Focus unit; D_II

For technicians and partly for sales managers!

 

 

                Enhancements

 

Construction of DESK_II's focus unit has changed in relation to the focus unit of DESK; since summer 2016 main modifications are:

Shutter mechanics removed

Because the scanner unit of the DESK is never able to scan slides in FL scan mode, the mechanical shutter is also not required.

In DESK, the shutter unit was implemented for the Hardware limits and to keep compatibility with SCAN and MIDI type scanners.

 

Hardware limits not required

Because the shutter wire is removed, the rotation of the ex-center is no longer limited. The ex-center may rotate unlimited

 

Ex-center position visualization pin

In the position of the shutter arm of the ex-center, now only a visualization pin is implemented to make the ex-center position adjustment possible.

 

Stepper motor type exchanged

In the DESK, there was a 2-phase stepper motor with 200 full-steps/revolution used, so a resolution of the 3200µ-steps /revolution is possible.

In the DESK_II a 5-phase stepper motor is implemented, this guarantees a higher full-step resolution,  and more smooth movement of the rotor and so, vibration and noise are also reduced.

One revolution of the of the stepper motor is now 6400µ-steps/revolution.

 

Increased slide thickness

The modified focusing solution allows now focusing of slides with a thickness of 0.95mm to 1.20mm. 

 

Modified focusing mechanics

To focusing slides with a thickness of 1.20mm, the movement range of the focus pin had to be increased.

To reach this, the relation of the divider was modified and the stepper motor's step angle was decreased.

 

 

       Enhancements_2016

 

 

 

 

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 in thickness to each other, 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 DESK type scanners 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 will be exchanged, the specimen holder must be moved away from the objective to avoid collision.

 

During slide actions, except focusing, the objective must be protected against touching the cover slip and the focus pin and the 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, controlled via the X-Y-stage unit.

 

  • Components and construction of the focus unit is mainly defined by the requirements of the objective.

 

 

Objectives

The default objective magnification in the DESK scanners is 20x, but the user can order the 40x magnification also, explicitly.

The focus unit is able to hold 2 objective types.

 

  • 20x magnification and
  • 40x magnification

The worst case is defined by the parameters of the 40x objective.

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.

 

  • The objective exchange is realized manually, so an adjustment of the objective position is required after exchange.

 

 

 

 

 

 

 

 

                   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 value, the quality of the scanned FOV may be reduced!

 

 

 

 

 

 

 

 

 

 

 

General

 

 

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

·       The focusing part  (driven by the focus stepper motor)

·       The objective with mounting nut and objective position fixing.

·       The condenser with its position fixing.

·       The focusing mechanics (Ex-center, 4:1 divider, Focus pin).

·       Dovetail unit mounting, so the correct mounting position is found again after the unit was removed and inserted again.

·       To adjust the ex-center position, a visualization pin (here named as "Pin") is implemented

 

 

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 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 chromatically and spherically corrections and delivers so an image of very high color trueness with very small spherically and chromatically 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 than 0.25mm, other kind of objectives can be used also but it is strongly not recommended!

 

·       Always check with 3DHISTECH first when a different objective should be used!

 

The exchange of the focus unit is possible

            - If the stepper motor or its electronics for the focus unit is faulty

            - If the shape of any part is deformed or a part is broken.

            - If the focus unit has any fault and you are unable to fix it.

 

 

Requirements

 

 

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!

 

 

 

 

                   Objective position

In Pannoramic type scanners the objective is mounted so, that the focus point of the objective is in the middle of the focus range, offered by the movement range of the focus pin during focusing.

 

  • It is very important, that the objective position is adjusted well; otherwise, if the objective's focus point 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 perpendicular 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.

To avoid collision of the objective with the slide during crossing the upper peak of the sine wave, 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.

 

 

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 6400µ-steps /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.

 

Remark

The connection of the ball headed joining is realized with the connection spring and the triangle (see ex-center) and not by a compression spring as shown.

 

 

 

 

 

 

 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 DESK_II has a resolution of 6400µ-steps/revolution.

 

 

Home1,2

The Home position of the entire mechanics is situated inside the focus range. The option Home1,2 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.

 

 

 

 

 

 

 

 

 

 

 

Hardware limits

 It is true, that the hardware limiters are removed from the focus unit, but to set limits for the software, the limits in the ini-file are set to -1700µ-steps and to 2650µ-steps respectively.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Visualization pin in vertical 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 focus plate mounting to adjust the visualization pin vertical, while the rotor stays in the position -1100steps.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   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 D_II.

