Data Overview:

A collection of fluorescence data collected using the phosphite (Phi) assay method described in "A Fluorometric Assay for High-throughput Phosphite Quantitation in Biological and Environmental Matrices". Excel files contain the fluorescence output data from the Spark 10m microplane reader (Tecan), unless otherwise noted, including the instrument's operating conditions, and the organization of fluorescence readouts by sample ID. Fluorescence values are averaged in triplicate, then the sample containing no 17X-PTDH is subtracted from all average values. Every sample has at least 3 points on a standard addition curve ranging from 0-5 nmol of spiked Phi (excluding example assays which have a wider range). Red boxes with "OVER" were samples that saturated the detector and were not used for any analysis.

Other supplied data include absorbance measurements collected to determine 17X-PTDH and LDH* activity.

Files

1. 3-1-22_LOD_3_1x_and_0x_PMS_annot.xlsx
2. 3-11-22_17X-PTDH_Activity_and_concentration.xlsx
3. LDH_activity_in_STET_Annot.xlsx
4. 9-23-21_E.coli_LB_LOD_1.xlsx
5. 11-16-21_e.coli_LB_LOD_2_and_3.xlsx
6. 12-2-21_Seawater_LOD_1_and_Recovery_Annot.xlsx
7. 11_9_21_Seawater_LOD_2_and_3_Annot.xlsx
8. 4-26-22_n.maritimus_pellet_LOD1_Annot.xlsx
9. 4-24-22_n._maritimus_LOD_2_Annot.xlsx
10. 3-13-22_pt_assay_kinetics_fig_2_annot.xlsx
11. 3-6-22_LDH_E._coli_comparison_in_LB_and_Ecoli_LOD_in_LB.xlsx
12. 3-4-22_Example_Std_Adn_and_LOD_1_annot.xlsx
13. 2-28-22_1x_PMS_and_LOD_2_annot.xlsx
14. 3-8-22_50__seawater_recovery_annot.xlsx
15. 2-16-22_n._maritimus_LOD_3_Annot.xlsx
16. 2-9-22__Radish_leaf_LOD_1_annot.xlsx
17. 1-26-22_Ecoli_LDH_comparison_in_MOPS_and_LOD_Annot.xlsx
18. 1-19-22_Artificial_seawater_Annot.xlsx
19. 1-14-22_Anaerobic_digester_LODs_and_Recovery_Annot.xlsx
20. 8-30-22_MS_radish_pt_assays_annot.xlsx
21. 4-1-22_radishes_annot.xlsx
22. 3-29-22_radish_data_annot.xlsx
23. 3-30-22_radish_analysis_annot.xlsx
24. 3-13-22_radish_leaves_and_stems_Pt_recovery_annot.xlsx
25. 2-8-22_2_root_extract_LOD_annot.xlsx

Data-Specific Information

3-1-22_LOD_3_1x_and_0x_PMS_annot.xlsx

This fluorescence data is for two experiments of the Phi assay: the first portion of the plate (A1-C3) shows the Phi assay prepared as described in Methods, using 100 uM PMS; the second portion of the plate (D1-F3) shows the Phi assay prepared with no PMS. Instrument parameters are detailed in the first 57 lines, then an initial reading (t = 0) was taken of the samples. Then the kinetic experiment begins, and the instrument parameters are detailed in lines 68-96.

Below the instrument acquisition, averages and standard deviation (lines 133-164) are taken for the triplicate measurements of the standard addition curve as well as the PTDH-free blank.

Finally, to determine the LOD of the assay, the final time point was used to generate a standard addition curve (lines 175-184).

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column (and row, for the t= 0 measurement) representing the plate's well location. In the averaged data (133-164), the header is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.
• Time [s] - time since the instrument began acquisition on the kinetics experiment.
• Cycle Nr. - Number of times the instrument has taken a measurement since the beginning of the kinetics experiment.

