Adjust the parallelogram

For experienced technicians!

This chapter describes the adjustment procedure to reach perpendicularity between the objective (optical axis) and the slide surface (specimen holder).

Because this adjustment needs special tools and might be time consuming, it should not be done in the field; an experienced technician may execute these steps in the service center.

The adjustment is based on the software version 1.15 and the scanner P250, but the principle and almost all procedures are the same in the scanners S_M_D.

Contents

General

Tools used for the adjustmen.. t

The parallelogram

Principle of the adjustm.. ent

Prepare the scanner for the adjustment

Adjust the parallelogram

Finishing the adjustment

Reassemble the scanner

Post adjustments and checks

Finishing

Adjustment techniques

Images of the adjustment steps

General

The optical axis of the objective and the glass slide needs to be perpendicular to avoid tilting and partially out of focus FOVs on digital slides.

As commonly known, in Pannoramic scanners, the slide is moved with the focus pin around the nominal focus position (around the position +500 steps; in S_M_D this position is 1600 steps nominal) during focusing. To shift the slide in the Z-direction (this means focusing) and not inclining or tilting the slide during these movements, a parallelogram is used. The parallelogram is mounted between the Y-carriage and the specimen holder.

Also important is the position of the parallelogram in relation to the X-Z-plane and the Y-Z-plane of the slide.

Best scanning results will be reached if the slide is perpendicular to the optical axis.

· The adjustment, described in this chapter, defines the position of the parallelogram so, that the inclination of the slide in the X-Z-plane and the Y-Z-plane will not exceed the allowed limits.

More information can also be found in the chapters:

· “Focus unit” and “Scan area”, “Principle of focusing”

· “X-Y-Stage unit”, “Parallelogram” , “Specimen holder”

· “Optics and illumination”, “Stage skew check”

Ideal position of the slide in the specimen holder

In the ideal position of the slide, the optical axis is perpendicular to the slide surface and by moving the focus pin in the range of -300 steps to +1300 steps or by selecting any field of view inside the scan area, this perpendicularity will not be lost; the LASER beam (reflected by the slide surface) would be always reflected in the same angle onto the same position of the screen during movements of the slide.

1) The ideal slide position would be reached if the field of view (FOV), near to the limit Y-min would be in the same plane as the field of view near to the position Y-max (shown with the line (Y))

And

2) The field of view, near to the limit X-min would be in the same plane as the field of view near to the position X-max (shown with the line (X)).

· If this requirement is fulfilled, the optical axis is perpendicular to the slide surface.

This definition assumes also:

· The slide (the glass) itself is flat, and not tilted or slant in any direction and the slide thickness does not vary.

· The force of the preload spring of the parallelogram is not much; otherwise, the pressure of the focus pin against the slide may tilt the slide surface in positions of the field of view, far away from the holding mechanics (c).

Real position of the slide

In practice, the ideal position will not be reached, but the inclination of the slide in X- and Y-direction may be reduced as much as possible.

· The slide will be inclined in the X-Z-plane, in the Y-Z-plane or in both planes at the same time, but always inside the allowed tolerances.

· In these cases, the Z-axis of the slide will be inclined, in relation to the optical axis.

The LASER beam will be reflected to a different position onto the screen (in relation to the ideal position of the slide).

· The inclination of the slide may result in unfocused parts of the virtual slide and these are best visible on the border of the actual field of view in relation to the neighbor FOV, if the inclination of any plane is too much.

Faulty inclination of the slide in the specimen holder

The real position of the slide is faulty if:

· The Z-axis of the slide is inclined, in relation to the optical axis; outside the allowed limits.

The inclination angle of the slide’s X-Z- or Y-Z-plane is too much, if the focus of the field of view differs (the focus difference is visible) on the border to the neighbor field of view in the virtual tissue and:

· The slide is correctly inserted into the specimen holder

· The slide is held properly in the specimen holder (the slide does not change its position during scanning)

· This behavior can be detected often on two neighbor FOV’s on different positions of the scanned tissue; the border of the FOV is visible (in relation to the focus).

