Wien | Website
The microflow cytometer Micro-PLUS Flow Cytometer (Apogee) can be used for the characterization of cells and bionanoparticles in the fermentation broth. Cell characterization ranges from size and shape determination of large populations with indications for cell-state (replicating and non-replicating cells) to transfection and infection efficiency screening for virus production development and transient recombinant protein production development. The instrument is capable of characterizing bionanoparticles as well as of reliably determining size of particles around 80nm and above, characterizing extracellular vesicles, virus like particles (VLPs) and viruses. Detection is done with light scattering detectors after excitation by laser and – if samples are labelled – with fluorescence detectors, which is especially important for the determination of transfection and infection efficiency. The device has different lasers (405 nm, 488 nm, 561 nm, 638 nm) and fluorescence detectors to cover a large spectrum of different fluorescence markers. The device is equipped with an autosampler.
Dr. Martin Trinker
acib offers extensive and reliable partnerships in national and international research projects, as well as the implementation of contract research.
Methods & Expertise for Research Infrastructure
The instrument can be used for several purposes. The ability to determine scatter profiles and fluorescence for individual cells enables the characterization of a cell population.
Determination of cell state and kind: The scatter profile determined by the instrument yields a distinction of cells due to the kind of cells (e.g. lymphocytes vs. myeloid cells), and a characterization of the replication state of the cells in their replication cycle (cell cycle analysis of G0/1 or G2 phase cells).
Determination of transfection/infection efficiency: By introducing a fluorescence label into the production system (e.g. coupling the recombinant protein or virus protein with green fluorescence protein) a transfection efficiency for transient expression of recombinant proteins, or an infection efficiency for the production using viruses can be determined. The instrument yields a population of transfected/infected cells as well as uninfected cells, and the amount of fluorescence is a direct indication of how much recombinant protein/virus protein is produced within the cell.
Characterization of VLP/virus population: Using the scatter profile of the violet laser (405nm) yields a characterization of particle populations, like VLPs. This population characterization can be used to track healthy and unhealthy cells through the composition of their VLP-population, as well as serve as a quality control during production and purification of VLPs and virus particles.
Allocation to Core Facility
2. Forment, J. V, Walker, R. V & Jackson, S. P. A high-throughput, flow cytometry-based method to quantify DNA-end resection in mammalian cells. Cytometry. A 81, 922–928 (2012).
3. de Almeida Santiago, M. et al. Flow Cytometry as a Tool for Quality Control of Fluorescent Conjugates Used in Immunoassays. PLoS One 11, e0167669–e0167669 (2016).
4. Homann, S. et al. A novel rapid and reproducible flow cytometric method for optimization of transfection efficiency in cells. PLoS One 12, e0182941–e0182941 (2017).
5. Blackstock, D. J. et al. Comprehensive Flow Cytometry Analysis of PEI-Based Transfections for Virus-Like Particle Production. Res. (Washington, D.C.) 2020, 1387402 (2020).
6. Tang, V. A., Renner, T. M., Fritzsche, A. K., Burger, D. & Langlois, M.-A. Single-Particle Discrimination of Retroviruses from Extracellular Vesicles by Nanoscale Flow Cytometry. Sci. Rep. 7, 17769 (2017).
7. Görgens, A. et al. Optimisation of imaging flow cytometry for the analysis of single extracellular vesicles by using fluorescence-tagged vesicles as biological reference material. J. Extracell. vesicles 8, 1587567 (2019).