Sensitive staining of PHA by BODIPY 493/503 [78], this probe was apparently not
Sensitive staining of PHA by BODIPY 493/503 [78], this probe was apparently not used for sorting. Several studies have been published on the sorting of plant protoplasts, e.g. for alkaloid production [e.g. [79]]. For more details the reader is referred to references in [80]. In some cases quite unspecific staining has been successfully used for sorting of overproducers. One example is gramicidin S production by Bacillus brevis, based on the observation that gramicidin S overproducing cells gave higher fluorescence signals after fluorescein-isothiocyanate (FITC) staining as compared to low or non producers. By sorting for high FITC fluorescence these authors were able to isolate a strain with appr. twofold morePage 8 of(page number not for citation purposes)Microbial Cell Factories 2006, 5:http://www.microbialcellfactories.com/content/5/1/gramicidin S production [81]. It should be noted that the potentials of sorting based on autofluorescence or unspecific staining are extensive, hence it is not the intention of this review to list every single application available in the literature.Competing interestsThe author(s) declare that they have no competing interests.Authors’ contributionsBoth authors contributed equally to this manuscript.ConclusionWhile cell sorting is still mainly applied for clinical purposes, there is an increasing interest in biotechnology to utilize its potential for library screening or strain development. The aim of this review was to structure the fields of cell sorting applications in biotechnology, and to highlight many of the examples published over the last PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/28381880 years. It turns out that the major fields of use are protein engineering and screening for protein overproduction, both with microbial and animal cells. However, the potential, especially for non-protein products, is still by far not fully utilized. To design a novel sorting strategy for the improvement of a product or a production strain, the development of a suitable analytical method describing the desired properties adequately is most critical. The full potential of flow BIM-22493 msds cytometry methods, based on autofluorescence, physiological and metabolite specific probes, immunofluorescence, etc. can be utilized for this purpose. After identifying cell populations with superior properties, these can be sorted by FACS, thus directly employing the flow cytometry protocol on a preparative scale. The multiparameter analysis option of flow cytometry is an especially appealing feature, as it enables the simultaneous screening for overproduction and desirable cellular properties like robustness under production conditions. The general screening rule “you get what you screen for” is especially true for cell sorting, which emphasizes the importance of development and design of analytical methods. By closing the loop back to molecular physiological characterization of sorted strains and to strain development, one can elegantly perform “inverse metabolic engineering” [82]. Gaining understanding of the genetic background of sorted phenotypes enables novel rational approaches for cell and metabolic engineering which may further improve the performance of production strains. Finally, one may envisage a further refinement of systems biology. At present, most physiological data fed into models were obtained as average values of cell populations. As it becomes more and more obvious that clonal cultures also evolve significant heterogeneities, one can postulate a significant role of f.
