Cytometry

Equipments

Cytoflex bench-top cytometer (Beckman-Coulter)

This Cytoflex is equipped with a 405-nm laser with 4 fluorescence detectors, a 488-nm laser with 2 fluorescence detectors, a 561-nm laser with 4 fluorescence detectors and a 640-nm laser with 3 fluorescence detectors, in addition to Forward- and Side-scatter detectors. Fluorescence detectors are avalanche photodiodes (APD), much more sensitive than photomultipliers (PMT). It is equipped with a plate reader and allows an absolute volumetric counting.

The optical diagram of the machine will help you choose your fluorochromes.

This modern, economical, user-friendly and intuitive cytometer is dedicated to autonomous users. Training by the engineer in charge makes it possible to understand the functionalities in a few hours.

The optical diagram of the machine will help you choose your fluorochromes.

MoFlo Astrio sorter (Beckman-Coulter)

This flow cytometer provides the technology and the power necessary for the simultaneous reading of more than 20 fluorescence parameters and for the sorting of events, even if they are rare, in a population of heterogeneous objects. It is equipped with 22 photomultipliers, and 5 laser lines (355 nm / 100 mW; 405 nm / 55 mW; 488 nm / 150 mW; 561 nm / 200 mW; 640 nm / 100mW). Sorting can be accomplished in six ways (harvest in 5ml or eppendorf tubes) or on a plate (6 to more than 96 wells) with a reading speed of 70,000 events per second. Since January 2015, the analyzer-sorter has benefited from a module allowing the detection of small events (≥200 nm).

Although it is difficult to train for sorting with occasional use, training in autonomy in analysis is possible and recommended for routine experiments.

The optical scheme of the machine will help you choose your fluorochromes.

Available software

Analysis station with the following software:

  • FlowLogic 7.2 (Miltenyi-Biotech / Inivai)

For the analysis of flow cytometry data with 3 modules:

Analysis (gateLogic): overlays, heat maps, cell cycle, kinetics, cell proliferation

Statistics (graphLogic): statistical analyzes and creation of graphs

Report (docLogic): layout, pdf generation

 

  • Kaluza Analysis Software (Beckman Coulter)

Complete cytometry software, with the possibility of exporting fcs data to csv

  • CytExpert 2.2 (Beckman Coulter)

Basic and free software that controls the cytoflex

 

  • Summit 6.3 (Beckman Coulter)

Basic and free software that controls the sorter

 

  • R and Bioconductor tools (3.6)

Services

To define the type of device most appropriate to your problem and the desired level of access, a first discussion with one of the members of the platform is recommended.

Three levels of access are offered to our users:

The provision of assisted service concerns work carried out by the platform on behalf of a user. It implements techniques and know-how mastered by the platform. The work done in this type of service is not inventive.

An analysis of the results obtained will be provided in PDF format. It implies recognition of the engineer’s investment in possible publications. The platform does not guarantee the retention of data at the end of the service.

The use of equipment requires prior user training by one of the platform members. This training results in the acquisition of autonomy allowing the user to reserve the equipment on which he has been trained.

Use of the equipment after training is suggested when the user’s needs require repeated use of the equipment.

Before being fully autonomous on the machines, the user must agree to comply with the platform access conditions by completing an “access request”. The user’s team leader must also agree by signing this document.

The platform must be thanked in any publications.

In addition to the training provided as part of our service offers to lead to the autonomy of use of equipment, Imagerie-Gif organizes training in the form of theoretical courses, practical work and tutorials in specific areas of cytometry : link to training page.

The purpose of scientific collaboration is to remove a methodological barrier, in direct collaboration with a user. Collaboration is particularly implemented when a user’s request requires adapting protocols or carrying out specific developments.

The engineer involved will be considered in the position of co-authors in a possible publication.

Acknowledgments

The funding requests necessary for the development of the Imaging-Gif division make it essential that the activity of the platforms be explicitly recognized.

The funding requests necessary for the development of the Imaging-Gif division make it essential that the activity of the platforms be explicitly recognized.

The present work has benefited from Imagerie‐Gif core facility supported by l’Agence Nationale de la Recherche (ANR-11-EQPX-0029/Morphoscope, ANR-10-INBS-04/FranceBioImaging ; ANR‐11‐IDEX‐0003‐02/ Saclay Plant Sciences )

In case of publication by co-authors: Indicate this address:

Cytometry / Electronic Microscopy / Light Microcopy Facility, Imagerie-Gif,, Université Paris-Saclay, CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), 91198, Gif-sur-Yvette, France

Remember to send the platform manager a PDF copy of the article or any other communication.

