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Flow Cytometry

The ZIK plasmatis uses different flow cytometers for daily measurements. There is proven experience in designing and compensating multicolor panels of 12-13 channels, both on the analyzers and on the cell sorter. A 4-laser (405nm, 488nm, 561nm, 633nm), 13-colour (+FSC/SSC) CytoFlex S (Beckman-Coulter) autosamples via a hydraulic pump from 96-well microtiter plates (flat-, V-, U-bottom). The optical arrangement of the lasers is parallel. The acquisition volume is 10-200μl per minute. For large sample sets, capacious 10l sheath and liquid waste containers are available. The device is frequently used for multiplex bead analyses of cell culture supernatants. Specially designed modules are available for evaporation protection when measuring small sample volumes. Data depth is 24bit (16 million channels) up to 10,000 events per second and a sensitivity of 200nm particle size. Build-in Avalanche photodiodes (APDs) are outperform regular photomultiplier tubes, since they have a higher sensitivity in the "red" fluorescence spectrum allowing excellent resolution of populations also with far-red dyes. Absolute volumetric measurements facilitate accurate cell / particle counts per microliter. The software (CytExpert) is available as freeware for the analysis of FACS data. Our 3-laser (405nm, 488nm, 638nm), 10-color (+FSC/SSC) Gallios (Beckman-Coulter) acquires samples from FACS tubes via compressor-driven hydrodynamic focusing. The optical arrangement of the lasers is parallel. The acquisition volume is 10-60μl per minute and for large samples sets a capacious 10l sheath and 20l liquid waste container is available. A 32‑tube autosampler provides the appropriate throughput, while more samples can for instance be stored in a second carousel in the refrigerator and measured immediately after. The free offline software allows generating acquisition settings and sample descriptors (worklists) at any office workplace for quick and simply loading of sample sets. Data depth is 20bit (1 million channels) at up to 25,000 events per second and a sensitivity of 400nm particle size. Our 1-laser (488nm), 4-color (+FSC/SSC) Attune Nxt Acoustic Focusing Cytometer (Applied Biosystems / Life Technologies) utilizes acoustic focusing to facilitate acquisition volume rates of up to 25-1000µl per minute. Interestingly, high volumes to not significantly affect the coefficient of variation (CV) of a (fluorescence) intensity of interest because in contrast to traditional cytometers the sample core stream is not becoming wider but faster. This is especially interesting for “no-wash” sample runs or samples were centrifuging should be avoided, while still providing sufficient data in a reasonable time. Moreover, HPLC pumps allow for the absolute counts of particles per microliter measured. We routinely use the Attune for accurate couting of viable cells among dozens of cell lines. This approach is not only very efficient and fast but also facilitates a “track record” of all counts per cell line and years back, which helps to context e.g. usual results. The Attune software for data analysis is freely available for office computers. Different software platforms are available for analyzing .fcs (FACS) data. In addition to above-mentioned freeware (Attune and CytExpert), our Summit software (Beckman-Coulter) helps analyzing sort data in the high 8-digit range (50 million events and more). This is especially important for the identification of rare populations (blood stem cells, etc.). A Flow‑Jo dongle is also available. However, our five Kaluza 2.0 (Beckman-Coulter) network licenses installed on all office and lab computers are analysis standard at ZIK plasmatis “plasma-redox-effects”.

Contact: Dr. Sander Bekeschus, sander.bekeschusinp-greifswaldde

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Kontakt

Leibniz-Institut für Plasmaforschung und Technologie e.V.

plasmatis – Plasma plus Zelle
Felix-Hausdorff-Str. 2
17489 Greifswald

Tel.: +49 3834 - 554 3319
Fax: +49 3834 - 554 301

plasmatisinp-greifswaldde
www.plasmatis.de/