====== Safety Considerations ======

The 2P system is custom-built but is designed to be treated as a closed-box system for laser safety purposes equivalent to a regular commercial confocal.

  * Do not attempt to use the system without basic training (from Erika, Istvan, or Greg)
  * Please review the [[http://intranet.lmb.internal/support-services/health-and-safety/radiation/lasers/|basic laser safety information]] and also the [[equipment:multiphoton:2p_risk_assessment]] with Greg
  * Only users have been trained in the [[protocols:aligning_the_2p_laser|laser alignment procedure]] and attended the university Class IV laser safety course may
    * open the system bolted enclosure covering the laser pathway on the table 
    * align the laser

====== Overview ======


The multiphoton system has become a mess of computers, wires, cables and other kit, fit to make a physiologist proud.

In principle it's quite simple to use the system once you've gotten over the slightly ridiculous learning curve. When all 
kit is working properly you'll have the Mac running MicroManager and Igor, and a PC running MATLAB/ScanImage. Basic use of
the multi-photon system can essentially be done through the PC and ScanImage.

If you're not familiar with 2P microscopy these tutorial videos will be useful:
  * [[https://www.youtube.com/watch?v=JSnh-btk22U]] - building a 2P scope in 8 mins
  * [[https://www.youtube.com/watch?v=CZifB2aQDDM]] - theory and tips in 2P imaging
  * [[https://www.youtube.com/watch?v=DLTaqzz8hJM&t=200s]] - beam paths in our Sutter MOM scopes
  * [[https://www.youtube.com/watch?v=hwCFtQ3WHoo]] - alignment tips for Sutter MOM
  * [[https://www.youtube.com/watch?v=hLmTZQFQU9o]] - Sutter MOM in detail

For advanced use and making changes in hardware first thing to do is read the Scanimage Concepts pages: [[http://scanimage.vidriotechnologies.com/display/SI2019/Concepts+Guide]]

====== Starting ScanImage ======
JefferisLab user, start Matlab from the desktop and run scanimage command.
ScanImage opens a dialog box requesting a Machine Data File and User Configuration. It defaults to the folder containing the most recently run
MDF/configuration, which is probably the correct one for the system (for example, MachineDataFile.m will most likely be the first thing
you'll see if you run ScanImage after I've been on the system. Select your config of choice. The MDF contains information about hardware configuration (which DAQ board channels correspond to what hardware, e.g. Scanners, PMTs, Pockels, Piezo, Stage motor), the user config defines the layout and presence of the windows of ScanImage while running.
ScanImage now opens a new dialog asking for the System Configuration file((and defaults to the Standard if you hit cancel, this is
bad, so don't hit cancel)), and again, it will default to the last opened file which is probably the correct one.

All of your ScanImage windows will now open and move themselves to the positions that are specified the User Config files.

====== Using ScanImage ======
To be quite honest, you should go read [[http://scanimage.vidriotechnologies.com/display/SI2019/Window+Reference+Guide|this]] documentation right now. It will save you a lot of time if you understand what all of these things mean. 
At the moment we are using ScanImage 5.6 from 2019, we might update to the Premium version to allow simultaneous imaging and optogenetic stimulation with a second laser.
==== Main Controls Window ====
This window has most of the useful controls and is the prime window for changing user settings and configurations. Much of what 
is useful in this window is fairly self-explanatory so I won't dwell on it for now.

==== Configuration Window ====
This window is essentially the guts of your configuration file. Here you control
  *Pixels/Per line
  *Lines/Frame
  *Uni/Bi-directional scanning
  *Ms/Line
  *Acquisition delays (important for calibrating bidirectional scanning to remove chatter)
  *Flyback settings of Y mirror when using bidirectional scanning 

One interesting note from the ScanImage documentation:\\
**Scan Amplitude X/Y :**
<code>
Adjust Infrequently
In general, the Scan Amplitude X/Y values are not intended to be varied frequently – they are typically set to the maximum 
half-amplitude the physical scanner supports in all CFG files maintained by a user. Adjusting the scan range is usually 
accomplished by the Zoom value instead. However, using the Scan Amplitude X/Y values to adjust the scan range allows scans
of: 1) areas which would require a fractional Zoom (e.g. 1.5x) and 2) non-square aspect-ratio areas.
</code>