Objective changer; P250

For technicians and partly for sales managers!

 

 

These instructions describe the procedures to install and adjust the focus unit and the objective changer of the scanner Pannoramic 250. The focus unit and the objective changer are assembled into a complex unit and these parts can not be separated in the field. To help resolve problems with the objective changer or the focus unit a hardware description of the implemented components and adjustment procedures are added. To increase the manageability, the parts are discussed separately.

Software relevant settings are based on the software version 1.15.

 

 

                Contents

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

General

The described focus unit is a component of the scanner Pannoramic 250 and includes the following main parts:

·       Focus unit

 

 The focus unit gives the possibility of focusing the FOV (field of view, seen by the scan camera) automatically during the scan process of the sample.

 

The objective changer unit is mounted onto the focus unit and allows the consecutive use of two, preinstalled objectives. The selection or exchange of the objective is initiated by software before a slide scan session is started. The movement of the objective holder disc is executed via a DC-motor and the position is sensed via Hall sensors. Each objective position has a separate hall sensor, so the software knows always which objective is actually in use. To guarantees the proper position of the objective in the light path, the final objective position is fixed via a form-fit mechanism.

 

The unit was developed for the use of objectives of the following types:

 

Plan-APOCHROMAT 20x/0.8             and     Plan-APOCHROMAT 40x/0.95

 

 

The objective type “Plan-Apochromat” contains several, special chromatic and spherical corrections and delivers so an image of very high color trueness with very small spherical aberration.

 

If the mechanical dimensions do not exceed the size of the Plan-APOCHROMAT 40x0.95 type objective, the mechanical mounting is identical and the focus distance of the objective to the tissue is not closer then 0.25mm, other kind of objectives can be used also but it is strongly not recommended! Always check with 3DHISTECH first if a different objective should be used!

 

The shutter mechanics covers the condenser and this way the bright field illumination path is broken during fluoresce scan sessions. The two commands, “condenser cover off” and “condenser cover on” are identical with the physical lower and upper hardware limit of the focus unit.

 

Automatic scan

In the slide scan table of the scan program “SlideScanner.exe” you can assign the objective with which the slide should be scanned. In other words, the objective can be changed any time between two slide scanning procedures. If the slide scan table does not contain information about the objective, the scan procedure will be done with the actual objective.

 

Remark

By default, the brightfield scan procedure is always done by the use of the 20x objective; during the fluorescent scan session can be done with the 20x or the 40x objective likewise.

 

Since the software version 1.16 the BF scan session may be executed with the 20x or the 40x objective likewise.

 

       Upgrade to the software version 1.16” and “What is new” “Brightfield scan with the 40x objective

 

 

Manual scan

 The objective can be changed any time, before the scan procedure is started.

 

The bus connector provides the power supply and Data information for the objective changer as well as for the turret unit via the cable DCJ-1; the stepper motor cables of the focusing part are connected to the connectors “Z” of the “X-Y-Z-ND and Flash” controller.

 

The exchange of the focus unit with the objective changer is possible:

 

  • If the stepper motor or its connection is faulty
  • If the DC motor or the electronics of the objective changer unit is faulty.
  • If the shape of any part is deformed or a part is broken.
  • If the focus unit or the objective changer has any fault and you are unable to fix it.

 

Requirements

  • Service program for slide scanners (SlideScannerService.exe) with actual license file
  • Slide scanner (version 1.15 or higher) and Slide scanner Viewer software (SlideScanner.exe, SlideScannerViewer.exe) with actual license files
  • 1.5, 2.5, 3 and 5 mm hex key wrenches,
  • A pair of special wrenches to adjust the objective position
  • Hardware and construction knowledge of the scanner Panoramic 250

 

Attention:       Do not mix the versions of SlideScanner.exe and SlideScannerService.exe! Always use these programs with the same version number. Otherwise the SlideScannerService.exe program could produce unwanted results and SlideScanner.exe does not work correctly or even freeze!

 

 

The objective nut and the lock nut of the objective mounting are handled by using the two wrenches shown on the right; these are delivered with the scanner Pannoramic 250 until spring of 2014.

 

  • To fix the objective position, hold still the objective mounting nut with one wrench and loosen or tighten respectively the lock nut with the other wrench.

 

 

 

 

 

 

Configure the objective changer unit

 

 

Since the software version 1.15 the units of the scanner are configured in the file “MicroscopeConfiguration.ini”, section [Microscope].

 

The actual version of the objective changer unit in the scanner Pannoramic 250 is “ObjectiveChangerType=OC_2Pos”.

 

 

·       The path of the file MicroscopeConfiguration.ini, in the software version with the operating system Windows® 7 is:

C:\ProgramData\3DHISTECH\SlideScanner\MicroscopeConfiguration.ini

 

 

[Microscope]

SerialNumber=xxxxx

MicroscopeType=3DMic9

ScanCameraType=

PreviewCameraType=CVrmc_m8_pPro

BarcodeReaderType=PreviewCamera

LoaderType=SL_9Mag_25Slide_Sensor_Vertical

CameraChangerType=CC_3DH_2Pos

ReflectorTurretType=RT_3DH_10Pos_Belt

BrightfieldLightSourceType=FlashLight2010

ObjectiveChangerType=OC_2Pos

ObjectGuideXYZType=OGXYZ_FLASH3

FlashUnitType=FlashUnit_Type2

NDFilterType=NDType2

PreviewLightType=PreviewLightUnitType_Type2

PowerSwitchBoardType=PowerSwitchBoard_Type1

 

If a different value is used for the objective changer unit, may be the unit is not recognized or is working wrong!

 

 

 

 

 

 

Functional overview of the focus unit

 

 

Physically, the focus position is defined by the distance of the objective to the tissue. If the tissue is in the focus of the objective, a sharp image is seen by the camera. Because the tissues are different from each other in thickness, and the thickness can change inside the same tissue also, the focus position must be checked and corrected always, during the scan procedure.

