Leica SP8 Scanning Confocal

Installed in January of 2016, the SP8 is a versatile scanning confocal with all the bells and whistles. This system has 4 sensitive HyD detectors and 2 PMT detectors enabling the simultaneous imaging of up to 6 different wavelengths. It has a White Light Laser (WLL) that can provide up to 8 excitation lines between 470 - 670nm) in addition to 405nm and 440nm lasers to provide almost any excitation wavelength and a prism spectrometer to control emission wavelengths to make this a very versatile system compatible with most combinations of fluorophores.

Startup SP8 without FLIM

  • Turn on key and 3 green switches (under table by your knee No. 1,3,2).
  • Turn on power strip on back, right side on microscope table (No.5).
  • Turn on lamp if needed (No. 6 on top self of rack to left of scope).
  • Sign-in to iLab. Use kiosk in LMIC or your own phone or computer.
  • Log in Windows. TCS user/password on yellow tape (IU040790)
  • Sign in to LMIC software to transfer images (IU username).Check to see what objectives are installed. Start Leica LAS X Hardware Config software if you need to change objective.
  • Start LAS X software.

Shutdown

  • Make sure all images are saved.Exit LAS X software.
  • Transfer images (Wall-E robot icon), Select Data Directory, add title, sign out. Wait until finished
  • Shut down computer. Wait until screen goes dark.
  • Turn everything off in reverse order from start up (Lamp, power strip, green switches + key).
  • Use lens paper and cleaning solution to clean objective.Cover scope.
  • Sign out of iLab - or you will keep getting charged.

Configuration

  • Leica DMi8 inverted microscope platform
  • Lasers: WLL (470-670nm), 405nm, 440nm.
  • Can simultaneously image fluorescence and brightfield or DIC
  • FRAP or FRET fully supported with rapid kinetic analysis
  • Picoquant system for FLIM or FCS
  • Controlled by Leica LAS-X software
  • Objectives:
    SystemCorrectionMagnificationImmersionN.A.W.D.CollarItem No.
    SP8HC PL Apo10xAIR0.4N/ANo506285
    SP8HC FLUOTAR25xWATER0.952.4No506285
    SP8HC PL Apo40xOIL1.30.24No506358
    SP8HC PL Apo63xOIL1.40.14No506350
    SP8HC PL Apo40xWATER1.10.65MOTORIZED 0,17-0,19506357
    SP8HC PL Apo20xAIR0.750.62No506517
    SP8HC PL Apo63xWATER1.20.3MOTORIZED506361
    SP8HCX PI Apo63xWATER1.2N/AManual 0.14-0.19506279

Technical Notes

  • Always use #1.5 coverslips!
  • A single coverslip in the middle of a slide works the best. This is an inverted scope. The coverslip will face down when imaging. You will need some space on either end of the slide to fit into the slide holder.

Leica documents: 

Leica SP8 confocal user-manual

Free Leica LAS-X is a free but limited version of the imaging software that will work for viewing, exporting and some processing of your .lif files. Works on PC only.

Opening .lif files in FIJI is one of the option to see and analyse your data.