· The neighboring FOV’s are scanned in the same focus motor step position

· Some parts of the field of view are out of focus while other parts of the same field of view are in focus

· This behavior can be found on different tissues (slides) also.

· By adjusting the parallelogram position, the inclination of the slide in relation to the optical axis will be minimized or eliminated; this is the task of the adjustment.

Remark

· During the tissue creation process the thickness of the sample itself may vary on small spots and this might also be visible as unfocused part of the virtual tissue. To decide, whether the thickness of the sample is varying or the inclination of the slide is the cause of the error, please check always the focus on the border of the FOV in relation to its neighbor FOV on different tissues also.

· The inclination of the specimen (holder) may also be checked in the live view and with a zoom factor of 2.73 of the scan program; see also: “Stage skew check”.

In all cases

If the scanned tissue shows mistakes or errors, that may occur in conjunction with non perpendicularity of the objective (Optical axis) and the specimen holder (slide surface) (some parts of the field of view are out of focus during other parts of the same field of view are in focus) and this behavior can be detected very often on two neighbor FOV’s and there is no difference in focus steps, the following checks have to be done first:

Please do the following checks before you set up the adjustment tools!

Check the correct tightness of the X-Y-stage unit; loosen it, check the dovetail mounting for unexpected behavior (smoothness, dust) move it into the correct position again and tighten the fixing bolt.
Check the correct tightness of the focus unit; loosen it, check the dovetail mounting for unexpected behavior (smoothness, dust) move it into the correct position again and tighten the fixing bolt.
Check the correct position and the tightness of the scanner plate; loosen it, move it into the correct position again and tighten the fixing bolts in the right sequence.
Check the correct position and the tightness of the turret unit; loosen it, move it into the correct position again and tighten the fixing bolts in the right sequence.
Insert a slide into the specimen holder and check the correct fixing of the slide in the specimen holder (the slide should not be inserted slanted!)
Check the hysteresis in X- and Y-direction.
Check a well visible part of the tissue in the “Live view” window as described in the chapter “Stage skew check”.
Scan the tissue of the well inserted slide and analyze it for slanted field of views.

Any mechanical reason or incorrectness named above may result in the incorrectness of the optical axis; if one component is not placed well in the optical axis, the result is the same as if the parallelogram would be adjusted wrong!!

When shell I check or adjust the parallelism between the focus- and the X-Y-stage unit?

To check the parallelism, use always the “Stage skew check” first, before you set up the adjustment tools!

If the “Stage skew check” failed or a faulty inclination of the slide in the specimen holder was detected.
If the parallelogram was removed.
If the parallelogram or the specimen holder was exchanged.
If the entire X-Y-stage unit was changed.
If the Focus unit was exchanged.
If any spare part was changed and this spare part is in connection with the parallelism or the straightness of the optical axis.
If the mounting bolt positions or the adjustment bolts position of the parallelogram was altered.

Attention

If the parallelogram or the specimen holder is deformed (bent) or any spring of the parallelogram is broken or deformed, this adjustment will not deliver the desired results!

Exchange the appropriate spare part first before doing the adjustment steps.

Important

If any modification on the parallelogram or the specimen holder mounting was done, please:

· Check or adjust the magazine unit position.

· Check or adjust the correct position of the specimen holder fixing fork.

· Check or define the scan area again before scanning tissues!

· If the scan area was modified, please execute the preview calibration also.

· Check the darkfield preview.

Requirements

· The tools, described in the chapter “Tools used for the adjustment”.

· Diverse hexagon key bolt drivers

· Service program “SlideScannerService.exe”

· Scan program (SlideScanner.exe)

· Viewer program (“SlideViewer.exe”)

· Required time may be 2 hours or more.