 

  • In Pannoramic scanners, as well as in the P250, the real focus position is found by moving the slide toward or away from the objective via the focus pin.

 

  • The focus position is influenced by both, the fixed (adjustable) objective position and the actual position of the focus pin.

 

Furthermore, if the slide as well as the objective will be changed, the specimen holder must be moved away from the objective to avoid collision.

 

If the shutter is changed from shutter off to shutter on or vice versa, the specimen holder is moved to a position where the focus pin is not in connection with the slide or the specimen holder. This way, the distance between specimen holder (cover slip) and objective is not affected if the ex-center crosses the upper peak of the sine wave.

During all these actions, except focusing, the objective must be protected against touching the cover slip and the focus pin and 4:1 divider must be protected against overstraining. This is realized by the focus unit, with the different positions of the focus pin and the positions of the specimen holder.

 

As you can see, the gaps between cover slip and objective are very small, especially if the objective Plan-APOCHROMAT 40x/0.95 is used.

Because both objectives can be used without adjustment of the objective position after exchange, the previously developed focus unit (the focus unit for SCAN, MIDI and DESK) must be modified for the consecutive use of the 20x and 40x objectives. The worst case is defined by the parameters of the 40x objective.

 

       Optical path and Field Of View

 

 

 

 

 

 

 

 

                   40x objective

On the scale of the 40x objective, the nominal thickness of the cover slip should be selected.

 

  • If the real thickness of the cover slip differs from the selected or adjusted value, the quality of the scanned FOV may be reduced!

 

 

 

 

 

 

 

 

                   Objective position

 

In Pannoramic type scanners the objective is mounted into the middle of the focus range, offered by the movement limits of the focus pin during focusing.

 

  • It is very important, that the objective position is adjusted well; otherwise, if the objective position does not match the focus range, offered by the focus pin movement range, a focused camera image can never be produced!
  • Because the slide thickness can vary from 0.95 to 1.20 mm, the thickness of the slide, used to adjust the objective position, is also important during the objective position adjustment.

 

            Adjust the objective position”. 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Principle of focusing

The slide is hold in the specimen holder (not shown; see the X-Y-Stage). To achieve a parallel movement of the slide in relation to the objective, the specimen holding mechanics contains a parallelogram. This guarantees the position of the tissue to be always perpendicular to the optical axis during focusing. The specimen holder has a mechanical preload, so the slide has always a connection to the focus pin, except during a shutter on qr off operation.

To avoid collision of the objective with the slide during the “shutter on” command, the X-Y-stage is moved to a place (X=27000 and Y=18000 steps) where the focus pin is not in connection with the slide or the specimen holder; otherwise the 40x objective would collide with the slide or specimen holder.

This principle is used also if an objective exchange procedure is in progress with the objective changer.

 

 

The focus pin is moved by a stepper motor via an ex-center and a 4:1 divider. The resolution of the stepper motor with 6400 steps per revolution, the characteristics of the ex-center and the 4:1 divider allows a very fine resolution of the focus steps and so tissue focusing.

 

 

 

 

 

 

 

 Focusing algorithm

 

The focus position of the FOV is found, if the camera sees a sharp image. To reach this, the scan camera takes images in different positions of the entire focus range and the software finds the best focus interval. When this interval is found the real focus position is found by iteration of the interval.

 

Stepper motor

In opposite to the DESK, MIDI and SCAN, the focus motor of the P250 has a resolution of 6400 steps/revolution.

 

Home1,2

The Home position of the entire mechanics is situated inside the focus range. Because the number of steps never exceeds the direction limit for Home1 (the number of steps is always less then 3200 steps in each direction) the home position is always found within Home1 only. The option Home2 will never be executed during normal scan operations and is used during system integration only or before  the ex-center fixing bolt will be tightened (if mechanical adjustments are done on the ex-center). In this case, the mounting of the ex-center starts after a Home1,2 execution. This solution is important for protecting the 40x objective.

 

 

 

 

 

 

 

 

 

Shutter arms are in horizontal position

This is a special position for adjusting the physical ex-center position on the motor axle. This adjustment defines the focus range; the sine wave crosses the X-axis at 500 motor steps (see above).

 

  • Use the position of the slot in the ex-center and focus plate mounting to adjust the shutter arms into the horizontal position.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                                      Hardware limits; shutter on and off

 

By the use of two shutter arms, one for shutter on, the other for shutter off operation, the hardware limits could be held inside the direction limit for Home1 (less then 3200 steps in each direction). The hardware limits will be reached only during shutter on (CondenserCoverOn = BF light path closed, fluoresce scan) or the shutter off (CondenserCoverOff = BF light path opened, brightfield scan) procedure. To prevent the touching of the 40x objective during the shutter on procedure, the action is executed at a special specimen holder position, where the focus pin is guaranteed between the specimen holder and the slide. This position is X= 27000 steps and Y=18000 steps. This way, the specimen holder practically does not move, during the ex-center crosses the upper peak of the sine wave, because there is no connection between focus pin and slide or specimen holder. Please take this into account if you are working with the service program!!

 

 

       How to define hardware limits

 

 

 

 

 

 

 

 

 

 

                   Focus range

In earlier developed scanner systems (DESK, MIDI, SCAN) the step number of the stepper motor was identical with the actual focus position and this was displayed by the software. The focus range in these systems is 800 steps and the positions of the ex-center in relation to the displayed focus range is identical; the physical number of steps is the same as the displayed focus position in the software.

 

Because the resolution of the motor is increased by twice (1 revolution of the motor axle is done in 6400 steps), the focus range is now 1600 steps, in the P 250.

Because the focus range is physically counted from -300 steps to +1300 steps, and negative numbers as focus position are unusual, the entire focus range is shifted in the software to the focus range from 1050 to 2650; add to the physical step number 1350. 

 

In the Pannoramic 250:    SW focus position = HW stepper motor position + 1350.

