Frequently Asked Questions - FCF

 

General information – FCF stands for Flow Cytometry Facility

1. Where exactly is the FCF lab-space located?

In the clinical complex UK Aachen, level 5, floor 41, room 07, accessible via elevators D1 or C1

2. How much does it cost to use FCF?

Users have to pay for the service of cell analysis and cell sorting.

The fee depends on the user group:

  • Group I: Members of RWTH Medical Faculty or IZKF
  • Group II: Members of any other RWTH Faculty or other public research institutes

Exact prices can be requested at FCF.

3. How are FCF user fees billed?

  • FCF services will be charged to the users.
  • Internal service charges and external invoices are billed every three months
  • To that end every user is obliged to fill in an order form (“Auftragsformular”) and to have it signed by his supervisor to whom the invoice will be addressed. The order form has to contain the correct address to ensure that the bill can be sent without problems.
  • The prices for the use of the devices depend on the duration of the measuring time. The calculation is based on the reserved measuring time or the actually required measuring time, in case this exceeds the reserved measuring time.

4. How does the payment process for internal cost allocation work?

  • You will receive an internal cost allocation from us. Please draw this "factually correct" and indicate from which internal order (“Kostenstelle”) the amount is to be paid.
  • Finally, you submit the internal cost allocation to Mrs. S. Kessel in the Financial Management Division.

5. What does compensation mean?

In FACS analyses, several fluorescent dyes are usually used at the same time. Signals of the individual dyes are measured by various detectors.

Unfortunately, most fluorescent dyes do not emit strictly one wavelength, but a whole spectrum. Thus, the fluorescence signal of one dye not only reaches the channel or detector provided for this dye, but also causes a (albeit weaker) impulse in other channels. This impairs the evaluation of the results, since in multicolor analyses it cannot be known which dye caused the detector signal.

However, since simple experiments allow to determine how strongly one dye scatters into another channel, this problem can be corrected mathematically. This process is referred to as ‘compensation’.

6. What do I need for compensation?

The following samples are required for successful compensation:

  • Negative control: cells/beads to be analyzed without any dyes
  • Single stainings: For each dye/antibody a single color staining must be provided. This should contain both, cells positive and negative for that dye

7. How long is the FCF's lab space open  service time?

The FCF service time is usually weekdays from 8am to 4pm. During this time the laboratory is open and accessible to everyone.

8. What do I have to do if I want to use FCF outside service hours?

Reliable users able to use FACS Canto independently can get access to the lab at any time.

In the corridor next to the laboratory door there is a safe which contains the key. The required pin code can be obtained from FCF.

9. Why do I have to filter my cells before use?

All samples have to be filtered before measurement to remove any cell aggregates that may have formed to prevent clogging of the instruments!

10. How should I filter my cells before use?

  • For filtration cell filters with pore sizes of 35 to 70µm should be used (for example Falcon Tube, 35µm Cell Strainer (Art. Nr.352235)).
  • When filtering cells, the cell size must be taken into account! Cells larger than 70µm have to be filtered with 100µm filters.
  • Except of sterile cell sorts there is no need to use sterifilters.

 

Cell analysis: Use of BD FACS Canto

1. What do I need to do when I want to use BD FACS Canto (cell analysis) for the first time?

  • Please contact FCF (and ask for an appointment for a briefing.
  • Read the user regulations that can be found on the FCF homepage.
  • You will receive an order form from FCF that has to be signed by your supervisor (that person to whom the invoices will be sent).
  • Bring the signed user regulations and the completed order form with you for the briefing!
  • Once all documents are complete and the briefing done, you will receive your own account, which will allow you to use FACS Canto independently.

2. How can I require a briefing for BD FACS Canto?

  • Please contact and ask for an appointment for the briefing.

3. What do I need to perform cell analyses using BD FACS Canto?

  • 5 ml FACS tubes (Art.Nr. 55.1579, Sarstedt)
  • Staining Buffer, for example PBS
  • Unstained sample (to serve as a negative control)
  • Stained sample(s)
  • If compensation is required: a probe (cells or beads) that is only stained with one respective fluorochrome.  Such probe is required for each fluorochrome used in the multicolour staining. The probe should contain both, cells/beads positive and negative for this fluorochrome.
  • Cell filter with a pore size of appr. 40µm
    depending the size of cells/beads in your sample; for example cell filter with attached tube, Falcon, Art. Nr. 352235

4. How can I make a reservation for FACS Canto?

  • Come to the lab space. Here (on top of FACS Canto) a calendar is available in which you can manually reserve your measuring time.
  • For users outside UK Aachen (upon previous agreement) appointments can also be made by phone call (+49 241 80 37654).
  • An online reservation system is currently being established.  
  • Please reserve only the amount of time you really need and arrive on time for your measurements.
  • Due to the high utilization of the device it is recommended to book the desired measuring time two weeks in advance.