Flow cytometry: mickael.bourge@i2bc.paris-saclay.fr

Electronic Microscopy: claire.boulogne@i2bc.paris-saclay.fr

Photonic microscopy: romain.lebars@i2bc.paris-saclay.fr

Conditions of Use

  • Access to equipment

Before any training or service, the user must complete an “access request” to agree to the proposed service and the proposed access terms. The user’s team leader must also sign this document. Upon signing, the team leader thus acknowledges having read and accepted the terms of use of the platform’s resources.

 

By signing this document, the user AND his/her team manager therefore agree to:

– respect the health and safety rules and the operating rules of the platform,

– provide all the information on the samples analyzed, in particular with regard to the potential risks for the manipulator and / or the material and inform the platform engineer of any change in biological material,

– respect the reserved hours of use,

– accept the billing conditions for services performed on the platform

– notify with acknowledgments any publication / presentation of data acquired on the platform

– enhance the investment of engineers

 

By signing this document, the platform for its part undertakes to:

– advise and guide the user towards the system (s) most suited to their needs,

– provide the user with maintained, checked and calibrated equipment,

– train users to support them towards partial or total autonomy in using the desired equipment,

– provide technical and / or scientific support,

– improve the quality of services offered by implementing a quality management system certified ISO9001 and NFX-50-900.

 

If these instructions are not followed, access to the various services may be called into question.

 

  • Sample compliance

The platform laboratory is only authorized to handle level-1 risk biological samples. Higher-level samples must be inactivated before being brought to the platform. The platform is not equipped to support the study of radioactive samples.

In certain cases, the platform engineer reserves the right to refuse a service if he judges that the samples present too many biological risks.

 

  • Machine reservation

Reservation is compulsory and is done via the GIPSI website.

Once trained in the use of one or more equipment, the user receives his identifiers by email in order to be able to connect to the intranet of the GIPSI website.

To access the reservation, simply log on to the left of the screen with the email address and password received. Those who have lost their password can request a new one by clicking on “Forgotten?…” Below the connection login.

To reserve equipment, simply select the platform of your choice in the “Platforms” tab, then go to the “Reservations” tab.

 

  • Storage of data and analysis results

The platform does not guarantee the retention of data at the end of each session. The user is therefore solely responsible for archiving and safeguarding his personal data.

 

The data acquired or processed can be retrieved by the user at the end of each service (autonomous or assisted by the engineer) on a USB key or on an external hard drive. In the case of a service performed by the platform engineer, a hard copy or computer copy (sent by email) can also be provided to the user at his request.

 

NOTE: The duration of the acquisition / analysis session MUST take into account the transfer time of acquired or processed data. No data should remain stored on the local drive.

 

  • Equipment maintenance

The platform monitors the stability and good condition of this equipment. This is essential to ensure our users have good reproducibility in their acquisitions.

For his part, the user is obliged to report any problem encountered when using the devices by completing an anomaly section made available on his reserved slot on GIPSI.

 

  • Intellectual property & confidentiality rules

Intellectual property

The know-how and knowledge used by Imagerie-Gif to provide the services remains the property of the CNRS; consequently any improvement in know-how will remain the property of the CNRS. The property of the results of the services belongs to the customer.

Retention of Title clause: CNRS RETAINED OWNERSHIP OF THE RESULTS UNTIL FULL PAYMENT OF THE PRICE OF THE SERVICES CONCERNED BY THE CUSTOMER. IN THIS RESPECT, AND IN PARTICULAR IF THE CUSTOMER IS THE SUBJECT OF AN ADJUSTMENT OR A JUDICIAL LIQUIDATION, THE CNRS RESERVES THE RIGHT TO EXERCISE THIS CLAUSE OF RESERVATION OF OWNERSHIP ON THE RESULTS OF THE SERVICES CONCERNED, TO THE COMPETITION OF THE ARE OWNED BY THE CUSTOMER.

 

Confidentiality rules

Each party (user and platform) undertakes not to publish or disclose in any way whatsoever the scientific, technical or commercial information of the other Party, and in particular prior knowledge belonging to the other party of which it may have knowledge during the performance of the services and this, as long as this information is not accessible to the public. This commitment will remain in effect for five years (5 years) from the date of issue of the invoice.

