Silica production - PDMPO
| Biogenic silica production (PDMPO) |
|---|
| Approach: fluorescent silica dye (PDMPO) incorporation |
| Context: incubation, in situ and lab |
| Spatial scale: point sample; single cell |
| Temporal scale: hours to days |
| Units: (x)mol Si cell-1 d-1 |
| Community captured: bulk, targeted size classes or taxa |
| Co-measurements: ambient [Si(OH)4], PAR, temperature, BSi, pH/pCO2 |
Method Overview
2-(4-pyridyl)-5-((4-(2-dimethylaminoethylaminocarbamoyl)methoxy)phenyl)oxazole (PDMPO) is a fluorescent dye that partitions specifically into sites of active silica deposition in diatoms and other silicifying organisms. During incubation, PDMPO is added to the sample and becomes incorporated into newly forming frustule silica, where it fluoresces strongly (excitation ~415 nm, emission ~480 nm) due to the acidic environment of the silica deposition vesicle. The fluorescence intensity, measured by flow cytometry or confocal microscopy, is proportional to the amount of silica newly deposited during the incubation. Calibration against a fluorescence standard derived from uniformly PDMPO-labelled diatoms converts fluorescence to Si production rates[1].
At the single-cell level, confocal microscopy resolves per-cell silica production rates and can identify individual silicifying cells within mixed communities.
Scale of measurement
Point samples requiring incubations of hours to days. At the single-cell level, production rates are resolved per individual cell.
Data generated
Per-cell and bulk BSi production rates (x)mol Si cell-1 d-1. Can distinguish actively silicifying cells from non-silicifying cells in a community.
Units & currency
Units are (x)mol Si cell-1 d-1. The currency is Si via calibrated fluorescence.
Sample size
Typical samples are ~60 mL in volume.
Repositories & databases
Limitations
The BSi:PDMPO molar conversion ratio (established at ~2916 mol BSi per mol PDMPO) must be verified for each study, as it can vary between diatom taxa and measurement conditions. PDMPO fluorescence decreases over time in stored samples, potentially reducing quantitative comparability across sample collections. Calibration requires a uniformly labelled standard, which is difficult to prepare consistently.
Example Applications & Protocols
Classic examples
- Shimizu et al. (2001) A novel fluorescent silica tracer for biological silicification studies [1]
- Leblanc & Hutchins (2005) New applications of a biogenic silica deposition fluorophore in the study of oceanic diatoms [2]
- McNair et al. (2015) Quantifying diatom silicification with the fluorescent dye PDMPO [3]
Recent applications
- Maniscalco et al. (2022) Diminished carbon and nitrate assimilation drive changes in diatom elemental stoichiometry independent of silicification in an iron-limited assemblage [4]
Common calculations/conversions
- ρSi (fmol Si cell-1 h-1) = (RFUcell / RFUstandard) × Sistandard / incubation time; where Sistandard is the known Si content of the labelled diatom standard.
References
- ↑ 1.0 1.1 Shimizu, K., Del Amo, Y., Brzezinski, M. A., Stucky, G. D., & Morse, D. E. (2001). A novel fluorescent silica tracer for biological silicification studies. Chemistry & Biology, 8(11), 1051–1060. https://doi.org/10.1016/S1074-5521(01)00072-2
- ↑ Leblanc, K., & Hutchins, D. A. (2005). New applications of a biogenic silica deposition fluorophore in the study of oceanic diatoms. Limnology and Oceanography: Methods, 3, 462–476. https://doi.org/10.4319/lom.2005.3.462
- ↑ McNair, H. M., Brzezinski, M. A., & Krause, J. W. (2015). Quantifying diatom silicification with the fluorescent dye PDMPO. Limnology and Oceanography: Methods, 13, 587–599. https://doi.org/10.1002/lom3.10049
- ↑ Maniscalco, M., McNair, H. M., Brzezinski, M. A., & Krause, J. W. (2022). Diminished carbon and nitrate assimilation drive changes in diatom elemental stoichiometry independent of silicification in an iron-limited assemblage. ISME Communications, 2, 6. https://doi.org/10.1038/s43705-022-00136-1