Remark

The required time depends highly on the misalignment of the parallelogram, the post adjustments and checks that has to be done (see above “Important”) and, nevertheless, the experience of the technician.

Tools used for the adjustment

Feedback target

The target is placed in a nominal distance of about 3m to 3.5m from the scanner onto the wall and is used as a screen.

The marker areas on it are used to check the adjustment result.

The momentarily adjustment is qualified nearly in the center of the scan area and in the focus positions:

-300 steps +500 steps and +1300 steps

If the travel of the LASER beam does not go out of the green area during the focus movements (if the distance is in the range between 3m - 3.5m), you have shot the bull’s eye.

If the distance is shorter than 3meters (e.g. only 2.5m) the allowed movements should be closer to the black circle in the center; the acceptable (green) area has to be defined smaller.

For best adjustment results a distance of at least 3m is recommended; further details will be discussed later.

· You may also create and print your own target by the help of any image editor that is able to draw circles or print the example target.

LASER source holder with beam mirror

As light source of the LASER beam a LASER pointer or any other kind of LASER source can be used; the mechanical construction may be important for the mounting possibilities.

Precautions

Never look directly into the LASER beam of the LASER source! The emitted LASER beam has only a power of about 0.01watts but this is enough to damage the retina of the eye!

The mirror is mounted at an angle of 45º onto the LASER source holder and may be used to reflect the beam sideways, onto the wall.

The LASER source holder can be mounted so, that the beam passes the opposite side (or any side) of the LASER source mounting where no mirror is mounted.

“Arrangement 1”

Objective mirror

Into the “objective position 1” nut of the objective changer the “objective mirror” is inserted (into the 20x objective position; after the objective was dismounted).

· If the objective position is already adjusted please do not loosen the objective nut (or the objective counter nut) during removing the objective; otherwise the adjusted objective position will be lost!

This mirror defines the plane of the objective pupil; the mirror is mounted perpendicular to the optical axis and is used to arrange the entire configuration for the real adjustment.

After the configuration is arranged, this mirror is not needed for the real adjustment.

See also “Arrangement 1” and “Arrangement 2”.

Remark

· Because the plane of the objective pupil has to be checked more times during the adjustment, we recommend to remove both objectives (the 20x and the 40x objective or the dummy objective also).

· By rotating the objective disc manually into the objective position 1 or 2 respectively, you can easily and quickly “switch” between the “Arrangement 1” and the “Arrangement 2”.

Another solution

A simple mirror may also be used to define the objective plane, instead of the objective mirror; see also “Arrangement 1C”.

Dismount the objective by unscrewing only the objective from the objective nut; do not modify the adjusted objective position (the wrenches should not be used).

The reference surface of the objective nut should be smooth; stickers / labels should not disturb the smoothness of the reference surface.
Put the mirror onto the reference surface of the objective nut as shown and find a well reachable place on the wall for the screen; use the laser beam.
Stick the screen onto the wall with the laser beam in the center of the marker.
Then remove the mirror.

Important

The actual reference surface position should be checked more times during the adjustment process.

Holding system

To mount the LASER source in the correct position over the focus unit, this holding system is used; the dovetail foot is mounted onto the truss, onto the position of the turret unit fixing clamp. Onto the other end of the holding system the LASER source holder fixing with mirror is mounted.

· You can also use any holding system that allows the position adjustment of the LASER source holder with mirror in the X-, Y- and Z-direction of the room co-ordinates.

Holding systems from NOGA “http://www.noga.com/”

Mirror slide

The mirror slide is used in the “Arrangement 2”; it deflects the LASER beam according to the inclination of the specimen holder; the angle of the inclination is added to the angle of the LASER beam and so the LASER beam is deflected more, less or equal in relation to the “Arrangement 1”.

Remark

The LASER beam will be reflected on the surface of a simple slide also; so it is not important to use a mirror slide in the arrangement 2; but the description refers always to “the mirror slide”!

