Zeiss LSM 710 User Manual
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Zeiss LSM 710 User Manual

Confocal microscope
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See also: Quick Manual
ZEISS LSM 710 CONFOCAL MICROSCOPE
USER MANUAL
START THE SYSTEM ............................................................................................... 2
START ZEN SOFTWARE .......................................................................................... 3
CONTROLLERS .............................................. 4
2
OBSERVATION AT OCULARS .................................................................................. 5
STATIF PRESENTATION .......................................................................................... 6
ACQUIRE ONE OR SEVERAL STAININGS .................................................................. 7
SAMPLING (IMAGE RESOLUTION) ........................................................................ 13
ACQUIRE A Z STACK ............................................................................................ 14
ACQUIRE A TIME SERIES ...................................................................................... 16
ACQUIRE A MOSAIC IMAGE ................................................................................. 17
ACQUIRE MULTIPLE STAGE POSITIONS ................................................................. 19
VISUALIZATION OF YOUR ACQUIRED IMAGES ....................................................... 20
SIGNAL QUANTIFICATION ................................................................................... 21
SAVE YOUR IMAGES ........................................................................................... 22
SWITCH OFF THE SYSTEM .................................................................................... 23
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Summary of Contents for Zeiss LSM 710

  • Page 1: Table Of Contents

    ZEISS LSM 710 CONFOCAL MICROSCOPE USER MANUAL START THE SYSTEM ....................2 START ZEN SOFTWARE ..................3 SET THE TEMPERATURE AND THE CO CONTROLLERS ..........4 OBSERVATION AT OCULARS .................. 5 STATIF PRESENTATION ..................6 ACQUIRE ONE OR SEVERAL STAININGS ..............7 SAMPLING (IMAGE RESOLUTION) ................

  • Page 2: Start The System

    START THE SYSTEM 1. Switch on « MAIN SWITCH » button. 2. Switch on « SYSTEM PC » button. 3. Switch on the PC and log in « USER » session. 4. Once Windows session starts, switch on « COMPONENT » button. 5.

  • Page 3: Start Zen Software

    START ZEN SOFTWARE 1. Start the software by clicking on ZEN 2011 icon on the desktop. 2. Choose START SYSTEM to access to the acquisition menu. 3. Zen software is divided in 3 parts: the left side is dedicated to acquisition parameters, central part is for displaying images and toolbar and the right side list acquired images.

  • Page 4: Set The Temperature And The Co Controllers

    SET THE TEMPERATURE AND THE CO CONTROLLERS 1. Open the INCUBATOR window. 2. Enter the temperature value that you need and tick the box. 3. Open the CO2 bottle. 4. Enter the percentage of CO and tick the box. Warning !!! At the end of your session, please untick all boxes and close the CO bottle.

  • Page 5: Observation At Oculars

    OBSERVATION AT OCULARS 1. Choose the 1 tab : LOCATE. 2. Choose the adapted configuration for fluorescence or transmission. According to the type of illumination that you use, you can : 3. Light on/light off/choose the intensity of the transmission lamp. 4.

  • Page 6: Statif Presentation

    STATIF PRESENTATION On the left side : 1. Focus knob 2. No fonction 3. No function 4. Optovar 1x 5. Previous fluorescence filter 6. Next fluorescence filter On the right side : 1. Focus knob 1. Set the objective at the lowest position to install your sample.

  • Page 7: Acquire One Or Several Stainings

    ACQUIRE ONE OR SEVERAL STAININGS 1. Choose the ACQUISITION tab. 2. Tick SHOW ALL TOOLS. 3. In the LASER toolbox activate the 561 laser if you need it (Power On). 4. Open SMART SETUP. 5. In DYE select the different dyes and their respective color.
  • Page 8 6. Several methods are available : -FASTEST : allows a simultaneous acquisition ; it is the fastest method but the cross talk from one color to the next channel could be important. -BEST SIGNAL : each channel is acquired separately. It is the slowest method but it efficiently avoids the Cross talk phenomenon.
  • Page 9 7. In the LIGHT PATH toolbox, each TRACK represents a sequence of one or several colors. 8. Add or remove sequences by clicking on +/- 9. Adapt wavelength detection by moving the window for each channel. 10. Add or remove channels with +/- for each sequence.
  • Page 10 14. In CHANNELS toolbox, adjust the pinhole at 1 AU (Airy unit) for each channel in order to get the best resolution to signal ratio. 15. Adjust the GAIN (MASTER) at 550 for each channel except for the T-PMT. 16. Acquire in LIVE. 17.
  • Page 11 22. In ACQUISITION MODE toolbox choose the resolution (be careful of the sampling) see p12. 23. Move in Z to find the plane of interest. 24. Choose the scan speed (faster it is, worse would be the signal to noise ratio). 25.
  • Page 12 31. You can zoom directly on your region of interest : stop the LIVE then click on CROP under the image. A square appears on the image. The blue line represents the top of the final image. You can move, rotate, enlarge or reduce the square. Then click on LIVE or SNAP. 32.

