Paradigm multi mode detection platform

In order to obtain growth characteristics of P. angustum S14 in batch mode, and in conditions equivalent to those used in the continuous culture experiment, we performed duplicate growth curves in the light (406.7 μE · m^–2 · s^–1) and in the dark using the photobioreactors, the “high” substrate medium and the same parameters used in the continuous culture experiments. Growth was estimated from OD_{620 nm} measurements as above.
The optimum pH for growth was tested in duplicate using the “high” substrate medium to which buffers MES (Sigma-Aldrich; for pH 5.5–6.5), HEPES (Sigma-Aldrich; for pH 7.0–8.0), or AMPSO (Sigma-Aldrich; for pH 8.5–10.0) were added at 2.0 mg/ml. pH was adjusted to a 5.5–10.0 range using 25% (v/v) concentrated HCl or 1 N NaOH. Growth was followed in 24 well plates (Evergreen Scientific, USA) incubated at 25°C in the dark at 100 rpm. OD_{620 nm} measurements were obtained using a Paradigm™ Multi-mode Detection Platform (Molecular Devices, USA).

Triplicate 1 ml samples were fixed with 1% glutaraldehyde (Sigma-Aldrich, USA), flash frozen in liquid nitrogen and stored at –80°C. Samples were diluted 100-fold and stained for 15 min with a 1:400 dilution of SYBR green I (Molecular Probes, USA). Cell counts were measured in triplicate by flow cytometry using a FACSCalibur flow cytometer (Becton Dickinson, USA) equipped with an air-cooled argon laser (488 nm, 15 mW) as previously described (Van Wambeke et al., 2011 (link)).
Final OD_{620 nm} measurements (shown in all figures, except for Figure 1) were taken in triplicate in 96 well microplates from the same fixed samples using a Paradigm™ Multi-mode Detection Platform (Molecular Devices).