 

 

 

 

 

                   Nominal focus position

Because the specimen holder of the scanner P250 can hold slides with a thickness of 0.95 mm to 1.2 mm, the nominal focus position was modified also. To ensure, that all kind of slides can be scanned in focus, the slide thickness is important during the adjustment of the objective position. Depending on the used slide thickness, the objective position should be adjusted at the relevant nominal focus position.

  • Measure the thickness of the slide with tissue to be used (but without the cover slip); use the caliper.
  • Use the “Objective position calculator” on the right to find the nominal focus position.
  • Insert the measured slide into the specimen holder and set the appropriate (calculated) focus position in the live view of the program SlideScanner.exe
  • Adjust the objective position to the preset focus position until a focused image can be seen in the live view.

 

       Check or adjust the objective position

 

 

 

 

Components and mechanical construction

 

                                 Focusing part

 

                   Stepper motor

All stepper motor relevant information, like step number and direction; go to Home1,2 or shutter on and off and the state of the sensors Home1 and Home2 are transferred via the stepper motor cables; these are connected to the “Z” connectors of the “X-Y-Z-ND-Flash” controller unit.

In opposite to the DESK, MIDI and SCAN, the focus motor of the P250 has a resolution of 6400 steps / revolution; the solution of the sensors Home1 and Home2 is not modified.

 

 

 

 

 

On the ex-center mounting the two shutter arms are situated for the shutter off and shutter on operation; so the focus motor can not do a full revolution in any direction.

 

 

       Stepper motor

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Ex-center

 

The motor axle of the stepper motor rotates the ex-center on which the focus plate is mounted and this moves the 4:1 divider.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Attention

The mountings of the focus plate should not be loosened; here the position of the focus plate and so the position of the 4:1 divider and the position of the focus pin is adjusted!

Because the movement of the focus pin is very small, the adjustment of the focus pin position is delicate; the focus plate mounting bolts position must not be altered.

·       The adjustment of the focus pin position can be done in the factory only!

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Focus plate and 4:1 divider

 

The connection between focus plate and divider is realized with a ball-headed joining because the ex-center does an up-down and additionally a left-right movement.

The connection spring ensures a slippage-free connection between focus plate and 4:1 divider.

 

The connection spring ensures a slippage free connection of the ball headed joining.

 

 

Attention

The mountings of the focus plate should not be loosened; here the positions of the focus plate, the position of the 4:1 divider and so the position of the focus pin are adjusted!

Because the movement of the focus pin is very small, the adjustment of the focus pin position is delicate; the focus plate mounting bolts position must not be altered.

·       The adjustment of the focus pin position can be done in the factory only!

 

 

 

 

 

 

The bearing of the 4:1 divider and the ball-headed pin of the focus plate eliminates the unwanted left or right movement.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   4:1 divider

 

On the other end of the 4:1 divider the focus pin is realized. The focusing of the tissue is done by moving the focus pin toward or away from the objective; the mechanical construction allows a focus pin movement of 0.3 mm over all. Because the focus pin touches the slide always during focusing, the distance of the slide with tissue to the objective is modified until the focus position is found. The focus pin movement between the focus limits is limited to be 0.15 mm around the nominal focus position (the 500 motor steps position).

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Objective changer part

 

 

The objective changer part of the focus unit consists of:

 

 

 

Important

The drive unit can be separated from the objective changer mechanics unhesitatingly, but do not remove the objective changer mechanics from the focus part; see Align the objective into the optical axisfirst!

 

 

 

 

                       Functional overview and construction

 

The following chapters, concerning the objective changer are true for the “MIDI with objective changer” (3Dmic10) and “SCAN with objective changer” (3Dmic11) also.

 

 

                   Drive unit (from top)

The transport disc is rotated by the DC motor; always in the mathematical positive direction, if we are looking onto the objectives (during normal work).

The transport disc moves the transport pins situated in the transport holes, by a half revolution.

Just before the working position of the objective is reached, the DC motor will be switched off by the appropriate sensor and the final movement, into the exact working position (into the optical axis), is done by the form-fit mechanism.

 

  • The transport disc can be rotated manually, but do it carefully!
  • The unit does not need maintenance or adjustments.

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Electronics cover

The spacer in the electronics cover keeps a distance between cover and PCB and avoids so a shortcut in the electronics.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Drive unit (from beneath)

 

  • The control electronics get its information via the cable DOJ-1 and the bus connector; the bus is prolonged to the turret unit via the cable FCJ-1.

 

 

  • The objective changer type is defined in the file “MicroscopeConfiguration.ini” in the section:

 

[Microscope]

.

.

.

ObjectiveChangerType=OC_2Pos

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Objective changer mechanics

 

  • The objective disc holds the objectives and is driven by the fitting pins.
  • The form-fit arm, together with the ball bearing of the fitting pin guarantees the proper and constant objective position in the light path, even if the objective has been exchanged.
  • The objective position can be adjusted with the objective mounting nut and is hold constant with the lock nut; see also “The objective nuts”.

 

  • The unit mounting of the objective changer mechanics should not be loosened or removed, because the working position of the objective in relation to the optical axis is adjusted here; see also Align the objective”.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Objective position sensors

 

As mentioned above, on the top of the transport disc two permanent magnets are situated; the distance to each other is exactly 180 degrees (not shown correctly).

Just before the working position of the objective is arrived and the dead point of the form-fit mechanism is even crossed, the permanent magnet arrives over the sensor and stops so the movement of the transport disc.

 

  • Each objective (position) has its own sensor and its own permanent magnet also; the positions are named as “Objective position 1” and “Objective position 2”.
  • In the objective position 1 always the 20x objective has to be mounted; in the objective position 2 the dummy objective or the 40x objective is situated.
  • The objective positions 1 and 2 of the objective disc are decided, if the drive unit will be mounted; see also “Mount the objective changer drive unit”

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Adjustable permanent magnet position

In scanners, manufactured since June 2013, the position of the permanent magnet is adjustable.

 

With this solution, and by adjusting the magnet position, the dead point of the form-fit mechanism can be always reached surely.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Position of permanent magnet

 

·      If one permanent magnet position is moved backward (or forward) the permanent magnet in the opposite position should be moved also backward (or forward) by the same distance!

