Instructions for using the on site equipment
Before using the analysis instruments (CytoFlex, Cytek Aurora, Navios, FACS Fortessa, FACS Accuri), you will receive an instruction ("driving license"). This briefing is device-specific, intended to ensure proper handling of the equipment and usually carried out and documented by the head of the facility. After the briefing and once registered in the booking system, you will get the permission to use devices and thus the authorization to access and manage your own time slots. This instruction does not replace the offered internal trainings.
Registration with the online booking system
Before making reservations you will have to register via the jointly provided internet booking system. For more informations, see Booked. To register, click here: https://tumemi0-booked.srv.mwn.de/. Please note that for billing reasons, it is important for us to get as much informations as possible about your working place.
Before you start your experiment, it would be usefull to contact the staff of the facility to help you with design, planing and the best possible staining combination of Fluorophores to prevent spectral overlaps or compensation problems.
- Appointments should be planned realistically and, in the interest of all, times entered should be adhered to.
- If your booked appointment for an analyzer cannot be met, delete the reservation in the online calendar as soon as possible, so that others get a chance of short notice booking.
- The scheduled times are binding for user and CyTUM staff and are "sin tempore".
- Additional cell material that extends the requested time will not be sorted.
- if your experiment setup takes longer than expected, call the sorting operator as soon as possible. Delays will only shorten the timeslot reserved for you. You will also be immediately informed by the operator of any malfunctions of the equipment and the implied time differences. In both cases, the CyTUM staff will try to find an appropriate time solution.
- Sorting appointments can only be scheduled 2 weeks in advance. if you need a fixed time slot for experimental reasons, please indicate so beforehand in your sort registration mail.
For new users, introductions to the analyzers ("driving license") are carried out regularly.
- Plan your appointments realistically and, in the interest of all, stick to your time slot.
- A "warm-up" time of 15 minutes on the first appointment of the day is mandatory for all equipment.
- For the FACS Fortessa and FACS Accuri, an additional cleaning time of at least 15 minutes must be included at the end of the session for each experiment.
- An entry of more than 3 consecutive hours in the core time of 10:00 - 16:00 is only allowed in exceptional cases and requires the agreement of the head of CyTUM.
The following basic rules are to be bindingly observed when using the online reservation system. These guidelines apply to flow cytometric analysis appointments:
- You must execute the switch-on and switch-off routines for the analyzer as explained in the instruction.
- Hand over the device directly to the following user or make sure, e.g. by phone call, that the next user will attend his analysis appointment. This is to prevent devices from running over night or in worst case over the weekend.
- By using a device, you accept bindingly the device-specific instructions on your own responsibility.
- Damage to the analyzer caused by your faulty behavior will have to be covered in full by you or your organisation.
- At the end of the analysis, make a control measurement with water (with settings like duration, measurement speed and treshhold according to the specification of the respective device) and document it by saving the analysis file.
- Document each analysis in the log book available at the device, as well as any error messages that occured or problems encountered while using the instrument. In addition, in case of malfunctions, you have to inform a CyTUM employee immediately.
- No samples containing radioactivity, or potentially dangerous germs for humans may be measured on the analyzer. Fixed samples are considered non infectious.
- Filter your samples before loading (nylon filter material - 100 µm)
- Always inspect your samples for clumps and aggregates. For analysis purposes, the cell density should not exceed a cell count of 1 x 108 cells/ml.
- Once done, you are responsible for sheath fluids refilling and waste disposal. Remove all sample materials present and clean the workplace with a wipe disinfection.
The use of the sorting devices (FACS Aria, FACS Fusion, Astrios and S3) is possible without instruction, whereby the actual device handling is being done by the operators of CyTUM. Appointments at MIH have to be requested with the CyTUM staff by e-mail (firstname.lastname@example.org) who will then manage the booking in the online reservation system. A filled out sort registration form (available as download on the CyTUM website) must be attached to the mail.
Basic rules for sorting
- The scheduled times are binding for you and the CyTUM staff and are "sin tempore".
