drive (116a) along the second longitudinal axis (L2) and / or the mirror frame (114), particularly the associated mirror (112), is pivoted by the mirror drive (114b), such that the mirror (112) and the detector (113) are aligned such that the third light beam (B3) reflected by the mirror (112) meets a pre-defined detection region of the detector (113), particularly a linear region of the detector (113), particularly the detector (113) and / or the mirror frame (114) are moved automatically keeping the third light beam (B3) in the detection region of the detector (113), particularly the linear region of the detector (113).

39. The method according to one of the claims 35 to 38, wherein a shape of the cantilever (202), a size of the cantilever (202), a length of the cantilever (202), a width of the cantilever (202) and / or a height of the cantilever (202) is determined, particularly determined automatically, by the third imaging system (450), and / or wherein the position of the tip (203) of the cantilever (202) is determined, particularly determined automatically, by the third imaging system (450).

40. The method according to one of the claims 35 to 39, wherein prior to contacting the sample by the tip (203) of the cantilever (202), the method comprises the step of 3D profiling of the sample using information obtained by the first imaging system (450), and / or using information obtained by the second imaging system (460), particularly information related to a deep focus image and / or a depth map, particularly using information obtained by the first imaging system (450), and / or using information obtained by the second imaging system (460) and calculations, particularly wherein the calculations comprise the step of a Laplacian regression.

41. The method according to one of the claims 35 to 40, wherein prior to contacting the sample by the tip (203) of the cantilever (202), the method comprises the step of an automated spot selection based on sample-related information, based on information obtained by 3D profiling of the sample, based on information obtained by deep focus imaging, based on information obtained by the depth map, based on information regarding the shape of the cantilever (202), based on information regarding the size of the cantilever (202), and / or based on the position of the tip (203) of the cantilever (202), providing a prediction of the quality of the measurement at the selected spot of measurement.

42. The method according to one of the claims 35 to 41 , wherein prior to contacting the sample by the tip (203) of the cantilever (202), a reference sample having a predefinedstiffness is contacted by the tip (203) of the cantilever (202), and the head (100) is moved by the actuator (140) over a moving distance while the tip (203) contacts the reference sample without indentation of the reference sample, and wherein the deflection signal of the detector (113) is obtained over the moving distance, and wherein a deflection sensitivity of the detector (113) is determined, particularly automatically, from the moving distance and the deflection signal, and / or wherein the deflection sensitivity of the detector (113) is determined in a contact-free manner particularly based on information obtained by a resonance curve, particularly the area below the resonance curve, wherein the resonance curve is taken in air or in liquid, and / or wherein the deflection sensitivity of the detector (113) is determined in a depth stepping standards manner particularly wherein prior to contacting the sample by the tip (203) of the cantilever (202), a surface of a standard sample comprising recesses with pre-defined depth is contacted by the tip (203) of the cantilever (202), and the head (100) is moved by the actuator (140) over a moving distance, and wherein the deflection signal of the detector (113) is obtained over the moving distance, and wherein a deflection sensitivity of the detector (113) is determined, particularly automatically, from the moving distance and the deflection signal.

43. The method according to one of the claims 35 to 42, wherein the deflection sensitivity is determined, wherein the device detects when the deflection sensitivity leaves a predefined interval and initiates a recalibration, particularly automatically, by re-aligning the beam and executing an additional deflection sensitivity measurement.

44. The method according to one of the claims 35 to 43, wherein the cantilever (202) is moved in the proximity of the sample or moved to the spot of measurement, particularly wherein the cantilever (202) is moved in the proximity of the sample or moved to the spot of measurement in an approach-free manner, based on sample-related information, and / or based on information obtained by 3D profiling of the sample.

45. The method according to one of the claims 35 to 44, wherein an adaptive optimization is performed to move the cantilever (202) in the proximity of the sample or to the spot of measurement, wherein information considering a deviation between a detected previous spot of measurement and a proposed previous spot of measurement is used, particularly by applying the deviation on the proposed spot of measurement.

46. The method according to one of the claims 35 to 45, wherein a vibration signal is obtained from the vibration sensor, and wherein the head (100) is moved away from the sample along the first longitudinal axis (L1), particularly automatically, in case thesignal indicates the presence of a vibration, particularly of a vibration having an amplitude or a frequency above a predefined threshold.