·      To reach a correct adjustment, the distance of the permanent magnets to each other has to be always 180º!!

 

 

With this solution, and by adjusting the magnet position, the dead point of the form-fit mechanism can be always crossed surely.

 

       Adjust permanent magnet position

 

Watch video:                        P250_Objective change

 

 

·      If one permanent magnet position is moved backward (or forward), the permanent magnet in the opposite position should be moved also backward (or forward) by the same distance!

·      To reach a correct adjustment, the distance of the permanent magnets to each other has to be always 180º!!

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Adjust permanent magnet position

With the service program rotate the Transport Disc in forward direction.

If the motor is switched off, the form-fit mechanics should act and the final objective position should be reached surely.

Otherwise, a small gap should also exist between transport hole and transport pin, if the objective stays in the optical axis.

 

Check this behavior in both objective positions several times with the service program!

 

Watch video:                        P250_Objective change

 

 

·      If one permanent magnet position is moved backward, the permanent magnet in the opposite position should be moved also backward by the same distance!

·      To reach a correct adjustment, the distance of the permanent magnets to each other has to be always 180º!!

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Objective disc

 

The objective disc realizes the exchange of the objective between two slide scanning sessions.

If the transport disc (not shown) rotates by a half revolution, the objective disc will be rotated also as long as the connection between transport hole and objective fitting pin exists. The force of the form-fit mechanism is defined by its spring and this holds so the objective (the ball bearing of the fitting pin) always in the proper position until a new objective change command is executed.

 

 

·       The objective disc can be rotated manually in any direction to reach the required objective position.

·       The form-fit mechanism should always fix the objective position; otherwise, the optical axis will be incorrect!

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Form-fit mechanism

 

The form-fit mechanism acts in the transport disc’s stop position; the final objective position (the real working position) will be reached and the fitting pins will be disconnected from the transport holes.

The force of the spring as well as the shape of the form-fit arm guaranties always the proper and constant position of the objective in relation to the optical axis.

 

 

            Align the objective

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

The disc is always driven in direction from the spring to the mounting of the form-fit arm; the reason for this is the shape of the form-fit arm.

The use of ball bearings on essential, important mountings guarantees the proper position of the objective in the image path.

 

If the ball bearing of the fitting pin arrives to the form-fit arm, and the dead point of the form-fit mechanism is even crossed, the transport wheel stops the movement and the ball bearing of the fitting pin will be locked in the shape of the form-fit arm by the force of the spring.

 

 

 

 

The transport hole of the transport wheel moves the objective disc with the help of the objective fitting pin until the appropriate magnet arrives over its sensor. This position is so defined, that the dead point of the form-fit mechanism is even crossed.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Objective nuts

 

The objective nuts holding the objectives, fixing the adjusted objective position and existing in 2 versions:

 

  • The first version is used for objectives with a thread of M27; this is the actual version, used for the 20x, 40x objectives and the dummy objective as well; and
  • The second version with Whitworth thread is used for earlier types of Plan-Apochromat 20x objectives. Because the production of Plan-Apochromat 20x objectives with Whitworth W0.8"-1/36" thread was discontinued, today only objectives with a thread of M27 are used.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

In newer versions (since 2014) the counter nut of the objective nut is replaced by a fixing clamp.

Now only 1 wrench is used to adjust the objective position!

 

 

 

 

·       By using fixing clamps, the correct objective position can be found easily.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Solution since spring 2014

In this version, the objective nuts are fully removed; the objective disc contains 2 drillings with M27 thread, so the objective can be mounted directly.

The distance of the objective to the focus pin (the objective position) is adjusted by using fitting rings between the objective mounting surface and the objective disc surface.

 

The nominal distance is 0.3mm.

By using and combining fitting rings of different thicknesses, the objective position can be adjusted individual (not recommended!).

 

If the objective was removed, please insert the 20x objective into the mounting with the marker again, the objective position was adjusted with this condition.

 

 

 

 

 

 

 

 

 

 

 

 

 

Set of objective fitting rings

 

Name:                         SHIMS SET FOR M27 OBJECTIVE

Article number:     AC-OBJ-SHM-0100

 

Content:                  Nominal objective position is reached if the thickness of the fitting ring is 0.3mm!

 

                   2x     0.3mm; already implemented

                   4x     0.1mm

                   2x     0.05mm

 

By using and combining appropriate fitting ring thicknesses the distance of the objective to the tissue can be adjusted in steps of 0.05mm.

 

·      Please use always the same distance of the objective to the tissue on both objectives!

·      Because the Objective distance is already adjusted (with the fitting rings of 0.3mm), the implemented fitting rings are not found in the set, delivered with the scanner!

 

 

 

 

 

 

 

 

 

                   Mount the objectives

 

Important

 

Principally, the objective position is already adjusted during the system integration process and modification of the objective position should be done only in exceptions; modification is strongly not recommended!

·      If the objective(s) are dismounted, please leave the fitting ring(s) on the objective (to avoid a mismatch)!

·      Always drive the 20x objective into the marketed position, see above!

·      Always drive in the objective until it stops!

 

The focus position of both objectives is found in the nominal focus position for the appropriate slide thickness within a tolerance of ±50 focus steps!

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Dummy objective

 

If the option 40x magnification is not used, the dummy objective is implemented in the objective position 2.

This way, a weight displacement of the objective disc will be avoid.

  • If a weight displacement of the objective disc would occur, the form-fit mechanism would not work correctly and the objective may be misaligned in the optical axis.

 

 

 

 

Mount the 20x objective always into the position 1 and the 40x or dummy objective into the position 2.

  • Because in all systems P250 always a 20x objective is used (the 40x objective is optional) the start-up procedure of the software calibrates the system with the 20x objective.
  • Furthermore, all the optical and preview adjustments are based on the 20x objective.