- if your experiment setup takes longer than expected, call the sorting operator as soon as possible. Delays will only shorten the timeslot reserved for you. You will also be immediately informed by the operator of any malfunctions of equipment and the implied time differences. In both cases, we will try to find an appropriate time solution.
- No samples containing radioactivity, carcinogenic, evaporative, harmful or irritating buffers (eg. 2-Mercaptoethanol or Trizol) will be sorted at the CyTUM facility.
- If you have samples of biosafety level S2 according to GenTG, you have to prepare them in rooms of biosafety level S2. In the event of a clogging in the device (danger of aerosols), you will have to leave the sort room immediately and may only return after decontamination measures have been completed by a CyTUM employee.
- The selection of the device and the settings of the fluidic system are the responsibility of the operator. As a rule of thumb, the cell diameter of the sample must not exceed 1/5 of the Nozzle diameter. By default, we use a 70 µm Nozzle and the sort sample is filtered through a 30 µm FILCON. Indicate any necessary changes of the Nozzle size on the sort registration form. The cell density should ideally be 1 x 108 cells/ml for cell sorting.
- discuss any necessary modification of the equipment for special experimental requirements beforehand with the CyTUM staff. The time needed for the changes will be charged.
- Changes in Nozzle size require adjustment times of the fluidic system. Increasing the sorting stream leads to a downtime of the instrument of 12 hours - if the sorting stream is reduced, the fluidic system is usually stable again after 2 hours.
- The usage fee is determined quarterly for each user, rounded down to full ½ hr. and invoiced to the respective workgroup/project leader. The invoice amount is due immediately and is to be transferred to the account specified in the invoice.
- With payment of the usage fee, all necessary operating and consumable materials, reagents for quality assurance and personnel costs are settled.
- By creating a reservation entry, you accept the CyTUM usage rules on your own responsibility. Violations thereof will first be reprimanded by the head of the functional unit. In case of repetition, the head of the functional unit reserves the right to exclude you from further use of the equipment.
- Short notice sort cancellations or missed appointments may be fully charged.
- Damage to the equipment, which is clearly caused by ones own faulty behavior, will be compensated by you or your organisation, according to the legal regulations. In that case, you or your organisation will have to pay for the costs of repair work and spare parts required.
- Any necessary modification to the equipment for special experimental requirements must be discussed beforehand with the CyTUM staff. The time needed for the changes will be charged.
- Changes in Nozzle size require adjustment times of the fluidic system. For FACS Aria an additional 1 hour of booking time will be charged. For MoFlo Astrios an additional 4 hours of booking time will be charged.
Rules for Acknowledgement and Collaboration with the Flow Cytometry Facility
Our services are charged by criteria of DFG Core Facility standards and partly covers the expenses for sorting. Some of our instruments are equally funded by the DFG and the Institute of Medical Microbiology, Immunology and Hygiene but most of them are purchased by the Institute solely. In addition, instruments costs, labour and consumables are provided by the Institute and are not covered by the billed prices. Therefore, all public presentations or formal publications containing data from work performed at our Flow Cytometry Core Facility must be identified. We would suggest the following statement: "We would like to acknowledge the support provided by the Flow Cytometry Facility MIH of the Institute for Medical Microbiology, Immunology and Hygiene".
Exception from the guidelines above are sorts under BSL3 conditions, which are always considered as collaboration with staff and leader of the CyTUM Core Facility. We expect that our project input in such cases results in a co-authorship in case of formal publication or presentation. Nevertheless, the acknowledgment for using our BSL3 sorter should be: "We would like to acknowledge the support and funding for Cell Sorting provided by SFB/TRR-179 - Determinants and dynamics of elimination versus persistent Hepatitis-Virus-Infections - and the Helmholtz Zentrum München – Deutsches Zentrum für Gesundheit und Umwelt.”
Further on, co-authorship is expected for extensive consultation on experimental design, data analysis or involvement of technological advances made for the project with help of the CyTUM facility