View source for Olympus LEXT OLS4000 Confocal uScope - Quick Start

<sup>Demis D. John –– 2018-07-19</sup>

== '''Do not use the LEXT unless trained first!''' ==
'''You MUST be officially trained on this tool before you are allowed to use it!'''  

'''These instructions are a quick-reference for users who have already received the full training.''' 

== Quick-Start General Procedure ==
In order to acquire a profile height measurement, you must do the following:

1)  Get the sample surface in focus with the regular “color” optical microscope & lock the focus knob

2)  Switch to “laser” mode & readjust the focus

3)  Specify the bottom and top range for the focus motor to scan over & acquire the scan data (in Fine/Manual mode)

4)  In the “Measure” tab, Specify a region on the image of which to take a profile

5)  Move the cursors to the desired step-height measurement positions

== Quick-Start Detailed Procedure ==

=== Initial Setup ===
Make sure microscope is raised and stage is away from the objectives in the “Sample Exchange” position.  Make sure software is started (should show a Login screen). If this is not the case, do the following:
# Open the software “'''''OLS4000'''''”. 
# Ensure no sample is inserted, and microscope objectives are lifted away from the stage – use the coarse focus knob to lift the objectives to a safe height & Lock it, if needed. Then press “OK” on the warning & wait for motor initialization.
# At the login screen, click “OK” to login as ''Guest''.
## [[File:Olympus LEXT - login guest.png|frameless|200x200px]]
## System should automatically display the “Acquisition” tab

=== Load & focus on sample ===
# Place your sample near the center of the stage.
# Make sure '''5x''' objective is selected and objectives will not hit stage if motors move down.  If unsure, click “''Cancel''” on the “''Move to Z reference position''” dialogue, and lift the objectives using the manual focus knob.
# Click the “Scan Position” button [[File:Olympus LEXT - Button Flag measurement position.png]] to move the stage to center 
# System should already be imaging live in Color mode.  If not, click the “Color” imaging button [[File:Olympus LEXT - Color imaging button.png|frameless|50x50px]], and ensure the “Live” window-tab has a halo around it by clicking on it, like so: [[File:Olympus LEXT - Live halo.png|frameless|95x95px]]
# Focus on the sample surface by using the black manual focus knob first, unlocking/re-locking if needed. This prevents the electronic focus motor from hitting it’s limit during imaging.
## Make sure to engage the focus “Lock” ring (finger-tight only), otherwise the focus will drift.
# Fine-Focus (electronic) by using the Mouse-Wheel (Right-click the live image to set wheel to Coarse focus), or by using the “Focus” up/down arrows on the right of the image.
# Use the joystick to drive to feature of interest.  
## '''Make sure sample's TOP surface is in focus before Stepping up in objectives/magnification, to prevent crashing.'''
## The higher the magnification, the better the height resolution.
# Click the Laser imaging mode button [[File:Olympus LEXT - laser imaging button.png|frameless|50x50px]] 
# Readjust electronic focus until the sample surface lights up.
## May need to adjust laser Brightness (vertical tab on image pane), for max brightness but not saturating  can use “Saturation Highlighting” to show this <code><<pic of saturation highlighting>></code>

=== Acquisition Settings / Scan Range ===
# Set the Acquisition Settings to [''Fine''▽] & [√] ''Manual, like so:''
## [[File:Olympus LEXT - acquisition settings - Fine, manual annot.png|200x200px]]
## [√] Color mode will scan the in color mode and then in laser mode, and superimpose the true-color image onto the 3D image.
# Expand the [+] on the right of the image and enable ''Saturation Highlighting'' 
## [[File:Olympus LEXT - Saturation Highlighting button highlighted.png|frameless|201x201px]]
# Focus ⇓on the lowest surface of your sample. Go a few mouse-wheel clicks below that focus point until lowest surface has reduced in brightness and click the [Bottom] button to save this Z height as the lower point of the scan range.
# Focus ⇑on the highest surface, and go a few clicks above that until brightness decreases, and click the [Top] scan range button.
## If you see any regions of the sample surface reach saturation anywhere in the scan range, lower the image brightness using the Brightness arrows on the right of the imaging panel.  <code><<pic of brightness buttons>></code>. 
## Higher brightness reduces noise, but saturation reduces accuracy of surface height detection.
## [[File:Olympus LEXT - Top-Bottom Setting.png|alt=Olympus LEXT - Top-Bottom Setting buttons|none|thumb|200x200px|Top & Bottom focus range buttons for Manual mode]]
# Click [Acquisition] to perform the scan [[File:Olympus LEXT - Acquisition Button.png|frameless|150x150px]]
## 200 height steps takes about ~30sec.  Watch out for excessively high scan steps that could take a long time.

=== Measure Height Profile ===
# Switch to the [Measure] tab at the top [[File:Olympus LEXT - Imaging + Measurement Buttons.png|frameless|222x222px]]
# Use Profile > 2-point mode[[File:Olympus LEXT - Profile 2-point Buttons.png|alt=Olympus LEXT - Measurement: Profile 2-point Buttons|none|thumb|254x254px]]
# Click two points on your image to extract profile
# Optional: level the surface (done automatically, but sometimes can be better)
## [Image Correction] > (•) 3-Point > ''Click 3 points on image on same surface''> [Execute]  [[File:Olympus LEXT - Image Correction button.png|alt=Olympus LEXT - Image Correction button|none|thumb|205x205px|[Image Corrections] button]]
## [[File:Olympus LEXT - 3-point Leveling Screen highlighted.png|alt=Olympus LEXT - 3-point Leveling Screen highlighted|none|thumb|284x284px|3-point Leveling Screen with relevant options highlighted]]
# Place cursors on height profile & observe “Height” value in corner.
## Optional: Click ▽under Profile buttons to average around the cursors
### [[File:Olympus LEXT - Cursor Averaging.png|alt=Olympus LEXT - Cursor Averaging Panel|none|thumb|247x247px|Cursor Averaging Panel]]
## Optional: Save [[File:Olympus LEXT - Save File button.png|frameless|45x45px]] as Screenshot or LEXT file (for later analysis)

=== Shutdown at End of Session ===
# [[File:Olympus LEXT - Imaging Button.png|frameless|99x99px]] [Imaging] at top > [[File:Olympus LEXT - Color imaging button.png|frameless|50x50px]] Color <<color icon>> > [[File:Olympus LEXT - 5x objective button.png|frameless|36x36px]] 5x > [[File:Olympus LEXT - sample load position button.png|frameless]]Sample Exchange
# Log Off (NOT the [X] button!) [[File:Olympus LEXT - log out-close buttons highlighted.png|frameless|88x88px]]
## This saves each user a few minutes of boot-up time
## “''No to All''” to discard unsaved scans.