Primary Use. Operating Principle

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1 Primary Use The Leica DVM6 is an optical microscope that has the ability observe samples at a high magnification at a high resolution. The microscope allows users to view their sample with up to a 2350x magnification while not losing out on resolution or quality. This instrument is capable of producing images up to 1 micron in accuracy. The accuracy and resolution make the microscope a useful product in many MSE projects. The microscope lets you manually adjust the tilt of the lense system as well as the rotation and movement of the sample stage. This allows the user to produce any angle of picture with ease. There are currently two objectives at varying working distance and magnification allowing the user to interchange the lense system to achieve different images. There are also two different stages: one solid fill and one with a controllable backlight. The system uses a computer software that allows for seamless interface with the microscope and easy manipulation of the image. The software comes equipped with a variety of tools that allows the user to find the perfect image of his/her sample. Operating Principle The Leica microscope uses a converging lense system to produce a magnified image. Using a digital camera, the sample being observed is viewed and analyzed on an attached computer/monitor. Light passes through a series of lenses which either spread out light rays making the sample appear larger or further magnify the image. The sample will be in focus when it is at a sufficient distance away from the objective lense, and can be altered by raising and lowering the stage. This microscope relies heavily on image processing. The capabilities expand beyond 2D analysis and can generate a 3D surface map of a sample. The software can automatically take images of the sample through a predetermined range of distance. Safety Procedures The Leica Microscope has a very low possibility of injuring the user, however there are several safety precautions to keep in mind so the device does not break. When coming close to the lense system or handling the objectives, gloves are needed to protect the delicate systems from oil and dust. This is also important when handling samples on and around the stage. Always be aware of the distance between the lense system and the stage in order to prevent collisions when focusing or tilting the system. When tilting the system, never forcefully tilt the system without pressing the black handle and the silver handle together on the back of the system. The tilting of the system should be done with care, place one hand on the handles and the other hand on the white part of the head so the system doesn't fall of break against the stage. Be aware the device produces beeps when it detects the head getting too close to the stage.

2 General Procedure 1. Steps for startup: a. Remove red cover over the microscope b. Turn on the microscope with the switch on the back and wait for the indicator light on the left side to turn green Figure 1a-b. The power switch and green light referred to in step 1b. c. Once the light is green, open the Leica LAS X Hardware Configurator Figure 2: Shortcut to Leica Hardware Configurator. d. Click the Hardware Setup button in the top left e. Make sure the Port and the Stage both match in saying DVM6 f. Click Test g. If it says it was successfully completed you can click Apply then OK and you can close out of the Hardware Configurator software i. If it says there was an error you might have a problem with the connection between the microscope and the computer ii. If you open the LAS X and an image is not being generated, then restart either the computer or the microscope until the connection is achieved

3 h. Next open the LAS X software and follow the steps in the popup menus to continue to the software 2. Adjusting the Magnification a. To control the magnification of the microscope, you can twist the big red wheel above the objective lense system b. Counterclockwise decreases the magnification, clockwise increases it c. You can also control the digital magnification with the scroll wheel in the software, however the digital magnification will not increase the resolution of the image Figure 3: The red magnification adjustment wheel highlighted in blue. 3. Focusing the Microscope a. After finding the magnification of your choice, you can focus the lenses by twisting the adjustment wheels towards the bottom of the microscope below the stage b. The bigger wheel is for rough focusing while the smaller wheel is for fine adjustments c. Secondary, if you left click and drag on the live image, a menu will pop up. Simply click the auto-focus and the highlighted area you have selected with adjust to focus

4 Figure 4: Focusing wheels on the microscope. 4. Movement of the Stage a. You can move the stage itself to move the location of your sample in either the X or the Y direction b. Also you can adjust the rotation of the stage by twisting it using the black knob on the bottom of the left side of the stage c. You can monitor the angle of rotation on the popup on the software 5. Movement of the Lense System a. To adjust the tilt of the lense system by slightly pressing the black and silver handle sticking up towards the back of the microscope, behind the lense system Note: Never try tilting the system without slightly pressing the two handles together b. You can monitor the angle of the tilt on the popup on the software Figure 5: Gripping Procedure for Tilting the Microscope.

