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- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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| 2. |
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| 3. |
- Kubler-Kielb, Joanna, et al.
(författare)
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Oligosaccharide conjugates of Bordetella pertussis and bronchiseptica induce bactericidal antibodies, an addition to pertussis vaccine.
- 2011
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 1091-6490. ; 108:10, s. 4087-92
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Tidskriftsartikel (refereegranskat)abstract
- Pertussis is a highly contagious respiratory disease that is especially dangerous for infants and children. Despite mass vaccination, reported pertussis cases have increased in the United States and other parts of the world, probably because of increased awareness, improved diagnostic means, and waning vaccine-induced immunity among adolescents and adults. Licensed vaccines do not kill the organism directly; the addition of a component inducing bactericidal antibodies would improve vaccine efficacy. We investigated Bordetella pertussis and Bordetella bronchiseptica LPS-derived core oligosaccharide (OS) protein conjugates for their immunogenicity in mice. B. pertussis and B. bronchiseptica core OS were bound to aminooxylated BSA via their terminal Kdo residues. The two conjugates induced similar anti-B. pertussis LPS IgG levels in mice. B. bronchiseptica was investigated because it is easier to grow than B. pertussis. Using B. bronchiseptica genetically modified strains deficient in the O-specific polysaccharide, we isolated fractions of core OS with one to five repeats of the terminal trisaccharide, having at the nonreducing end a GlcNAc or GalNAc, and bound them to BSA at different densities. The highest antibody levels in mice were elicited by conjugates containing an average of 8-17 OS chains per protein and with one repeat of the terminal trisaccharide. Conjugate-induced antisera were bactericidal against B. pertussis, and the titers correlated with ELISA-measured antibody levels (r = 0.74). Such conjugates are easy to prepare and standardize; added to a recombinant pertussis toxoid, they may induce antibacterial and antitoxin immunity.
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