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. 2014 Oct 28;9(4):1003–1013. doi: 10.1038/ismej.2014.199

Table 2. Criteria guiding data collection.

Data selection criterion Motivation and explanatory notes
Fe status of cells—only iron-limited cells We specifically probe the Fe uptake systems induced under iron limitation. This is environmentally relevant as Fe limitation is widespread across aquatic habitats.
Fe substrates—preferably chemically defined substrates, where only one Fe species is available for uptake To derive the uptake rate constant (equation 2), Fe substrate concentration and speciation must be known. Most selected experiments are of well-defined substrates such as Fe′ or strongly complexed Fe (siderophore bound). We also included less-defined substrates such as polysaccharides and colloidal Fe and discuss the limitations in their analysis.
Fe substrate concentrations—sub-saturating with respect to cellular Fe uptake sites Calculation of the uptake rate constant from uptake rates (equation (2)) is valid only for Fe concentrations below those that result in maximal uptake rate (that is, <Vmax). See Section 1 of Supplementary Appendix for more details
Growth phase—only exponentially growing cells Uniformity of phytoplankton physiological status in all experiments is extremely important. Phytoplankton physiology during stationary or lag phase may deviate significantly from that in exponential phase.
Cell density—low cell density to avoid changes in Fe concentration and speciation during the uptake experiments Quasi-equilibrium should be maintained between cells and Fe substrate. High cell density results in underestimation of uptake rate due to competition between cells over iron. We chose experiments where cells took less than 20% of the total Fe, or that the total uptake rate was at most 20% of the FeEDTA dissociation rate.
Extracellular Fe washed before measurements Iron tends to stick to cell surfaces and a wash (for example, titanium–EDTA wash or oxalate wash) is essential to prevent overestimation of intracellular iron concentration.
Minimal excess free ligand (for siderophore-bound Fe uptake). Free excess ligand can inhibit iron uptake rates due to interference with uptake mechanisms (Shaked et al., 2005). Therefore, uptake experiments with strong Fe complexes (not including EDTA) with minimal excess ligand were selected (Fe:L<1:5)

These criteria apply to both steady-state and short-term Fe uptake data.