3-11-22_17X-PTDH_Activity_and_concentration.xlsx

This data sheet shows three UV-Vis absorbance experiments taken on a Cary 60 from purified 17X-PTDH in order to determine its activity . The reaction mixture contained 1.2 ug protein and 3 mL of assay mix (100 mM MOPS, pH 7.25, 10 mM NAD+, and 10 mM Phi).

Columns A and B show the absorbance values at 340 nm taken in a kinetics experiment with the above reaction mixture stirred at 25C. Columns H and I show the absorbance values at 340 nm taken in a kinetics experiment with the above reaction mixture stirred at 37C. Columns Q and R show the absorbance at wavelengths of 500 to 200 nm. Instrumental parameters are reported at the bottom of the reported absorbance values for each column.

Variables:

• Abs [A.U.]- Absorbance values reported by the instrument at the given wavelength of incident light.
• Time [min] - Minutes since data acquisition began.
• Wavelength [nm] - Wavelength of light incident on the sample.

LDH_activity_in_STET_Annot.xlsx

This data sheet shows two UV-Vis absorbance experiments taken on a Cary 60 from purified LDH* in order to determine its activity. The reaction mixture contained 1 ug protein and 3 mL of assay mix (100 mM NaCl, 10 mM Tris-HCl, pH 8.0, 1 mM EDTA, 5% v/v Triton X-100, 0.5 mM NADH, 30 mM pyruvate).

Columns A and B show the absorbance values at 340 nm taken in a kinetics experiment with the above reaction mixture stirred at 25C. Columns C and D show the absorbance values at 340 nm taken in a kinetics experiment with the above reaction mixture stirred at 25C, with the purified enzyme undergoing 1x freeze/thaw. Column E shows the absorbance values in column D, adjusted so that Abs @ t = 0 is 0, for plotting purposes.

Variables:

• Abs [A.U.]- Absorbance values reported by the instrument at the given wavelength of incident light.
• Time [min] - Minutes since data acquisition began.

9-23-21_E.coli_LB_LOD_1.xlsx

This fluorescence data shows an experiment to determine the LOD of the Phi assay on E. coli lysate grown in LB without LDH treatment. This fluorescence data was taken on a Duetta Fluorescence and Absorbance Spectrometer (Horiba). The excitation wavelength was 535 nm (5 nm bandpass) and the emission range was 550 nm to 700 nm (5 nm bandpass), and the integration time was 0.1 s. Full wavelength scans are shown for each sample, and are averaged under the column indicating the sample name based on the amount of Phi spiked (e.g. 3 nmol). In columns CE-CJ, averaged fluorescence values at 585 nm are assembled and labeled by the amount of Phi spiked for clarity. The propagated error, slope of the standard addition line, and LOD of the sample standard addition curve is shown here.

Variables:

• Wavelength [nm] - Wavelength of light incident on the sample.
• Fluorescence [A.U.] - fluorescence values are reported by the instrument for a given sample.
• Smoothed Fluorescence [A.U.] - fluorescence values in the column to the left at a 5 nm rolling average to smooth the data according to the bandpass.

11-16-21_e.coli_LB_LOD_2_and_3.xlsx

This fluorescence data shows two experiments to determine the LOD of the Phi assay on E. coli lysate grown in LB without LDH treatment. These two runs represent two biological replicates run under the same conditions. This fluorescence data was taken on a Genios Pro 96/384 Fluorescence Absorbance Microplate Reader (Tecan). The excitation wavelength was 535 nm (5 nm bandpass) and the emission wavelength collected was 585 nm (5 nm bandpass), and the integration time was 0.1 s. The values of fluorescence were recorded from the instrument directly to the provided data sheet. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Average" and the PTDH-free blank is subtracted under "Adjusted", and the standard deviation is shown under "Std Dev". The propagated error, slope of the standard addition line, and LOD of the sample standard addition curve is shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument for a given sample.

