IGC

Leica HCS (High Content Screening) | Reservations

  • LeicaHCS
  • The new Leica HCS microscope is a fully automated microscopes specially designed to do high troughput and screening of samples by itself for a more efficient image acquisition. Equiped with the new Fast Filter Wheels for both emission and excitation and the new Fluorescence LED light source, the microscope can change channels much faster than a regular widefield and keep high levels of signal. The microscope is equipped with a Hamamatsu Flash Orca LT 4.0, a sCMOS camera with 4.0 MPixels with a big field of view and capable of maximum of 30 FPS at full frame (2048x2048).

    Location : UIC Room 2 (0B03), Bartolomeu Dias Wing

    Microscope : Leica DMI6000

    Year Installed: 2014

    Optics : Fluorescence and brightfield (with DIC)

    Camera : Hamamatsu Flash Orca 4.0 sCMOS 2048*2048 px

    Operating conditions : Temperature Controlled

    Keywords: widefield, live imaging, HCS, screening, high troughput, inverted microscope, high content screening


Suggestion for description in "Materials and Methods":

Images were acquired on a commercial Leica High Content Screening microscope, based on Leica DMI6000 equipped with a Hamamatsu Flash Orca 4.0 sCMOS camera, using a 100x 1.44 NA objective, DAPI + GFP fluorescence filtersets and controlled with the Leica LAS X software.

Suggestion for "Acknowledgments":

We would like to thank the technical support of IGC's Advanced Imaging Facility (AIF-UIC), which is supported by the national Portuguese funding ref# PPBI-POCI-01-0145-FEDER-022122, co-financed by Lisboa Regional Operational Programme (Lisboa 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER) and Fundação para a Ciência e a Tecnologia (FCT; Portugal).

Maintenance log:

  • 26-03-2015 - Lumencor LED power software control installed.
  • 13-01-2015 - LED fluorescent emission installed at the microscope to replace Metal Halide mercury lamp.

Booking Calendar details

Configuration
Value
Booking type
Hidden
Responsible
Unidade Imagiologia
Minimum booking time (in min)
30
Schedule tolerance (in days)
4
Detele tolerance (in hours)
1
Booking restriction status
Disabled

Available objectives

Place mouse over Magnification to see the objective.
Magnification
Objective type
NA1
Immersion
Working distance (mm)
Resolution2 @525nm in XY (µm)
Pixel Size (µm)
Nyquist sampling @525nm in Z (µm)
Link
5x
HC PLAN FLUOR
0.15
-
13.7
2.135
1.00
3.203
10x
PLAN FLUOR
0.30
-
16.0
1.068
0.50
1.601
20x
PLAN APO
0.7
-
0.59
0.458
0.25
0.686
40x
HCX PLAN APO
1.3
Oil
0.22
0.246
0.13
0.370
63x
HC PLAN APO
1.4-0.60
Oil
0.14
0.229
0.08
0.343
100x
HC PLAN APO
1.44
Oil
0.10
0.222
0.05
0.334

(1) NA - Numerical Aperture. For more information, follow this link.

(2) These values were calculated using the Resolution formula adapted to confocals as seen in the Olympus micro website: R = 0.4*λ/NA

Installed filtersets

Filter cube
Excitation filter
Dichroic
Emission filter
Filter cube wheel position
Filter ID
DAPI
BP 360/40
400
BP 470/40
#1
A4
EGFP
BP 480/40
505
BP 527/30
#2
L5
TRITC
BP 546/12
565
BP 600/40
#3
RHO
CY5
BP 620/60
660
BP 700/75
#4
Y5
FFW
DAPI
350/50
Quad
430/505/575/670
455/50
#5
QAS
GFP
490/20
525/36
TRITC
555/26
605/52
Cy5
645/30
705/72

IMPORTANT: Take note that the Fast Filter Wheel (FFW) uses the same cube and a Quad dichroic, which can be translated into a bleedthrough of some fluorophores into the other channels.

FFW filters cannot be used to focus the sample through the eyepieces!

Spectra, lines & filters:

Setup

Turn on procedures

  1. If you are the first person of the day, turn on the Light source power button.

    • Located on the right side. Click image to magnify.

    mercury

    mercury

  2. Turn on the LED button.

    mercury

    on/off switch

  3. Turn on the Camera. This should be done before turning on the microscope control box.

    Note: a LED ligt should start blinking. Only turn the microscope on when the LED stopped blinking and it is orange.

    mercury

  4. Turn on the microscope control box.

    on/off switch

  5. Start the Leica LAS X software available at the Desktop.

    Leica LAS X shortcut

  6. Optional: Start Sola GUI application to control LED light source and select COM7 if not selected.

    Sola GUI

  7. Set up your experiments on the microscope.

Turn off procedures

  1. Close the software and Log-off from Windows. If you are NOT the last user of the day, you don't need to do the following steps.
  2. Turn off the microscope control box.
  3. Turn off the Camera.
  4. Turn off the Lamp LED button (2).
  5. Turn off the Lamp power supply button (1).
  6. If you used any immersion lenses be sure to clean them.

Acquisition

  • Grid Colection / TileScan

    It is possible to acquire grids and tiles of big samples through Leica LAS X software. For a more detailed information, please check the following link or click in the image to the right.

  • Tile Scan

  • Adaptive Focus Control (AFC)

    AFC is Leica's version of Autofocus through hardware. This funcionality allows continuous focus of the sample through coverslip position, keeping the sample focused during the whole acquisition with the main objective of preventing drift in Z over long periods of time. For more detailed information, please visit the following link.

  • Leica AFC


Extra Info