Bioimaging Wiki

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<a href="/facility/bioimaging/lib/exe/fetch.php?media=zeiss_cell_observer_wiki_2022.png"><img src="/facility/bioimaging/lib/exe/fetch.php?media=zeiss_cell_observer_wiki_2022.png" width=300></a>
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<b>Location</b>: <a href="/facility/bioimaging/lib/exe/fetch.php?media=zeiss_cell_observer_map_new_p2a24.png">Room P2-A-24</a> (<img src="/facility/bioimaging/lib/exe/fetch.php?media=phone_neg.png" width=15> 47219) <br> 
<b>Manufacturer</b>: <a href="http://www.zeiss.de/micro" target="_blank">ZEISS Microscopy</a><br>
<b>Model</b>: Axio Observer<br>
<b>Nickname</b>: "Cell Observer"<br>
<b>Software</b>: ZEN 3.0 (blue edition)<br>
<b>Year</b>: 2015 <br>
<b>SN</b>: 3851002396<br>
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Data will be deleted after: <b>15 days</b>
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&rarr; <a href="/facility/bioimaging/doku.php?id=zeiss_cell_observer_quality"><img src="/facility/bioimaging/lib/exe/fetch.php?media=list-icon.png" width=18> Zeiss Cell Observer Quality Control</a>
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&rarr; <a href="/facility/bioimaging/doku.php?id=zeiss_cell_observer_usage"><img src="/facility/bioimaging/lib/exe/fetch.php?media=chart_line.png">  Zeiss Cell Observer Usage Statistics</a>
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===== Microscope overview =====
{{  :widefield_lightpath.png?0x300}}The Zeiss Cell Observer is a fully motorized inverted //widefield fluorescence microscope// ideal for live-cell imaging with fast acquisition speeds. It is equipped with a large cage incubator and a stage top incubator for temperature control and CO2 supply and with Definite Focus for hardware focus control during long time-lapse acquisitions. Using the ZEN software you can control the motorized filters, shutters and stage to simultaneously set up multi-color time-lapse imaging experiments in multiple stage positions. The stage is furthermore equipped with a piezo for fast Z displacement. You can choose between a HXP metal halide lamp or fast-switching LEDs for illumination. Its cooled [[https://asset-downloads.zeiss.com/catalogs/download/mic/a8e5b4d7-25f6-4069-8f03-c6713d044c87/EN_product-info_Axiocam-506_mono.pdf|Axiocam 506m]] camera allows you to capture weak fluorescent signals and minimize photobleaching/photodamage in light sensitive samples. It is also equipped with a high sensitivity [[https://www.photometrics.com/products/prime-family/primebsiexpress|Photometrics Prime BSI Express]] sCMOS camera for high resolution imaging with maximized signal detection.

  * **Microscope**: Zeiss Axio Observer
  * **LED Illumination**: [[https://www.zeiss.com/microscopy/en/products/accessories/colibri.html|Zeiss Colibri 7]]
  * **Camera** (left port): [[https://asset-downloads.zeiss.com/catalogs/download/mic/a8e5b4d7-25f6-4069-8f03-c6713d044c87/EN_product-info_Axiocam-506_mono.pdf|Axiocam 506 mono CCD]]
  * **Camera** (right port): [[https://www.photometrics.com/products/prime-family/primebsiexpress|Photometrics Prime BSI Express sCMOS]]

{{::phenocycler.png?260  |}}This system is also set up to work together with the [[https://www.youtube.com/watch?v=OQGJX45Wsgw|AKOYA PhenoCycler]] fluidics instrument for automated iterative imaging cycles of whole slide tissues for [[https://www.akoyabio.com/why-spatial-biology/|Spatial Biology]] applications, such as image-based quantification of biomarkers to predict immunotherapy response, or quantification of cells by lineage and variant with single-cell resolution in the context of an intact tissue sample.