 

  • The objective positions 1 and 2 are decided, if the drive unit will be mounted; see also “Check / Set the working position 1”

 

Remark

Since the software version 1.16 the BF scan session may be executed with the 20x or the 40x objective likewise.

 

       Upgrade to the software version 1.16” and “What is new” “Brightfield scan with the 40x objective

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

The software setup of the objectives is done in the dialog “Microscope settings”, there will be defined if the 40x objective is present or not.

 

       Pannoramic_250_1.15_User's_Guide_EN_Rev1_FINAL.pdf

 

 

Remark

In the dialog “Microscope settings”, any objective (20x or 40x) can be set to any position (objective position 1 or 2), but we recommend the use of the objective 20x in the position 1 and the 40x or dummy objective in the position 2, because some adjustments (e.g. the preview calibration procedure) requires the existence of the 20x objective in the position 1.

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Objective position

 

The objective position can be modified according to the requirements of the used objective.

 

  • To tighten or loosening the objective nut, hold still the objective mounting nut with one wrench and loosen or tighten respectively the lock nut with the other wrench.
  • By rotating the objective nut the objective position will be modified and adjusted.
  • Measure the thickness of the slide (without cover slip) to be used to adjust the objective position and calculate the nominal focus pin position, this should be used as focus step number in the live view of the scan program.
  • Insert the measured slide, create the live view, find the tissue and set the calculated number of focus steps.
  • Rotate the objective nut toward or away from the tissue, until the tissue becomes in focus.
  • Tighten the lock nut; in newer solutions, tighten the objective position fixing clamp.
  • Find the real focus position by executing the option “Auto focus”.
  • Correct the objective position as necessary by repeating previously explained steps.

 

 

 

 

 

After finishing the objective position adjustment check the tightness of each objective!!

 

 

      Adjust the objective position

 

       Setup and define the implemented objectives

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Adjustments of the objective changer unit

 

The mechanics of the objective changer unit does not need adjustments; the positions of the Hall sensors are fixed as well as the positions of the permanent magnets.

 

The adjustment is limited to the following parts:

 

 

 

Check the working position 1

1.      With the service program set the Y-stage to Home1,2.

2.      Set the X-stage to Home1,2.

3.      In the tab “Objective changer”, select the sensor 1 and press the button “Forward”; the 20x objective should move into the working position of the objective changer unit, in the specified movement direction and the form-fit mechanics should snap in correctly.

 

Set the working position 1

 

1.      If the working position 1 is not correct after the check, remove the entire focus unit with objective changer.

2.      Remove the objective changer drive unit (probably the stepper motor cables must not be disconnected).

3.      Rotate the objective disc by a ½ revolution so, that the 20x objective stays in the working position.

4.      Mount the objective changer drive unit.

5.      Mount the entire focus unit with objective changer.

 

 

 

 

 

Remove the objective changer drive unit

 

1.      Remove the focus unit.

2.      Remove the mounting bolts of the objective changer drive unit.

3.      Easily separate the drive unit from the objective changer mechanics.

 

 

 

 

Mount the objective changer drive unit

  • With the mounting of the drive unit the position of the 20x objective will be defined also.

 

1.      Connect the cable DOJ-1 to the bus connector.

2.      In the tab “Objective changer”, of the service program select the sensor 1 and press the button “Forward”; see above.

3.      Rotate the objective disc manually so, that the 20x objective is in the working position of the objective changer unit.

4.      Fit the objective changer drive unit onto the objective changer mechanics and drive in the mounting bolts.

5.      Mount the entire focus unit with objective changer.

6.      Check the correctness of the objective changer and its mechanics manually and with the service program also.

7.      If the form fit mechanism does not snap in during the normal exchange operation, modify a little bit the drive unit mounting position until the dead point of the form fit mechanism is crossed correctly.

 

 

 

 

 

 

 

 

 

 

                   Dismount or mount the objective

 

  • The following procedure assumes that the focus unit with objective changer is mounted and should not be dismounted.
  • If the focus unit with objective changer is already removed from the scanner plate, you can remove the drive unit and so dismount or mount the objectives.

 

1.       With the program SlideScannerService.exe Set the X-Y-stage to the positions Y-Home1,2 and X-Home1,2.

 

2.      In the tab “Objective changer” move the objective (position), to be dismounted or mounted into the working position.

 

3.      Select the option “Time out”, set a numerical value of 1300 ms and press the button Backward.

 

 

4.      Dismount or mount the objective. Hold the mounting nut with the delivered wrench and loosen or tighten the objective by hand. Before mounting a brand new objective, use a little bit grease on the mounting thread.

 

5.      If the objective nut position was previously not adjusted, check the distance of the objective to the focus pin. If the focus pin seems to touch the objective, correct the distance by rotating the objective nut; the focus pin should never touch the objective!

 

6.      Adjust the distance of the objective to the plane of the focus pin until a gap of approx. 4 mm is reached; this prevents the objective against touching the specimen holder.

 

7.      Select the appropriate sensor and press the button forward; the actual dismounted position or the mounted objective should move into the working position.

 

8.      Repeat the steps 2 to 7 with the other objective (position), if required.

 

9.      If the 40x objective was mounted, check and set the thickness of the cover slip; see also “The 40x objective.

 

 

 

 

 

                   Check and adjust the objective position

  • This adjustment assumes that the ex-center position of the focus unit is adjusted well and so, the sine wave crosses the X-axis at +500 steps and the appropriate objective is mounted.

To adjust the focus position you can use a slide with a thickness in the range from 0.95 to 1.20 mm. Important is, that the slide thickness, used to adjust the objective position, is measured and known. The focus position will be calculated with the table “Objective position calculator”. The cover slip is excluded from the slide thickness measurement!

 

10.  Start the program SlideScanner.exe with the service password.

 

11.  Clean the cover slip and the slide bottom, then insert the measured slide with the known thickness.

 

12.  Check the correct holding of the slide in the specimen holder!

 

13.  Select a FOV of the tissue inside the preview window with the positioning tool.

 

14.  Select the focus tab and live view.

 

15.  Press auto focus.

 

16.  If the found focus position is equal with the calculated focus position for that slide (a deviation of + - 25 steps is possible) the objective position does not need adjustments; the check is finished.