· Please check the shape and the surface of the no-coated slide carefully before using it during the adjustment!

Parallelogram

This component ensures the parallel shifting of the X-Y plane if the focus pin moves the slide toward or away from the objective (in Z-direction).

· The position of the component in relation to the X-Y-plane of the slide defines the perpendicularity of the optical axis in relation to the slide surface (tissue).

Parallelogram

Connection surfaces

The connection surfaces to the Y-carriage and the specimen holder are grinded to ensure the parallelism of both connected components.

Parallelogram movement limiter

The allowed movement of the parallelogram is limited to be 0.8 mm over all in P250 and S_M_D_II-type scanners (in opposite to S_M_D = 0.5mm). This limit is constant and can not be adjusted separately. To adjust the correct limiter position a fitting plate is used. The thickness of the used fitting plate defines the correct position of the entire parallelogram and so the position of the parallelogram axle in relation to the limiter also.

How to check the acting range of the parallelogram and the movement limiter?

1) Move the stage with the service program nearly into the middle of the scan area.

2) Set the focus motor to Home 1,2

3) Set the focus motor to the position -300 steps.

4) Check the position of the parallelogram axle in relation to the limiter; in this position a small gap should exist and the focus pin should be in connection to the slide.

5) Set the focus motor to the position +1300 steps.

6) Check the position of the parallelogram axle in relation to the limiter; in this position a small gap should also exist.

· If a small gap on both limit positions can be found, the focus pin is always in connection with the slide and the inclination angle adjustment of the slide is finished (the adjustment of the parallelogram position), the movement limit of the parallelogram is well defined.

· During the adjustment procedure of the parallelogram position please check this behavior more times; even if the inclination angle of the slide in X-direction was modified!

· If there is no gap in the appropriate limit position or the slide is not in connection with the slide over the entire focus range, the inclination of the slide in X-direction may be wrong or the thickness of the fitting plate may be selected incorrectly.

Preload spring

The preload spring is used to create a force of the specimen holder against the focus pin, so a mechanical preload is created. With this solution the slide is always in connection with the focus pin during focusing or specimen holder movements in the scan process.

Check the parallelogram

Before you mount the parallelogram (even if a used one should be mounted) you have to check it of functionality, visually. The parallelogram is correct if all 8 mounting spring leafs are not bent or broken. The flat springs are the most sensitive parts of the parallelogram.

· Compare the look of the springs to each other!

Fitting plate

The fitting plate is used to define the parallelogram limiter so, that the limiter does not limit the parallelogram movement in the range between -300 and +1300 focus steps; the thickness of the selected fitting plate is correct if there a small gap on the limiter exists if the focus position of -300 steps is reached and even so, there have also to be a small gap if the focus position of +1300 steps is reached.

· The fitting plate is mounted between the Y-carriage and the parallelogram.

· The drillings and edges have to be burred; the surface should be smooth.

· The nominal thickness of the fitting plate can be 0.1 mm, 0.2 mm, 0.3 mm, 0.4 mm, 0.5 mm or any combination of two fitting plates.

· The thickness of the fitting plate is often found between 0 (no fitting plate used) and 0.5 mm.

· The fitting plate has to cover the entire mounting surface of the parallelogram; see also below “Mount the parallelogram”.

· In exceptions, the fitting plate might be mounted between the specimen holder and the parallelogram (if a negative value for the fitting plate thickness have to be reached); in this cases, there have not to be a fitting plate on the mounting surface between parallelogram and Y-carriage; the adjustment bolt(s) are always used on the side, where the fitting plate is placed.

Inclination angle adjustment in X-direction

This adjustment bolt is used to define the angle of the parallelogram and so the angle of the specimen holder (slide). By screwing the X-direction preload angle bolt, the inclination angle of the parallelogram can be adjusted to zero (the angle of inclination between X-min and X-max of the scan area).

