Development of Flow Cytometry Flow cytometry was initially developed for medical applications, but has since evolved into a versatile tool used across various fields such as cell counting, classification, and biomarker detection [7]. Although the first study on using flow cytometry for plant cell nuclear analysis was published in 1973 [8], it wasn’t until later that plant DNA analysis using this technique gained significant traction. Since then, researchers have consistently applied flow cytometry in plant science. Botanists primarily use it to measure the DNA content within plant nuclei. For a detailed explanation of its principles and applications, refer to Reference 9. Figure 2a generally shows how a flow cytometer suspends dyed or targeted particles (with sizes ranging from 0.2 μm to 150 μm [7]) in a stream of hydrodynamically focused liquid through an electronic detection system. In plant studies, nuclei are first stained with a fluorescent dye, such as DAPI or propidium iodide. Each suspended nucleus is then exposed to a light beam, typically a laser or UV light, and the scattered light is detected to analyze fluorescence intensity, which corresponds to DNA content. The data from thousands of intact nuclei provide insights into the DNA content of tissues, as seen in the peaks of Figure 1. The quality and purity of the solution used for sample preparation and analysis are crucial, as the physical and chemical properties of thousands of particles are analyzed simultaneously every second. Using low-quality or inappropriate water can lead to contamination, causing non-sample particles to fluoresce and create "noise," interfering with results and leading to inaccurate assessments. Therefore, high-purity ultrapure water is essential for reliable flow cytometry experiments. Ultrapure water produced by an ultrapure water system meets ASTM Class I standards. Sartorius (Göttingen, Germany) provides such systems. In this study, ArUPH2O from the arium® pro VF system was used to evaluate its impact on flow cytometry results. The research focused on analyzing the reproductive pathways of angiosperms, particularly their seed development, by comparing results obtained using ArUPH2O and standard sheath solutions (0.04% sodium azide, 0.01% detergent) from Partec GmbH. The arium® pro VF system (Figure 3) produces ultrapure water by removing all contaminants from pretreated drinking water. It ensures continuous circulation and stable flow rates via a pressure-controlled pump system. Conductivity is monitored at both the inlet and outlet ports, or directly at the production port. The system described in this article uses two different filter cartridges filled with activated carbon and mixed bed ion exchange resin to produce ultrapure water with very low total organic carbon (TOC). It also includes an integrated UV lamp that operates at 185nm and 254nm to oxidize organics and kill bacteria. In addition, the arium® pro VF system features an integrated ultrafiltration module, acting as a tangential flow filter. This membrane traps colloids, microorganisms, endotoxins, RNA, and DNA. A 0.2μm end filter is installed at the outlet to remove particulates and bacteria after water purification. Refer to Figure 4 for the full purification process. Materials and Methods Seed samples were collected from hexaploid Ranunculus under free pollination conditions. Seeds were crushed in a plastic petri dish with 300 μl of extraction buffer (CyStain UV Precise P, Partec GmbH), incubated for 10 minutes at room temperature, and then transferred to a 5-mL plastic tube with 30 μl of CellTric® suspension. After adding 1.2 mL of staining buffer (CyStain UV Precise P), samples were analyzed using the blue fluorescent channel of a flow cytometer (CyFlow Space, Partec GmbH; see Fig. 2b). Results A total of 61 individual seeds were analyzed to reconstruct their reproductive pathways. According to standard procedures, 30 seeds were suspended in sheath fluid, while 31 were suspended in ArUPH2O (conductivity: 0.055 μS/cm or 18.2 MΩ·cm at 25°C). On average, 2,329 nuclei were tested per sample, accounting for 73% of total counted particles. The remaining 27% represented background signals or G2 phase nuclei. For all seeds, the average peak position of embryos and endosperms was calculated based on the total enrichment of each peak. Discussion The test results showed minimal differences, confirming that the arium system is an efficient and cost-effective alternative for ploidy analysis in plant materials. High-quality ArUPH2O proved effective in achieving reproducible results in lost cell seed screening techniques. However, additives like antibiotics and detergents are often recommended to prevent biofilm formation and maintain system integrity. While these substances may have long-term or short-term effects, they are commonly added to homemade sheath fluids. In summary, ArUPH2O is instantly available for flow cytometry in plant cells. As lost cell technology becomes more critical in applications like tumor cell detection, cell cycle analysis, DNA-RNA content estimation, and apoptosis detection, ArUPH2O offers new opportunities in emerging flow cytometry technologies. View full story References 1. Johri, BM Embryology of Angiosperms. Springer-Verlag: Berlin, Germany, 1984. 2. Nogler, GA Gametophytic Apomixis. In: Embryology of Angiosperms; Johri, BM, Ed. Springer-Verlag: Berlin, Germany; pp 475–518. 3. Battaglia, E. The male and female gametophytes of angiosperms—an interpretation. Phytomorphology 1951, 1, 87–116. 4. Asker, SE; Jerling, L. Apomixis in Plants. CRC Press: Boca Raton, FL, 1992. 5. Hojsgaard, DH; MartÃnez, EJ et al. Competition between meiotic and apomictic pathways during ovule and seed development results in clonality. New Phytologist 2013, 197, 336–47 (and supporting information in the online version of this article). 6. Dittrich, W.; G?hde, W. Patent DE 1815352, Flow-through Chamber for Photometers to Measure and Count Particles in a Dispersion Medium; 1968. 7. en.wikipedia.org/wiki/Flow_cytometry. 8. Heller, FO DNS-Bestimmung an Keimwurzeln von Vicia faba L. mit Hilfe der Impulscytophotometrie. (DNA estimation on radicles of Vicia faba L. using pulse cytophotometry; translation of the original German title by Dr. Herbig.) Berichte der Deutschen Botanischen Gesellschaft 1973, 86, 437–41. 9. Dolezel, J. Flow cytometric analysis of nuclear DNA content in higher plants. PhyTOChem. Anal. 1991, 2, 143–54. 10. Matzk, F.; Meister, A. et al. An efficient screen for the reproductive pathways using mature seeds of monocots and dicots. Plant J. 2000, 21, 97–108. 11. Paun, O.; H?randl, E. Evolution of hypervariable microsatellites in apomictic polyploid lineages of Ranunculus carpaticola: directional bias at dinucleotide loci. Genetics 2006, 174, 387–98. 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