 

17.  If an adjustment of the objective position is required, set the calculated number of focus steps in the live view, loosen the lock nut of the objective mounting and drive the objective mounting nut so, that the tissue becomes into focus (toward the cover slip or away from it respectively).

 

18.  If the tissue becomes into focus, tighten the lock nut; press the button “Auto focus” and repeat from step 15.

 

19.  Check the result again by pressing the button “Auto focus” some times.

 

20.  If there are more then 25 steps difference to the nominal (calculated) focus position, repeat the adjustment from step 15.

 

21.  Check the focus position on different positions of the tissue.

 

22.  Change to the other objective position (tab “Service”, “Microscope control” select the other objective position and go back to the tab “Focus”, adjust the exposure time) and check or adjust the focus position for the other objective by repeating the previous adjustment from step 13.

 

23.  If the 40x objective was adjusted, check / select the thickness of the cover slip; see also “The 40x objective

 

24.  After finishing the objective position adjustment check the tightness of each objective!!

 

25.  If the adjustment is done, please move the objective changer to the opposite objective position and again into the actual position (since the software version 1.16 it is possible in the scan program) and start the option “Auto focus” again. Only this way you can be sure that the actual objective is right aligned in the optical axis.

 

Remark

Because we handled the objective manually with the wrenches, it is easily possible to misaligning the optical axis!

In earlier versions of the file “SlideScanner.exe” you should start the scan program with the service password and then, in the tab “Service” and “Microscope control” you can exchange the objective by software also. Then check the focus position again with the function “Auto focus”.

 

 

 

 

 

 

                  Remove the condenser

  • Remove the focus unit (optional).
  • To remove the condenser, loosen its fixing and drive the condenser out; away from the objective.

 

 

                  Mount the condenser

·         To mount the condenser, put it into its holder and drive the condenser in; toward the objective.

·         Mount the focus unit.

 

 

 

 

 

 

 

 

 

 

 

 

Condenser position adjustment tool

 

·         The condenser position adjustment tool is used to find the optimal condenser position.

·         The adjustment tool is put onto the collimator lens housing during the condenser position adjustment and if the center drilling of the adjustment tool is in focus, the condenser position is nearly found.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Adjust condenser position

The adjustment of the condenser is important for the bright, uniform and optimal illumination of the FOV. This reduces so the exposure time of the camera and increases the quality of the scanned tissue.

The position of the condenser affects the following:

  • The size of the visible FOV (color shading)
  • The image contrast
  • The image resolution (the numerical aperture) and
  • The exposure time.

 

  • We assume that the focus unit and the objective changer are already adjusted; the position of the objective is correct.

 

  1. Use the program SlideScanner.exe and insert a slide with cover slip, create a live view and click inside the tissue.
  2. Use the tool “Auto focus”.
  3. If the tissue is in focus, find a “clean” FOV (without tissue or dust). Use the positioning tool  and click outside the tissue but inside the cover slip of the preview window in the program “SlideScanner.exe”.

 

  1. Loosen the condenser’s fixing bolt and put the condenser position adjustment tool over the collimator lens holder.

·      If you can not see the hole of the condenser position adjustment tool, calculate the exposure time and move the adjustment tool on the optical axis until the image of the drilling or a part of it can be seen in the live view.

 

  1. Rotate the condenser toward to the objective, so the start position for the adjustment will be found; the brightness will increase.

 

  1. Rotate the condenser backward and look at the live view. Beware of the condenser cover (shutter), don’t close it and don’t bend it. You will see, that the drilling of the condenser position adjustment tool comes into focus. While rotating the condenser you can also observe that the brightness decreases.

 

 

 

 

 

 

 

 

  1. If the center drilling of the adjustment tool is in focus, the condenser position is nearly found, a small correction of the condenser position have to be done, because the focus position of the drilling is not the real position of the collimator lens. Drive the condenser by about ¼ revolution more backward and stop the move of the condenser; tighten its fixing bolt.

 

  1. Check the correctness of the adjustment in the live view and by scanning a tissue or a part of it.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Alternatively, if the focus position of the tissue is correct, you may stick a slice of paper onto the collimator lens holder.

·       Rotate the condenser, until the fibers of the paper are in focus.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

  • The unevenly illumination of the “clean FOV” will be corrected by software (the right border is a little bit darker than the left side).

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Shutter mechanics

 

 

                   General

The mechanical shutter is implemented as a part of the focus unit and the shutter guide is situated between condenser and 4:1 divider.

 

During fluorescent scanning the shutter must be closed and insures so a dark background. Other, unwanted fluorescent materials (e.g. painting, optics) can not reflect the fluorescent light or can not fluoresce and so they will not disturb the fluorescent view.

 

During bright field illumination the shutter must be fully open to ensure a bright and evenly illumination of the FOV.

 

 

 

 

 

 

 

 

 

 

                   Principle of shutter mechanics

 

The shutter has only two usable states, shutter on and shutter off. These positions are identical with the hardware limits of the focus unit; in these positions the bright field illumination should be fully opened or fully closed respectively. All other possible positions of the shutter are not defined, therefore faulty positions and not allowed during the scan process.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

During rotation of the stepper motor axle in direction to the negative limit of the focus unit, the shutter wire will be touched by the shutter arm and is so moved to the shutter off state (bright field illumination allowed). If the stepper motor axle is rotated in direction to the positive limit, the other shutter arm will touch the shutter wire and moves it so to the shutter on position (bright field illumination disabled).

In practice it means, that the shutter wire together with the shutter arms is the limiter of the focus mechanics and with this limiter the shutter on / off option is realized.