12-2-21_Seawater_LOD_1_and_Recovery_Annot.xlsx

This fluorescence data shows two experiments to determine the LOD and Phi recovery of seawater in the Phi assay. Seawater from the SB Channel was coarsely filtered and spiked with 100 uM Phi prior to analysis. The instrument parameters are reported in lines 1-41. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Average" and the PTDH-free blank is subtracted under "Adjusted", and the standard deviation is shown under "Std Dev". The % recovery calculation, propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data the "nmol Phi added" is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

11_9_21_Seawater_LOD_2_and_3_Annot.xlsx

This fluorescence data shows two experiments to determine the LOD of seawater in the Phi assay. Seawater from the SB Channel was coarsely filtered and spiked with 100 uM Phi prior to analysis. The instrument parameters are reported in lines 1-41. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Average" and the PTDH-free blank is subtracted under "Adjusted", and the standard deviation is shown under "Std Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data the "nmol Phi added" is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

4-26-22_n.maritimus_pellet_LOD1_Annot.xlsx

This fluorescence data shows an experiment to determine the LOD of concentrated N. maritimus culture in the Phi assay. The culture was grown as described in Methods, and was concentrated 1000x using tangential flow. To lyse the cells, the samples were subjected to three freeze/thaw cycles. The instrument parameters are reported in lines 1-45. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Average" and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "Std Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data (133-164), the "nmol Phi added" is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

4-24-22_n._maritimus_LOD_2_Annot.xlsx

This fluorescence data shows an experiment to determine the LOD of concentrated N. maritimus culture in the Phi assay. The culture was grown as described in Methods, and was concentrated 1000x using tangential flow. To lyse the cells, the samples were subjected to three freeze/thaw cycles. The instrument parameters are reported in lines 1-45. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Average" and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "Std Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data the "nmol Phi spiked" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

3-13-22_pt_assay_kinetics_fig_2_annot.xlsx

This fluorescence data is to determine the kinetics of resorufin fluorescence in the Phi assay prepared as described in Methods, using 1 ug 17X-PTDH and 100 uM PMS. Instrument parameters are detailed in the first 57 lines, then an initial reading (t = 0) was taken of the samples. Then the kinetic experiment begins, and the instrument parameters are detailed in lines 65-93. Below the instrument acquisition, averages and standard deviation (lines 130-154) are taken for the triplicate measurements of the standard addition curve as well as the PTDH-free blank. Outlier fluorescence values in F2, F3, and E10 were removed for clarity (an occasional problem in kinetics experiments due to the misalignment of the plate and taking only one measurement per well) these conditions were excluded from the analysis and the figure presented in the paper.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column (and row, for the t= 0 measurement) representing the plate's well location. In the averaged data (133-164), the header is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.
• Time [s] - time since the instrument began acquisition on the kinetics experiment.
• Cycle Nr. - Number of times the instrument has taken a measurement since the beginning of the kinetics experiment.

3-6-22_LDH_E._coli_comparison_in_LB_and_Ecoli_LOD_in_LB.xlsx

This fluorescence data shows an experiment to compare the effect of LDH* treatment on E. coli lysate. The culture was grown, split into two aliquots, and one was treated with LDH* and pyruvate as described in Methods. The instrument parameters are reported in lines 1-45. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Intensity" and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "Std Dev". The propagated error and LOD of the LDH*-treated sample's standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

3-4-22_Example_Std_Adn_and_LOD_1_annot.xlsx

This fluorescence data shows an experiment to demonstrate the standard addition curve and how it can be used to read the Phi content of a sample. In the first experiment, the buffer of the Phi assay mix was spiked with 100 uM Phi. In the second experiment, the Phi assay mix was prepared as described with MOPS buffer. The instrument parameters are reported in lines 1-45. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Intensity" and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "Std Dev". The propagated error and LOD of the standard addition curve is shown here, as well as the read Phi concentration by the standard addition curve.

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

2-28-22_1x_PMS_and_LOD_2_annot.xlsx

This fluorescence data is presented to show an example of the Phi assay run with assay buffer with 100 uM PMS, 2 ug 17X-PTDH, and spiked Phi to determine the kinetics of resorufin fluorescence over time. Instrument parameters are detailed in the first 57 lines, then an initial reading (t = 0) was taken of the samples. Then the kinetic experiment begins, and the instrument parameters are detailed in lines 65-93. Well B10 gave outlier fluorescence values for cycles 20-25 (an occasional problem in kinetics experiments due to the misalignment of the plate and taking only one measurement per well), and was excluded from the analysis and the figure presented in the paper.