  * [[https://www.youtube.com/watch?v=OQGJX45Wsgw|Understanding the PhenoCycler Technology]] 
  * [[https://www.akoyabio.com/webinar/a-beginners-guide-to-codex/|A Beginner’s Guide to CODEX]]
  * [[https://www.akoyabio.com/webinar/spatial-phenotyping-a-revolutionary-approach-to-biomarker-discovery-for-cancer-immunotherapy/|Spatial Phenotyping: A Revolutionary Approach to Biomarker Discovery for Cancer Immunotherapy]]

{{:warning.png?25|}} Data files older than **15 days** will be automatically deleted on this system, please copy your data to the iMM server using the desktop link.

===== Booking Rules =====
  - **Users** can book at maximum **8 hours per week**
  - This usage restriction does not apply for **weekends** and for working days **before 9:00** and **after 21:00**
  - Exceptions to these rules require **approval from [[joserino@medicina.ulisboa.pt|José Rino]]**.

===== System components =====

==== LEDs  ====
^  Line  ^  Wavelength  ^
|  UV  |  385 nm  |
|  B  |  475 nm  |
|  G  |  555 nm  |
|  Y  |  590 nm  |
|  R  |  630 nm  |

==== Filtersets ====

^  Position  ^  Filterset  ^  Reference  ^  Excitation  ^  Dichroic  ^  Emission  ^
|  1  |  HE GFP  |  [[https://www.micro-shop.zeiss.com/?s=23108437692794b&l=en&p=us&f=f&a=v&b=f&id=489038-9901-000&o=|FS38HE]]  |  450-490 nm  |  495 nm  |  500-550 nm  |
|  2  |  HE DsRed  |  [[https://www.micro-shop.zeiss.com/?s=23108437692794b&l=en&p=us&f=f&a=v&b=f&id=489043-9901-000&o=|FS43HE]]  |  538-562 nm  |  570 nm  |  570-640 nm  |
|  3  |  Cy5  |  [[https://www.micro-shop.zeiss.com/?s=23108437692794b&l=en&p=us&f=f&a=v&b=f&id=488050-9901-000&o=|FS50]]  |  625-655 nm  |  660 nm  |  665-715 nm  |
|  4  |  Analyzer DIC  |||||
|  5  |  QUAD  |  [[https://www.micro-shop.zeiss.com/en/us/shop/filterAssistant/filtersets/489090-9100-000|FS90HE]] |  370-400 nm  |  405 nm  |  410-440 nm  |
|  :::  |  :::  |  :::  |  450-488 nm  |  493 nm  |  499-529 nm  |
|  :::  |  :::  |  :::  |  540-570 nm  |  575 nm  |  580-605 nm  |
|  :::  |  :::  |  :::  |  615-648 nm  |  653 nm  |  659-759 nm  |
|  6  |  DAPI  |  [[https://www.micro-shop.zeiss.com/?s=23108437692794b&l=en&p=us&f=f&a=v&b=f&id=488049-9901-000&o=|FS49]]  |  335-383 nm  |  395 nm  |  420-470 nm  |

==== Objectives ====
^  Magnification  ^  Model  ^  Immersion  ^  NA  ^  WD (mm)  ^  Reference  ^
|  10x  |  EC Plan-Neofluar Ph1  |  Air  |  0.30  |  5.20  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420341-9911-000/Objective-EC-Plan-Neofluar-10x-0.30-Ph1-M27|420341-9911-000]]  |
|  20x  |  Plan-Apochromat  |  Air  |  0.80  |  0.55  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420650-9901-000/Objective-Plan-Apochromat-20x-0.8-M27|420650-9901-000]]  |
|  20x  |  EC Plan-Neofluar Ph 2  |  Air  |  0.50  |  2.00  |  [[https://www.micro shop.zeiss.com/en/de/shop/objectives/440341-9904-000/Objective-EC-Plan-Neofluar-20x-0.5-Ph2|440341-9904-000]]  |
|  40x  |  EC Plan-NeoFluar  |  Air  |  0.75  |  0.71  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420360-9900-000/Objective-EC-Plan-Neofluar-40x-0.75-M27|420360-9900-000]]  |
|  63x  |  Plan-Apochromat DIC  |  Oil  |  1.40  |  0.19  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420782-9900-000/Objective-Plan-Apochromat-63x-1.4-Oil-DIC-M27|420782-9900-000]]  |
|  63x  |  Plan-Apochromat Ph3  |  Oil  |  1.40  |  0.19  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420781-9910-000/Objective-Plan-Apochromat-63-1.4-Oil-Ph3-M27|420781-9910-000]]  |