 

  • During focusing the shutter wire will never be touched by the shutter arm.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

A = Focus pin

B = Shutter guide

C = Shutter wire

D = Condenser

E = Condenser fixing

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Shutter guide

 

If the shutter wire is touched by the shutter arm, the torsion is transmitted via the shutter wire to the slider of the shutter guide. The slider brings so the shutter guide into the proper position.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Adjustment procedures

 

Adjustments of the focus part and shutter mechanics

 

 

 

                   Adjust the ex-center position

This adjustment must be done only if the ex-center was dismounted or the fixing bolt was loosened. With this adjustment the focus position is set to be +500steps nominal.

Because in this position the fixing bolt can not be loosened or tightened, we use the focus stepper motor position of 2100 steps for this adjustment.

 

 

1.      Loosen the ex-center fixing bolt so, that the ex-center can be moved on the motor axle.

 

2.      With the service program set the focus unit to Home1,2.

 

3.      Go forward to the focus motor position 2100 steps.

 

4.      Bring the ex-center into the horizontal position (the shutter arms shows to the left and to the right; use the slot of the ex-center mounting to find the proper position. Before you are tighten the ex-center fixing bolt please check the position of the shutter arm in relation to the shutter wire also. The shutter arm should touch the shutter wire nearly in the middle of the hook. To reach this, bring the ex-center in proper position on the motor axle and tighten the fixing bolt.

 

 

 

 

 

 

 

 

Attention

 

A small gap between motor mounting surface and ex-center mounting should exist, otherwise, if the ex-center mounting rubs on the motor mounting surface, the focus unit may lost rotor movements and so the scanned tissue may be out of focus or blurry, even if the focus rotor direction had changed.

 

5.      Go to the positions Home1,2.

 

6.      Go forward to the focus motor position +500 steps.

 

7.      In this position the shutter arms should showing exactly up and down. A deviation of approx. 10 steps from the vertical direction is allowed. If the deviation is too much, repeat the adjustment from step 1.

 

 

 Find the hardware limits for the focus unit

This procedure must be done only if the ex-center was dismounted or its fixing bolt was loosened. Before the following procedure is done, the previous adjustment “Adjust the ex-center position” should be correct.

·       The path of the file MicroscopeConfiguration.ini, in the software version with the operating system Windows® 7 is:

C:\ProgramData\3DHISTECH\SlideScanner\MicroscopeConfiguration.ini

 

 

       How to define the hardware limits

 

 

 

Find the negative limit

 

8.      With the service program set the focus unit to Home1,2.

 

9.      Go forward to the focus motor position -1750 steps.

 

10.  Go backward +1750 steps.

 

11.  Press Home1 (only). There should be not more then +-2 steps difference to Home1.

If there are more steps lost, decrease the actual number of steps by 10 and repeat from step 8. In the steps 9 and 10 use the actual number of steps!

 

12.  If there are not more then 2 steps difference to Home1, increase the number of steps by 10 and repeat from step 8. In the steps 9 and 10 use the actual number of steps!

 

13.  The negative limit is found correctly if the motor movement has no steps lost and the actual number of steps, increased by 10 would produce lost steps.

The found negative limit can differ by more ten steps from unit to unit. The reasons are the tolerances of the components and the used position of the ex-center in relation to the motor axle.

 

14.   Update the parameter value of the parameter “FocusDeviceMin” with the found number of the actual steps in the file “MicroscopeConfiguration.ini” section [HardwareLimits].

 

15.  Update the parameter value of the parameter “CondenserCoverOff” with the found value in the file “MicroscopeConfiguration.ini” section [Focus].

 

 

Find the positive limit

 

16.  With the service program set the focus unit to Home1,2.

 

17.  Go forward to the focus motor position +2750 steps.

 

18.  Go backward 2750 steps.

 

19.  Press Home1 (only). There should be not more then +-2 steps difference to Home1.

If there are more steps lost, decrease the actual number of steps by 10 and repeat from step 16. In the steps 17 and 18 use the actual number of steps!

20.  If there are not more then 2 steps difference to Home1, increase the number of steps by 10 and repeat from step 16. In the steps 17 and 18 use the actual number of steps!

 

21.   The positive limit is found correctly if the motor movement has no steps lost and the actual number of steps, increased by 10 would produce lost steps.

The found positive limit can differ by more ten steps from unit to unit. The reasons are the tolerances of the components and the used position of the ex-center in relation to the motor axle.

22.  Update the parameter value of the parameter “FocusDeviceMax” with the found value in the file “MicroscopeConfiguration.ini” section [HardwareLimits].

 

23.  Update the parameter value of the parameter “CondenserCoverOn” with the found value in the file “MicroscopeConfiguration.ini” section [Focus].

 

 

 

 

 

 

 

Dismount; mount the focus unit

 

 

 

 

                   Focus unit mounting

 

The mounting of the focus unit with objective changer and the mounting of the X-Y-stage is realized with dovetails; these are hold by dovetail fixing clamps.

 

  • Rotate the screw driver clockwise to open (loosen) the dovetail clamp.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   Dismount the focus unit

 

  • Make sure, that the X-Y-Stage is not in the X-home position, because in this position the focus unit can not be removed.

 

1.      With the service program go forward to the position 10,000 steps in +X direction (3 full turns of the motor axle).

 

2.      Disconnect the bus cables DOJ-1 of the objective changer and FCJ-1 of the turret stepper motor.

 

3.      Disconnect the cables of the focus motor (optional).

 

4.      Loosen the fixing bolt of the focus unit mounting; turn the screw driver clockwise!

 

5.      Remove the entire focus and objective changer unit by pulling the unit backward.

 

            Watch Video:           Remove focus unit (SCAN)

 

 

 

                   Mount the focus unit

 

6.      Insert the focus unit with objective changer into the dove tail of the scanner plate until the physical limit is reached; take care on the objectives.

 

7.      Tighten the fixing bolt of the focus unit mounting; turn the screw driver counter clockwise!

 

8.      Connect the bus cables DOJ-1 of the objective changer and FCJ-1 of the turret stepper motor.

 

9.      Connect the cables of the focus motor.

 

            Watch Video:           Mount focus unit (SCAN)

 

 

 

 

Setup and define the implemented objectives

 

 

 

 

If the objective(s) are implemented and the program “SlideScanner.exe” finished the start up procedure, select the option “Microscope Settings” in the menu “Options”.