5. What do I have to do if I cannot attend a booked appointment?

  • We kindly ask you to delete reserved dates that you cannot use from the calendar. This gives other users the chance to hop in.
  • For users outside UK Aachen appointments can be cancelled by phone call (+49 241 80 37654).
  • If agreed deadlines cannot be met, the user can cancel the appointment free of charge within two days before the measurement. Otherwise, cancellation costs will be charged according to the reserved measurement time.  

 

Cell sorting: Use of BD FACS Aria

1. What do I need to do when I want to use BD FACS Aria (cell sorting) for the first time?

  • Please contact  and ask for an appointment for a briefing.
  • Read the user regulations that can be found on the FCF homepage:
  • You will receive an order form from FCF that has to be signed by your supervisor (that person to whom the invoices will be sent).
  • You will receive a document from FCF that requests details of the sort, such as the type of cells and conditions of the sort. This document must be provided to FCF at least one day ahead of the sort (pdf file or printout).
  • Bring the signed user regulations, the required information regarding your cell sort and the completed order form with you at the date of your cell sort.

2. What do I need for cell sorting at BD FACS Aria?

  • All required and completed documents (see above)
  • 5 ml FACS Tubes, for example Art.No. 352054, Falcon oder Art.No. 55.1579, Sarstedt
  • Staining Buffer, for example PBS or cell culture medium without FCS/BSA
  • Sort Buffer (for example cell culture medium with FCS/BSA) to store the cells after the sort
  • Unstained probe (to serve as a negative control)
  • Stained probe(s)
  • If compensation is required: a probe (cells or beads) that is only stained with one respective fluorochrome.  Such probe is required for each fluorochrome used in the multicolour staining. This probe should contain both, cells positive and negative for the fluorochrome.
  • A cell filter with a pore size of appr. 40µm (depending on the size of your cells/beads; for example filter with attached tube Falcon, Art. No. 352235), sterile if required
  • Collecting tubes for the cells post sorting
    • 1.5mL Eppendorfer cups: up to 4 populations of cells, with a maximum of 0.5 Mio. cells
    • 5mL FACS tubes:  up to 4 populations of cells, with a maximum of 2 Mio. Cells
    • 15mL Falcon tubes: up to 2 populations of cells with a minimum of 1 Mio cells

3. How can I make reservations for BD FACS Aria?

  • The Online calendar will give you an overview about available time slots.
  • Please contact FCF and agree upon a date for cell sorting

4. What do I need to do if I cannot attend a booked appointment?

Within two days in advance the user can cancel the agreed date free of charge (via  or phone call: +49 241 80 37654).

Otherwise, cancellation costs will be charged according to the reserved measurement time. 

 

Spectral cell analysis: Use of FACS Aurora (Cytec)

1. What do I need to do when I want to use FACS Aurora (Cytec) (spectral analysis) for the first time?

  • Please contact  and ask for an appointment for a briefing.
  • Read the user regulations that can be found on the FCF homepage:
  • You will receive an order form from FCF that has to be signed by your supervisor (that person to whom the invoices will be sent).
  • Bring the signed user regulations and the completed order form with you for your briefing!
  • Once all documents and the briefing are successfully completed, you will receive your own account, which will allow you to use FACS Aurora independently.

2. How can I require a briefing for FACS Aurora (Cytec)?

  • Please contact  and ask for an appointment for the briefing.

3. What do I need to perform cell analysis with help of FACS Aurora (Cytec)?

  • 5 ml FACS tubes (Art.No. 55.1579, Sarstedt)
  • Staining Buffer (for example PBS)
  • Unstained probe (to serve as a negative control)
  • Stained probe(s)
  • If compensation is required: a probe (cells or beads) that is only stained with one fluorochrome. Such probe is required for each fluorochrome used in the multicolour staining. The probe should contain both, cells positive and negative for this fluorochrome.
  • Cell filter with a pore size of appr. 40µm (depending on the size of your cells/beads; for example a filter with attached tube, Falcon, Art. No. 352235)

4. How can I make a reservation for FACS Aurora?

  • Upon briefing each user gets access to an Outlook calendar. Measuring times have to be reserved in this calendar in advance.
  • Each user is only permitted to administrate his own measuring times and it is strictly prohibited to change or delete measuring times of other users.
  • Upon previous agreement users outside UK Aachen can also make reservations by phone call.
  • An Online reservation system is currently being established.
  • Please reserve only an amount of time that you really need and arrive on time for your measurements.

5. What do I have to do if I cannot attend a booked appointment?

  • We kindly ask you to delete reserved dates that cannot be used from the calendar to give other users the chance to hop in.
  • For users outside UK Aachen appointments can also be cancelled by phone call.
  • If agreed deadlines cannot be met, the user can cancel the appointment free of charge within two days before the measurement. Otherwise, cancellation costs will be charged according to the reserved measurement time.