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 DESK_II:    SW focus position = HW stepper motor position + 1350.

 

 

 

 

 

 

 

 

                      Nominal focus position

Because the specimen holder of the scanner DESK_II 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”, shown 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 / adjust the objective position

 

 

 

 

 

 

 

 

Components and mechanical construction

 

 

 

 

                   Stepper motor

All stepper motor relevant information, like step number and direction; go to Home1,2 and the state of the sensors Home1 and Home2 are transferred via the stepper motor cable; this is connected to the USB controller unit.

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

 

 

On the ex-center mounting the shutter arm is removed, only a visualization pin is situated for the shutter position adjustment; now the focus motor can not do a full revolution in any direction.

 

    5-Phase stepper motor

 

Watch video:            Stepper motor

 

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 steps and so the scanned tissue may be out of focus or blurry, even if the focus rotor direction had changed.

 

 

 

 

                  Adjust ex-center position

 

1.      Go to the positions Home1,2.

 

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

 

3.      In this position the visualization pin should showing exactly up. A deviation of approx. 10 steps from the vertical direction is maximal allowed.

4.      If not, loosen the excenter mou nting bolt, and rotate the visualization pin exactly upward and tighten the mounting bolt.

5.      Check the correct position with the service program.

6.      If the deviation is too much, repeat the adjustment from step 1.

 

 

 

 

 

 

 

                   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 only with special tools!

 

 

 

Watch video:            Ex-center and focus plate

 

 

 

 

 

 

 

 

 

                   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.

 

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).

 

Watch video:            Focus mechanics

 

 

 

 

 

 

 

Scan area

 

The scan area defines the part of the slide on which the tissue, scanned by the scan camera, is expected to be. The entire scan area is captured by the preview camera in three sections and is shown in the preview area of the program SlideScanner.exe.

The scan area is limited by the mechanical construction of the specimen holder and should be defined as large as possible; it is not defined in relation to the cover slip.

In all cases, the specimen holder should never be touched by the focus pin or the objective! In other words, the scan area is that area of the slide, on which the focus pin and the objective (especially 40x magnification) can be moved seemingly during scanning the tissues, without touching the specimen holder.

 

 

 

 

 

 

 

 

 

 

              Exchange of the objective

As mentioned before, the focus unit may be equipped with a 20x or a 40x objective. Because the focus position of the 40x objective is very close to the tissue, the scan area, defined for the 20x objective is no longer valid, to avoid collision of the objective with the specimen holder.

If the objective will be changed from a 20x to a 40x magnification, the scan area must be defined again and therefore the preview calibration must be executed again also. During defining the scan area for the 40x objective, the position of the focus pin and the position of the objective pupil should be checked to avoid touching of the specimen holder! Adjust the limits in the nominal focus position!

If the objective is changed from the 40x to a 20x magnification it is not bound to define the scan area again, if the smaller area can be accepted. In all cases, if the scan area was modified, a preview calibration is bound to do.

 

       Working distances” and “How to exchange the objective in DESK

 

Watch video:            Exchange objective

 

  1. Remove cables from the camera as required.
  2. Loosen the mirror tube mounting bolt.
  3. Remove the mirror tube rearward.
  4. Loosen the fixing bolt of the objective mounting.
  5. Remove the objective nut with objective.
  6. Exchange the objective.
  7. Insert the objective nut with objective until to the focus position.
  8. Tighten the fixing bolt of the objective mounting.
  9. Mount the mirror tube; move it frontward until is stops.
  10. Tighten the mirror tube mounting bolt.
  11. Reconnect removed cables.
  12. Adjust the focus position.
  13. Define scan area
  14. Execute preview calibration.

 

 

Hint

If the exchange of the objective is often done, the technician may create two ini files, one for the parameter values with the 20x objective and one for the values with the 40x objective.

By easily exchanging of the ini file content, many limit definitions may be left out.

The focus position adjustment of the objective is bound to do always!

 

Important

Please make a security backup of the ini file before modifying values!

 

 

 

 

 

 

 

 

 

                   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.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

                   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 is 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.

·      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.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

  • If the “clean FOV” is evenly illuminated the condenser position is correct.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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 mount focus unit (D_II)

 

 

 

                   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:           Remove mount focus unit (D_II)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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 D_II

  • 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!