IMPORTANT: The limit of the parallelogram movement limiter is affected also and has to be checked always after driving the bolt.

“How to check the acting range of the parallelogram”

Inclination angle adjustment in Y-direction

With this adjustment the angle of inclination in Y-direction is reduced / eliminated; it means the inclination angle between Y-min and Y-max of the scan area.

· By loosening the mounting bolt and tightening the adjustment bolt or by loosening the adjustment bolt and tightening the mounting bolt, the angle of the inclination in Y-direction is modified.

· The adjustment is done only on one side; the adjustment bolt of the opposite side should be fully loosened (or removed)!

· If the adjustment on the actual side does not deliver the desired result, change the adjustment bolt to the opposite side and execute the adjustment there.

Mount the parallelogram

On this side the parallelogram is mounted together with the fitting plate (if needed).

· Place the fitting plate so, that the entire (grinded) mounting surface of the parallelogram is covered by the fitting plate.

· If the thickness of the fitting plate has to be modified, dismount the entire parallelogram with specimen holder, exchange the fitting plate and mount the parallelogram again (the two mounting bolts).

On the opposite side of the parallelogram the specimen holder is mounted, usually without fitting plate and without adjustment bolts.

In exceptions, the fitting plate is mounted between the specimen holder and the parallelogram (if a negative value for the fitting plate thickness have to be reached); in this cases, there have not to be a fitting plate on the mounting surface between parallelogram and Y-carriage; the adjustment bolt(s) are always used on the side, where the fitting plate is placed.

Principle of the adjustment

In “Arrangement 1” the position of the entire scanner, more precise, the position of the objective nut will be defined; in “Arrangement 2” the position of the specimen holder in relation to the previous arrangement will be defined. In other words, the plane defined by the objective mirror is used as reference plane and the plane of the specimen holder with the mirror slide will be adjusted to the reference plane. Therefore it is important, that the found position of the reference plane during the “Arrangement 1” must not to be altered until the entire adjustment is finished successfully!

This means:

· The position of the components must not be altered until proper alignment is reached.

Please do not move

· The table

· The entire scanner or the truss

· The holding system (it may occur if the parallelogram bolts are driven)

· The LASER source (it may occur if the parallelogram bolts are driven) and

· The screen until the entire adjustment is not finished correctly.

If one of the named components changed its position, please rotate or insert the objective mirror into the optical axis again, remove the target and stick it again onto the wall in the right beam position; or start with “Arrangement 1” again.

Arrangement 1A

Change the objective to the objective mirror.

Mount the LASER source holder with the holding system over the objective nut.

Adjust the position of the laser source holder so, that the LASER beam is reflected by the objective mirror to the LASER beam mirror in an angle that is nearly perpendicular.

The angle of the laser beam, reflected by the objective mirror should be as small as possible; so the error of the adjustment will be small also.

· To reach this, the LASER beam source holder should be mounted at a large distance from the objective nut.

By positioning the entire scanner and the holding system find a well reachable and visible part on the wall (use the LASER beam) and stick the screen onto the wall with the LASER beam in the center of the marker position.

Arrangement 1B

The only difference to the arrangement 1A is the use of the LASER beam mirror.

· In the arrangement 1B the LASER beam mirror is not used.

Known errors

· Important: The LASER beam source holder should be mounted at a large distance to the objective nut.

Arrangement 1C

· The LASER beam mirror may be used or not like shown in the arrangement 1A or 1B.

· The main difference to the arrangement 1A or 1B is the use of a “simple mirror” on the objective nut instead of the objective mirror.

This version needs more accuracy during the preparation of the adjustment 1”!

Known errors

· Important: The LASER beam source holder should be mounted at a large distance to the objective nut.

Arrangement 2A

Rotate the objective disc into the objective position 2 (there is no objective present).

If arrangement 1C is used, remove the “Simple mirror” and check the LASER beam position in relation to the “illumination hole” of the focusing part.