Below the instrument acquisition, averages and standard deviation (lines 133-164) are taken for the triplicate measurements of the standard addition curve as well as the PTDH-free blank.

Finally, to determine the LOD of the assay, the final time point was used to generate a standard addition curve (lines 175-184).

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column (and row, for the t= 0 measurement) representing the plate's well location. In the averaged data (133-164), the header is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.
• Time [s] - time since the instrument began acquisition on the kinetics experiment.
• Cycle Nr. - Number of times the instrument has taken a measurement since the beginning of the kinetics experiment.

3-8-22_50__seawater_recovery_annot.xlsx

This fluorescence data shows an experiment to determine the Phi recovery of spiked seawater. The seawater was concentrated to 50% its original volume on a vacuum concentrator and was spiked with 190 uM Phi (9.5 nmol detected in 50 uL assay sample volume). The instrument parameters are reported in lines 1-45. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Fluor. Intensity" and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "Std Dev". The % recovery, propagated error, and LOD of the LDH*-treated sample's standard addition curve are also shown here. Wells with saturated fluorescence ("OVER") were not relevant to the experiment.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data the "Phi added" [nmol] column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

2-16-22_n._maritimus_LOD_3_Annot.xlsx

This fluorescence data shows an experiment to determine the LOD of concentrated N. maritimus culture in the Phi assay. The culture was grown as described in Methods, and was concentrated 1000x using tangential flow. To lyse the cells, the samples were subjected to three freeze/thaw cycles. The instrument parameters are reported in lines 1-41. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Average" and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "Std Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Pt added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

2-9-22__Radish_leaf_LOD_1_annot.xlsx

This fluorescence data shows an experiment to determine the LOD of radish leaf extract in the Phi assay. Radish leaves were lysed by a mortar and pestle cooled with LN2. Cells were extracted using STET buffer (4mL/g wet weight), then diluted 1:100 prior to the assay. The instrument parameters are reported in lines 1-41. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Average" and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "Std Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Pt added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

1-26-22_Ecoli_LDH_comparison_in_MOPS_and_LOD_Annot.xlsx

This fluorescence data shows an experiment to determine the LOD of the Phi assay on E. coli lysate grown in MOPS Minimal Medium with and without LDH treatment. Cell growth, lysate extraction, and LDH treatment are detailed in Methods. The instrument parameters are reported in lines 1-41. The two samples were run in two separate runs of acquisition, and are shown in two Excel sheets. Note that this was because of the gain adjustment, which allowed the fluorescence to be read in the linear range for both experiments. The instrument adjusted gain based on a well of the relevant experiment's sample with the highest nmol Phi spiked (4.5 nmol in this case). The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg" and the PTDH-free blank is subtracted under "Adj. Int.", and the standard deviation is shown under "Std Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Pt added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

1-19-22_Artificial_seawater_Annot.xlsx

This fluorescence data shows an experiment to determine the effect of concentration and individual presence of salts in artificial seawater on Phi assay performance. The formulation of artificial seawater is described in Methods. The instrument parameters are reported in lines 1-55. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Intensity" and the PTDH-free blank is subtracted under "Adj. Int.", and the standard deviation is shown under "Std Dev".