Any publication or communication from the user relating to the results of the services must indicate that these results were obtained by the platform. It must in no case disclose the know-how implemented by the platform.

 

  • Imagerie-Gif platform charter
  1. Daily hours, opening of the laboratory, access to the premises

Users can choose their arrival time within the range 9:00 am – 6:00 pm.

Access to the I2BC outside of these ranges may be expressly and by name authorized by the technical manager of the Platform.

Users must comply with the health and safety instructions posted in the premises made available to them (see also paragraph 3.4 Isolated work of this charter).

 

The Platforms operate throughout the calendar year. An engineer absence table is clearly displayed, users must take these absences into account when scheduling supported work. Any closure periods are decided by the scientific manager of Imagerie-Gif.

 

  1. Dissemination of scientific results
    • Confidentiality

Everyone is required to respect the confidentiality of the work entrusted to them as well as that of their colleagues. In case of disclosure of information (publications, patents, communications based on the use of the resources of a Platform), it is strongly recommended to submit the Materials and Methods / Results parts obtained on the Platform’s equipment to the assessment of the technical manager of the Platform. The results and data from research carried out on the Platforms are not properties of the Platform but are subject to the regulations of the various Institutions covering each user. On the other hand, the originals of the laboratory notebooks of the engineers involved in the support must be kept on the Platforms.

Platform engineers are authorized to use the results acquired on the Platform in their actions to promote the platform under cover of the authorization of the project authors.

 

  • Publications

Publications involving results obtained on one of the Imaging-Gif Platforms must specify this in the acknowledgments. If engineers appear as a co-author, their membership in the unit and their attachment to the supervisory bodies must appear in the address.

A copy of all publications (articles, journals, theses, etc.) for which all or part of the work has been carried out within the platforms must be submitted as soon as they are published to the technical manager of the Platform.

 

  1. Hygiene and safety
    • security

Platform staff must ensure that user agents have received security training and, where applicable, specific training adapted to their work on the Platform. Otherwise, he agrees to have them follow these training courses.

 

  • No smoking

Smoking is prohibited in the workplace (Decree 92-478 of May 29, 1992).

 

  • Isolated work

Isolated work must be exceptional and devoted to tasks that do not present any risk. In the event that hazardous work must necessarily be carried out outside of official opening hours, it is compulsory to be accompanied. The line manager and the technical manager of the Platform must be notified.

 

  • M.O.: Specific measures

The use of GMOs (Genetically Modified Organisms) is regulated. Official information on their use can be found on the following sites:

http://www.cnrs.fr/SDV/ogmcircul.html; http://www.cnrs.fr/SDV/ogmrecap.html

The main rules for the handling of GMOs at I2BC are as follows:

  • All GMOs must be handled, stored, decontaminated and disposed of in such a way as to:
  1. Do not harm the health of users
  2. Avoid contamination of the biological material of other users
  3. Avoid dissemination
  • GMOs include microorganisms (bacteria, viruses, yeasts, etc.), plants and transgenic cell cultures
  • Before disposal, all GMOs must be sterilized by treatment with bleach or autoclaving
  • Transgenic plants must be cultivated and handled in a greenhouse or in enclosures according to the rules which can be consulted in these places and on the Intranet.
  • The level of containment (L1, S1, S2 and S3) required for each type of experiment has been notified for each project by the Commission for Genetic Engineering (CGG). This information is available for consultation in the application files for the contained use of genetically modified organisms (groups I and II) submitted to the I2BC secretariat and available on the Intranet
  • Any new type of experiment must be the subject of a request for modification of the GMO contained use approval file.

 

  • Radioactivity: Specific measures

The user using radioactive markings must notify the technical manager of the platform from the first appointment.

 

  1. Training

Engineers inform and advise users for their training needs and requests. They will eventually set up specific training appropriate to the needs expressed.

 

  1. Use of IT resources

Each user of the Platforms hosted at the I2BC and using the I2BC’s IT resources must be aware of the I2BC IT charter.

It is strictly forbidden to use the computers dedicated to the management of the Platform equipment for purposes other than the acquisition or analysis of data related to the equipment (prohibition of Internet or intranet consultation, email, installation of programs, etc.).

 

  1. Use of collective technical resources

Engineers are responsible for maintaining equipment and laboratories. However, it is essential that the conditions and rules of use are strictly applied by each user. Platform engineers must ensure that users are trained and informed about security rules. They reserve the right to prohibit access to the Platform in the event of non-compliance with rules of behaviour.

Rates

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Training

Training soon
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