5 6. Opening a Previous Image a. The Explorer Menu lets the user navigate through computer files and open previous images b. Open the Explorer Menu in the top left on the screen c. Open the Folder or location where the image is located and open the Multifocus Image to use the full capabilities of the 3D image Figure 6: The explorer menu (top-left) for step Finding the Correct Exposure and Settings a. In the Image Menu, the top left button turns on the auto exposure. This is used in most situation to create a good image quick and easy b. To go in further detail or create a perfect image, the exposure, the brightness, and the contrast can be manually adjusts Figure 7: The image menu for step 7.

6 8. Controlling the illumination of the sample a. To get a easy image of a basic sample, the default ring light should be used. The intensity can be controlled as well as which lights are illuminated by picking the correct preset under the stage drop down b. To get a better overhead light, the Coaxial light should be used. This works well on metal samples where grain boundaries need to be analyzed c. The Backlight can be put on the stage to give the sample better light from the bottom. This works well with thin films and transparent samples Figure 8: The illumination menu for step Creating a multifocus image using the Start Up method a. Click on the Start Up button b. Switch on the Calculate an Extended Depth of Field switch c. Set the maximum number of steps to 60. If necessary, the user can raise the number of steps to 100 d. Close the menu and get the lowest point of the image into focus e. Click the Z-Stack button on the bottom toolbar of the software f. Monitor the process and stop the process once the highest point of the image just goes out of focus to insure the full image has been captured

7 Figure 9: The start-up multifocus menu for step Creating a multifocus image using the Custom method a. Click on the Custom button b. If there is a previous preset in the menu, click the trash button in the menu c. To start, get the lowest point of the image in focus and click begin d. Then get the highest point of the image in focus and click end e. Close out of the menu and click the Z-Stack button on the bottom toolbar of the software to run the program 11. Using a multifocus image to create a 3D model a. After an image has been Z-Stacked, click the 3D model button on the right side toolbar to open the 3D model b. A creat map can be created with different intensities as well as adding axis scallings, scale bars, and a heat map legend c. Different overlays and measurements can be added in the measurement tab Different step heights, volumes, and profiles can be created as overlays d. In order to return to the 2D view, click the 2D button on the bottom toolbar

8 Figure 10a-b: The button used to access 3d view (left) and the 3d workspace (right). 12. Stitching together multiple images together to create a photo of the entire sample a. Click the Stage Menu button to begin. b. Click the Spiral button c. After closing the menu, click the Stage icon button on the bottom toolbar d. This will begin the process and it will finish automatically e. Now a high resolution image has been created of the whole sample Figure 11. The stage menu button for step Saving an image a. To save an image, click the capture button on the bottom toolbar b. In order to save the image with overlays, select the stamp button on the top of the right toolbar. If the capture button has a stamp on it, then the overlays will be saved

9 Figure 12. Location of the save image button for step Create overlays on an image a. The top right of the toolbar has a menu to add and control overlays b. To measure distance, click the distance to distance button and click on two points that are being measured c. A scaler bar can be added through the scale bar menu and can be customized d. Annotations can be added that can have text added to point out certain features of the image e. The overlay menu can control which overlays are shown and if they will be saved or not Figure 13. The overlay menu for step Changing the objective a. Put on nitrile gloves b. Loosen locking screw on the left side of the microscope head c. Grip the objective with one hand and slowly pull directly outward until fully clear of the microscope head d. Without setting the objective down, place it in its designated box (located in clear containers beneath the Mark-10 e. Carefully lift the new objective out of its box and align it to the tracks on the microscope head

10 f. Slowly push the objective in all the way and tighten the locking screw Figure 14: The locking screw highlighted in blue.