**Upon request:**
^  Magnification  ^  Model  ^  Immersion  ^  NA  ^  WD (mm)  ^  Reference  ^
|  1.25x  |  EC Epiplan-Neofluar  |  Air  |  0.035  |  4.00  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/422310-9900-000/Objective-EC-Epiplan-Neofluar-1.25x-0.03-M27|422310-9900-000]]  |
|  5x  |  EC Plan-NeoFluar  |  Air  |  0.16  |  18.50  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420330-9901-000/Objective-EC-Plan-Neofluar-5x-0.16-M27|420330-9901-000]]  |
|  10x  |  EC Plan-Neofluar  |  Air  |  0.30  |  5.20  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420340-9901-000/Objective-EC-Plan-Neofluar-10x-0.30-M27|420340-9901-000]]  |
|  25x  |  LCI Plan-Neofluar Corr DIC  |  Oil/Glyc/W  |  0.80  |  0.21  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420852-9973-000/Objective-LCI-Plan-Neofluar-25x-0.8-Imm-Corr-DIC-M27|420852-9972-000]]  |
|  40x  |  Plan-Apochromat Corr  |  Air  |  0.95  |  0.25  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420660-9970-000/Objective-Plan-Apochromat-40x-0.95-Corr-M27|420660-9970-000]]  |
|  40x  |  Plan-Neofluar Ph3  |  Oil  |  1.30  |  0.20  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/440451-9903-000/Objective-EC-Plan-Neofluar-40x-1.30-Oil-Ph3|440451-9903-000]]  |
|  100x  |  Plan-Apochromat DIC  |  Oil  |  1.40  |  0.17  |  [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/440782-9902-000/Objective-Plan-Apochromat-100x-1.40-Oil-DIC|440782-9902-000]]  |

==== Camera ====
^  Model  ^  Type  ^  Frame Size  ^  Pixel Size (µm)  ^
|  [[https://www.zeiss.com/microscopy/int/products/microscope-cameras/axiocam-506-mono.html|Axiocam 506 mono]]  |  CCD  |  2752 x 2208  |  4.54 x 4.54  |

===== System Turn On Procedures =====

  * Turn on the HXP 120 fluorescent light source (if used)
{{zeiss_axio_observer_hxp.jpg?120|}}
  * Turn on the temperature and CO2 controller - white power strip (if you need them).
  * If you need CO2, open the Cell Observer CO2 valve
  * Turn on the motorized stage controller (**1** on remote control)
{{zeiss_axio_observer_a.jpg?height=150|}}
  * Turn on the microscope power supply (**2** on remote control)
{{zeiss_axio_observer_b.jpg?height=150|}}
  * Turn on the computer
  * Login in to Windows with your Agendo credentials
  * Start the **ZEN** software
  * Select HXP or LED Workspace in ZEN (top left)
{{::workspace_selection.png?250|}}

===== System Turn Off procedures =====
If there is another user for this microscope is the __next hour__:
  * Leave the fluorescent light source on
  * Clean up immersion objectives
Else:
  * Exit the ZEN software
  * Turn off the computer (not the UPS!)
{{:cell_observer_pc_vs_ups.png?250|}}
  * Clean up immersion objectives
  * Turn off the microscope power supply (**2** on remote control)
  * Turn off the motorized stage controller (**1** on remote control)
  * Turn off the Temperature and CO2 controller - white power strip (if used)
  * Close the Cell Observer CO2 valve (if used)
  * Turn off the HXP 120 fluorescent light source (if used)

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