  • Page 13: Sampling (Image Resolution)

    SAMPLING (Image resolution) In order to get an optimal resolution for your images, the image voxel size must be equal to the half of the objective resolution (Nyquist criterion). On a light microscope, the lateral resolution (in XY) is better than the axial resolution (in Z).

  • Page 14: Acquire A Z Stack

    ACQUIRE A Z STACK 1. Proceed in the same manner as in the ACQUIRE ONE OR SEVERAL STAININGS section and adjust the parameters in the brightest plane. 2. Tick Z-STACK to make the toolbox appears in the MULTIDIMENTIONAL ACQUISITION menu. Two methods : If you know about your sample thickess 3.
  • Page 15 10. For colocalisation acquisition, click on MATCH PINHOLE in order to adjust automatically pinholes size to get the same voxels size in all channels. 11. To adjust the acquisition sequence, in LIGHT PATH go to SWITCH TRACK EVERY and choose Z-STACK to change the channel after the acquisition of the entire stack.

  • Page 16: Acquire A Time Series

    ACQUIRE A TIME SERIES 1. Proceed in the same manner as in the ACQUIRE ONE OR SEVERAL STAININGS section and adjust the parameters in the brightest plane. 2. Tick TIME SERIES to make the toolbox appears in the MULTIDIMENTIONAL ACQUISITION menu.

  • Page 17: Acquire A Mosaic Image

    ACQUIRE A MOSAIC IMAGE 1. Proceed in the same manner as in the ACQUIRE ONE OR SEVERAL STAININGS section and adjust the parameters in the brightest plane. 2. Tick TILE SCAN to make the toolbox appears in the MULTIDIMENTIONAL ACQUISITION menu.
  • Page 18 10. To make a mosaic including different fields of interest, go to BOUNDING GRID option. 11. Find fields of interest in LIVE and save their coordinates by clicking on ADD. To remove a position, select the coordinates on the list and click on REMOVE.

  • Page 19: Acquire Multiple Stage Positions

    ACQUIRE MULTIPLE STAGE POSITIONS 1. Proceed in the same manner as in the ACQUIRE ONE OR SEVERAL STAININGS section and adjust the parameters in the brightest plane. 2. Tick POSITIONS to make the toolbox appears in the MULTIDIMENTIONAL ACQUISITION menu. 3.

  • Page 20: Visualization Of Your Acquired Images

    VISUALIZATION OF YOUR ACQUIRED IMAGES Under the image you can find a panel where you can select channels you want to see on the screen. 1. Tick the name (white on blue) of channels you want to see in the image.

  • Page 21: Signal Quantification

    SIGNAL QUANTIFICATION To quantify a fluorescence signal, the different experimental conditions must be acquired with the same acquisition conditions (same laser power, same gain and offset for PMTs). If possible, acquisition conditions must be adjusted on the brightest sample. In this way, the other samples won't be saturated.

  • Page 22: Save Your Images

    SAVE YOUR IMAGES Select one image then save it in .lsm FILE – SAVE in USER/année/mois/jour/nom d’utilisateur folders. You can select all acquired images with a right clic on the set of images then SELECT ALL and SAVE. You can also delete them by selecting DELETE.

  • Page 23: Switch Off The System

    SWITCH OFF THE SYSTEM Warning !!! If you used the temperature and CO controllers, please untick all boxes and close the CO bottle. Check the booking schedule online to know if the system is booked after your session. If the system is booked after your session or 4h later, don't switch off completely, but proceed like this : 1.

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