A total of 3 × 105 HEK293 cells/well were plated into 96-well culture plate 24 h before bacterial inoculation. The HEK293 cells were cultivated in DMEM/high glucose medium, containing 10% FBS at 37 °C, 5% CO2. The medium was changed before inoculation of K. pneumoniae to the following media: (1) 90% DMEM/high glucose medium, containing 10% FBS; (2) 89.99% DMEM/high glucose medium, containing 10% FBS, 0.01% Bacto Proteose Peptone (BD Biosciences, Franklin Lakes, NJ, USA); (3) 49.99% DMEM/high glucose medium, containing 50% FBS, 0.01% Bacto Proteose Peptone (BD Biosciences, Franklin Lakes, NJ, USA). Cell-free media served as control. K. pneumoniae UHI 1090 (104 CFU/well) was inoculated, and the incubation was prolonged at 37 °C, 5% CO2. OD values were measured 24 h after inoculation at 620 nm. Each measurement was performed in six replicates.
Bacto proteose peptone
Manufactured by BD
Sourced in United States
Bacto Proteose Peptone is a general-purpose peptone used in the preparation of microbiological culture media. It serves as a source of nitrogen, carbon, sulfur, and other nutrients required for the growth of a wide range of microorganisms.
Automatically generated - may contain errors
4 protocols using bacto proteose peptone
1
Evaluating K. pneumoniae Growth in HEK293 Cells
2
Cultivation and Characterization of Tetrahymena pyriformis
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T. pyriformis was donated by O. Numata (University of Tsukuba, Japan). The cells were cultured in growth medium [1.2% (w/v) Bacto Proteose Peptone (Becton, Dickinson and Company), 0.6% (w/v) Paticase (Kyokuto), and 0.2% (w/v) Bacto Yeast Extract, (Becton, Dickinson and Company)] at room temperature (20° to 25 ° C) with aeration (e-AIR6000WB, GEX). Serial transfer of the cells was performed twice per week. Before observation, the cells in midlog phase were washed three times with observation solution [10 mM Mops/tris (pH 7.2), 1 mM KCl, 1 mM NaCl, and 1 mM CaCl2] (26 ) and equilibrated with observation solution for more than 1 hour before observation.
The shape of the cells used in the experiments was approximated as an ellipse, where the mean major length was 52.3 μm and the mean minor length was 24.8 μm. The mean swimming speed far from a wall without external flow was 281.4 μm/s.
The shape of the cells used in the experiments was approximated as an ellipse, where the mean major length was 52.3 μm and the mean minor length was 24.8 μm. The mean swimming speed far from a wall without external flow was 281.4 μm/s.
3
Culturing and Preparing Tetrahymena pyriformis
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T. pyriformis was kindly gifted by Osamu Numata, University of Tsukuba, Tsukuba, Japan. The cells were cultured in growth medium [1.2% (wt/vol) Bacto Proteose Peptone (Becton, Dickinson and Company), 0.6% (wt/vol) Paticase (Kyokuto), and 0.2% (wt/vol) Bacto Yeast Extract (Becton, Dickinson and Company)] at room temperature (20–25 °C) with aeration (e-AIR6000WB; GEX). Serial transfer of the cells was performed twice per week. Before observation, the cells in midlog phase were washed three times with observation solution [10 mM 3-(N-morpholino)propanesulfonic acid/Tris (pH 7.2), 1 mM KCl, 1 mM NaCl, and 1 mM CaCl2] (49 ) and equilibrated with observation solution for more than 1 h before observation.
4
Francisella Growth and Enumeration
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F. novicida (U112) CIP56.12 (Centre de Ressources Biologiques de l'Institut Pasteur, Paris, France) and F. philomiragia (CHUGA-FP47) were grown either on solid or in liquid media, as indicated. The solid medium corresponds to pre-made Polyvitex-enriched chocolate agar (PVX-CHA) plates (Ref#43109 BioMérieux, France) incubated at 37°C in a 5% CO2 incubator with humidified atmosphere and used both to revive glycerol stocks and for CFU counting. Liquid cultures were carried out in Modified Mueller-Hinton broth (MMH) at 37°C under shaking. MMH corresponds to Mueller Hinton Broth (Ref#275730 Grosseron) supplemented with NaCl (Sigma; 5 g/L), BactoProteose peptone (Ref#211693 BD Biosciences; 5 g/L), BactoTryptone (Ref#211705 BD Biosciences 5 g/L) and L-cysteine (Ref#C7477 Sigma; 1 g/L).
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