 

  • The setup of the objective(s) is done mostly together with the install of the present cameras.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

In the dialogue “Base Settings” the implemented objectives are selected and parameterized.

 

By default and in the scanner P250

  • Select the type “Plan-Apochromat” (1) and the magnification of 20x (2) in the objective position 1
  • Select the type “Plan-Apochromat” (3) and the magnification of 40x (4) in the objective position 2.

§         If there is only a 20x objective implemented into the scanner, select “<none>” (3) in the objective position 2.

 

Remark

Of course, the dialogue accepts any kind of magnification in any objective position, but in the Pannoramic 250 the 20x objective should be always in the objective position 1 because this is the default objective and some adjustment procedures (e.g. the preview calibration process) uses the 20x objective in the objective position 1!

 

 

 

 

 

 

 

 

Align the objective into the optical axis

 

 

 

Attention

The alignment of the objective position to the optical axis is already done and finished; this is not a usual adjustment.

 

The following calibration should be done only, if the objective changer unit mounting was loosened, altered or the objective changer unit was separated and reassembled to the focus unit; so the optical axis will not be strait.

 

  • The following procedure will be used to fit the objective (position) into the optical axis correctly.
  • In any cases, check the alignment first, before starting the adjustment.
  • Check the correct working of the form-fit mechanics also.

 

Remark

The aligning procedure is advised and described for the 20x objective and the objective position 1.

If the 20x objective position is equipped with an objective nut using Whitworth thread, you can use unhesitatingly the 40x objective position to align the objective changer unit!

 

 

 

 

 

 

 

                   Alignment tools

The alignment is done mechanically; the objective pupil position (represented by the drilling of the objective aligner) will be fit into the optical axis by the help of the condenser aligner pin. During this fitting operation the objective changer unit mounting is barely loosened (except you are checking the correctness of the alignment). The unit mounting bolts will be tightened, if both adjustment tools fitting easily and the form-fit mechanics fix the objective position correctly.

 

  • In all cases, the lock nut of the objective mounting nut as well as the objective aligner should be fully tightened during the aligning procedure.
  • The alignment tools can also be used to check the objective position alignment if the objective changer unit mounting is tightened!

 

 

 

 

 

 

 

 

 

                   Check the alignment

1.      With the service program set the objective changer into the default position by pressing the radio button “Sensor 1” and the button “Forward” of the tab “Objective changer”; the objective 20x should be in the working position.

2.      Disconnect the cables of the objective changer and the stepper motor cables.

3.      Remove the entire focus unit with the objective changer.

4.      Remove the drive unit.

5.      Rotate the objective disc manually by about a ¼ revolution and remove the 20x objective; do not loosen the lock nut or the objective mounting nut; otherwise the objective position has to be adjusted again!

6.      Drive the objective aligner into the objective mounting of the 20x objective until it stops.

7.      Mark the condenser position (e.g. with a pen), then remove the condenser.

8.      Make sure, the mechanical shutter is open.

9.      Insert the condenser aligner into the condenser mounting.

10. If the condenser aligner moves easily in / out in the condenser mounting (if it would falling out by gravity) and the form-fit mechanics fix the objective position correctly, the adjustment is correct.

11. If the condenser aligner can not be inserted or moves strong, the objective is misaligned; execute the aligning procedure.

12. If the aligning result is acceptable, remove the condenser aligner and mount the condenser again (until the previously signaled position).

13. Remove the objective aligner and mount the 20x objective again.

14. Put the 20x objective into the working position manually and mount the drive unit.

15. Mount the entire focus unit with objective changer and connect the cables.

16. With the service program check the functionality of the focus unit and the objective changer unit as well, the 20x objective have to be in the working position if ‘Sensor 1” was selected.

17. Check or adjust the objective position.

18. Check the focus position

19. Check or adjust the condenser position.

 

 

 

 

 

                   Aligning procedure

 

1.      With the service program set the objective changer into the default position by pressing the radio button “Sensor 1” and the button “Forward” of the tab “Objective changer”; the objective 20x should be in the working position.

2.      Disconnect the cables of the objective changer and the stepper motor cables.

3.      Remove the entire focus unit with the objective changer.

4.      Dismount the drive unit.

5.      Rotate the objective disc manually by about a ¼ revolution and remove the 20x objective; do not loosen the lock nut or the objective mounting nut; otherwise the objective position has to be adjusted again!

6.      Drive the objective aligner into the objective mounting of the 20x objective until it stops.

7.      Mark the condenser position (e.g. with a pen), then remove the condenser.

8.      Loosen the unit mounting bolts of the entire objective changer unit so, that the unit becomes barely moveable.

9.      Make sure, the mechanical shutter is open.

10. Insert the condenser aligner into the condenser mounting and adjust the entire objective changer unit position in relation to the focus unit.

11. If the condenser aligner moves easily in / out in the condenser mounting (if it would falling out by gravity) and the form-fit mechanics fix the objective position correctly, the unit mounting bolts should be tightened.

12. Check the easily movement of the condenser aligner in the condenser mounting again; hereby the form-fit mechanics should fix the objective position correctly also.

13. If the condenser aligner moves strong, repeat this procedure from step 8.

14. If the aligning result is acceptable, remove the condenser aligner and mount the condenser again (until the previously signaled position).

15. Remove the objective aligner and mount the 20x objective again.

16.  Put the 20x objective into the working position manually and mount the drive unit.

17. Mount the entire focus unit with objective changer and connect the cables.

18. With the service program check the functionality of the focus unit and the objective changer unit as well, the 20x objective have to be in the working position if the radio button “Sensor 1” and the button “Forward” of the tab “Objective changer” was pressed.

19. Check / adjust the objective position.

20. Check the focus position

21. Check / adjust the condenser position.