Developmental Biology
a worm

Utrecht University Large-Particle Flow Cytometry Facility (UU-LPC)

General Information

Utrecht University Large-Particle Flow Cytometry Facility (UU-LPC) General InformationThe selection and purification of molecules, cells, tissues, and organisms of interest are critical yet often time-consuming aspects of biomedical research. Flow cytometers have revolutionized the sorting and analysis of large numbers of individual cells. However, many objects are too large or too sensitive for conventional flow cytometry. For this reason, Union Biometrica has developed large particle flow cytometers able to handle a wider range of object sizes (1 – 1500 µm).The UU Large-Particle Flow Cytometry Facility provides the life sciences research community in the Netherlands with access to the latest model large particle flow cytometer: the BioSorter. The facility represents the first placement of a BioSorter in The Netherlands, and was established with financial support from the Netherlands Organisation for Scientific Research (Investment Grant NWO Medium), and from Utrecht University (Support Core Facilities, Life Sciences).Examples of materials that can be analyzed and sorted with the BioSorter are combinatorial chemistry beads and particles, microcolonies of Aspergillus and filamentous bacteria, plant seeds, zebrafish embryos, C. elegans embryos and larvae, Drosophila embryos and imaginal disks, large cells, cell clusters, cultured organoids, and embryoid bodies.

Fig.:Examples of organisms sorted with the BioSorter: C. elegans , Aspergillus niger with GFP and dTomato expression (Brand Rector, Utrecht University) and Arabidopsis thaliana seeds (Tatyana Radoeva, Wageningen University)

Facility equipment

The large-particle flow cytometry core provides sophisticated facilities for work with a broad range of cell culture and model organisms. These facilities allow preparation of samples that cannot readily be transported and need treatment on-site before or after analysis and sorting.

Tissue Culture facilities

Tissue culture facilities, including a tissue culture hood, CO2 incubator, water bath, fridge, freezer and centrifuge are available for use. In addition, stereomicroscopes are available to examine materials before and/or after sorting.

Tecan robotics

The facility provides access to a Tecan Freedom EVO 100 platform equipped with a 96-channel pipetting head.

Microscopy support

Low power microscopes are available in the facility, and a dedicated high power microscope is located in an adjoining room. Moreover, the facility is housed on the same floor as the Biology Imaging Center of the Institute of Biodynamics and Biocomplexity, which provides access, support and training in advanced light and fluorescent microscopy techniques for research groups within the department of Biology, as well as for researchers from other institutes within or outside of Utrecht University.

BioSorter details

The BioSorter flow cytometer is a continuous flow system capable of analyzing and sorting objects based on size, optical density, and fluorescence. Relative size and optical density are measured by a PIN photodiode. Fluorescence intensities can be simultaneously measured at three different wavelengths by three high-sensitivity PMTs. Sorting and dispensing decisions are based on user-selected ranges using the FlowPilot™ Pro software. The BioSorter flow cytometer can be placed in a tissue culture hood for sterile operation.Object size range The BioSorter uses easily exchangeable fluidics modules optimized for a particular size range. The following modules are available in the facility:

FOCA

The FOCA is an assembly of flow cell, pre-aligned beam focusing optics and fluidics components. Depending of the size of the sample the correct FOCA will be built into the BioSorter.

  • FOCA  250
    • size range: 20 – 150 µm
  • FOCA 1000
    • size range: 200 – 700 µm
  • FOCA 2000
    • size range: 500 – 1500 µm

Fig.: 250 , 1000 and 2500 FOCA

Excitation

  • 405 nm / 100 mW laser
  • 488 nm / 50 mW laser
  • 561 nm / 50 mW laser

Emission (exchangeable filter sets)

  • DAPI, BFP, GFP, YFP, DsRed

Sample introduction

The sample can either be introduced by a sample cup or an automatic Large Particle Sampler. Both allow sample introduction into variuos solutions. Incidentally, density of your sample determines the sorting time and efficiency. For sample analysis a rate until 500 events/s is possible; for sorting 10 events/s result in high recovery (~95 %).

  •   50 ml sample cup
  • 750 ml sample cup (coming soon)
  • Automated Large Particle Sampler

Sample Cup

The sample cup is provided with a suspended stirrer with alterable velocity and switching rotation.

Large Particle Sampler (LPS)

The automated Large Particle Sampler (LPS) allows gently aspirating of samples from multiwall plates or petri dishes. Prior the transfer, the sample will be suspendend. Than the samples will be automatically delivered via a flow system to the BioSorter where they will be analyzed and sorted. The anaylysis of the contents of a full 96 well plate requieres approxiamtely 45 min (100 ul volume per well).

Fig.: LPS

Sorting templates

An X-Y-Z stage allows dispensing into 24-, 48-, 96-, or 384-well microtiter plates, tubes and bulk receptacles. The Z-direction allows adjustment for different height plates and tubes. Further, calibration of individual sorting templates is possible.

Access and information

Fees will be applied to cover repairs and part replacements following the warranty period, to cover the materials used and to compensate for operator support. Users from the UU will be charged 20€ per hour. We will charge external users from academia 50 € per hour, and users from industry 100 € per hour. These amounts will be determined in part on total user time and exact terms can be discussed. Billing will be automated according to the booking information. No show after booking will be billed as a consequence of obstruction by use of other parties.

For more information and to arrange access to the facility, please contact Dr. Juliane Teapal