47. The method according to one of the claims 35 to 46, wherein a spring constant of the cantilever (202) is determined, particularly determined automatically, based on data provided by the third imaging system (440), particularly a length of the cantilever (202), a width of the cantilever (202) and / or a thickness of the cantilever (202), based on the temperature value obtained by the temperature sensor, based on the environmental humidity obtained by the environmental sensor and / or the environmental pressure obtained by the environmental sensor, a resonance frequency of the cantilever (202), and / or a quality factor of the cantilever (202).

48. The method according to one of the claims 35 to 47, wherein the spring constant of the cantilever (202) is detected repeatedly during the measurement and / or a value related to the deflection sensitivity calibration is detected repeatedly during the measurement, wherein, when the spring constant of the cantilever (202) leaves a pre-defined range, the cantilever (202) is exchanged and / or a re-calibration is executed, and / or wherein, when the value related to a deflection sensitivity calibration leaves a pre-defined range, the cantilever (202) is exchanged and / or a re-calibration is executed, wherein the recalibration comprises a re-alignment of the first beam relative to the cantilever, particularly the tip (203) of the cantilever, and an additional measurement of the value related to the deflection sensitivity, and / or wherein a quality value of the cantilever is detected repeatedly during the measurement, wherein, when the quality value of the cantilever (202) leaves a pre-defined range, the cantilever (202) is exchanged.

49. The method according to one of the claims 35 to 48, wherein the cantilever (202), particularly the cantilever holder (310), is picked up, particularly picked up from the sample stage (300), particularly from the second slot (302) comprised in the sample stage (300) for receiving the cantilever holder (310) comprising the cantilever (202), when the head (100) determines a pre-defined force value executed by the cantilever (202) to be picked up, particularly the cantilever holder (310) to be picked up.

50. The method according to one of the claims 35 to 49, wherein a correction factor related to a buffer surrounding the sample is determined, wherein prior to contacting the sample by the tip (203) of the cantilever (202), the deflection signal is detected while the cantilever (202) is moved along the first longitudinal axis (L1) towards the sample, wherein a position of the cantilever (202) along the first longitudinal axis (L1) is determined when the deflection signal is lost.

51. The method according to one of the claims 35 to 50, the atomic force microscope, particularly the AFM controller, particularly the field programmable gate array, monitors at least one monitoring signal, wherein a monitoring signal is one of:- a cantilever sum and the deflection signal,- a vibration signal,- a signal of the accelerometer,- a signal the geophone,- a signal obtained by a microphone, wherein the atomic force microscope:- interrupts any motion in the atomic force microscope to protect the probe, when the monitoring signal leaves a pre-defined range,- interrupts the force spectroscopy measurement, when the monitoring signal leaves a pre-defined range particularly in the case of external vibrations or external acoustic noise,- interrupts an optical measurement by the first optical system and / or the second optical system and / or the third optical system, when the monitoring signal leaves a pre-defined range, particularly in case of external vibrations or external acoustic noise, and / or- uses the monitoring signals as “gates” to only trigger measurements, when vibration and noise levels are low.

52. The method according to one of the claims 35 to 51 , wherein the deflection sensitivity in a liquid, particularly the buffer, is determined based on the deflection sensitivity determined in air and positional information regarding the tip (203) of the cantilever, particularly information regarding the position of the tip (203) of the cantilever with respect to the first longitudinal direction, the second longitudinal direction and / or the third longitudinal direction.

53. A method to obtain a refence position of the light beam using the atomic force microscope (1) according to one of the claims 1 to 24 or the system (600) according to one of the claims 25 to 34, comprising the following steps: a. directing a first light beam (B1) along a first longitudinal axis (L1) at the first optical target (402), b. detecting the intensity of the first reference light beam reflected by the first optical target (402) by the controller (410), c. detecting the position of the head (100) perpendicular to the first longitudinal axis (L1) by the controller (410),d. taking an optical image of the first optical target (402) by the first imaging system (450) and / or the second imaging system (460), e. determining a position of:- the head (100) with respect to the first imaging system (450), - the head (100) with respect to the second imaging system (460),- the first imaging system (450) with respect to the second imaging system (460),- the head (100) with respect to the first light beam (B1),- the first imaging system (450) with respect to the first light beam (B1), and / or- the second imaging system (460) with respect to the first light beam (B1).

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