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

1-14-22_Anaerobic_digester_LODs_and_Recovery_Annot.xlsx

This fluorescence data shows an experiment to determine the LOD and % Phi recovery of anaerobic digester sludge. The sludge was stored anaerobically and filtered with a 0.22 um syringe filter as described in Methods. The instrument parameters are reported in lines 1-55. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Mean" and the PTDH-free blank is subtracted under "Adj. Int.", and the standard deviation is shown under "Std Dev". Fluorescence values that were not relevant to the experiment are highlighted in grey (Plate row A). The % recovery, propagated error, and LOD of the anaerobic digester sludge sample's standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Pt" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

8-30-22_MS_radish_pt_assays_annot.xlsx

This fluorescence data shows several experiments to determine the Phi content of radish seedlings grown with various P sources and the presence or absence of Phi-oxidizing P. stutzeri. Radish seedlings were grown in MS minimal plant media and lysed using a LN2-cooled mortar and pestle, as described in Methods. Cell contents were extracted using 4 mL STET buffer/g wet weight, and the extract was diluted 1:100 prior to assay. This experiment was run on a Tecan M200 Infinite Pro. The experimental parameters are detailed in lines 1-50. As detailed there, multiple reads were taken per well in a square pattern and averaged. This was the case for all samples, except well G4, where only one measurement is used for the well, due to its large difference from the other three measurements. The triplicate measurements are averaged under "Avg Intensity" and the PTDH-free blank is subtracted under "Adj. Int.", and the standard deviation is shown under "Std Dev".

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.
• Well - Well of the plate being measured.
• Mean [A.U.] - Average of the four fluorescence readings taken per well.
• StDev (instrument output, not in analysis) - Standard deviation of the four fluorescence readings taken per well.
• 0;1, 1;1, 1;0, 0;0 - Coordinates of the square pattern that the instrument is following to take the four fluorescence readings of the well.

4-1-22_radishes_annot.xlsx

This fluorescence data shows an experiment to determine the LOD of radish leaf extract in the Phi assay. Radish leaves were lysed by a mortar and pestle cooled with LN2. Cells were extracted using STET buffer (4mL/g wet weight), then diluted 1:100 prior to the assay. The instrument parameters are reported in lines 1-53. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Inten." and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "St Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

3-29-22_radish_data_annot.xlsx

This fluorescence data shows an experiment to determine the LOD of radish leaf and root extract in the Phi assay. Radish leaves and roots were lysed by a mortar and pestle cooled with LN2. Cells were extracted using STET buffer (4mL/g wet weight), then diluted 1:100 prior to the assay. The instrument parameters are reported in lines 1-53. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Fl. Int." and the PTDH-free blank is subtracted under "Adj Inten.", and the standard deviation is shown under "St Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

3-30-22_radish_analysis_annot.xlsx

This fluorescence data shows several experiments to determine the Phi content of radish seedlings grown with various P sources and the presence or absence of Phi-oxidizing P. stutzeri. Radish seedlings were grown in MS minimal plant media and lysed using a LN2-cooled mortar and pestle, as described in Methods. Cell contents were extracted using 4 mL STET buffer/g wet weight, and the extract was diluted 1:100 prior to assay. The instrument parameters are reported in lines 1-53. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Fl. Int." and the PTDH-free blank is subtracted under "Adj Inten.", and the standard deviation is shown under "St Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

3-13-22_radish_leaves_and_stems_Pt_recovery_annot.xlsx

This fluorescence data shows an experiment to determine the % recovery of radish leaf and root extract in the Phi assay. Radish leaves and roots were lysed by a mortar and pestle cooled with LN2. Cells were extracted using STET buffer (4mL/g wet weight), then diluted 1:100 prior to the assay. Samples were then spiked with 100 uM Phi (5 nmol detected in 50 uL assay sample volume). The instrument parameters are reported in lines 1-55. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Fluor. Int." and the PTDH-free blank is subtracted under "Adj Fluor. Inten.", and the standard deviation is shown under "St Dev". The propagated error, LOD, and % recovery of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.

2-8-22_2_root_extract_LOD_annot.xlsx

This fluorescence data shows an experiment to determine the LOD of radish leaf and root extract in the Phi assay. Radish leaves and roots were lysed by a mortar and pestle cooled with LN2. Cells were extracted using STET buffer (4mL/g wet weight), then diluted 1:100 prior to the assay. The instrument parameters are reported in lines 1-55. The fluorescence values for each well are placed in the row corresponding to the amount of Phi spiked for that sample. The triplicate measurements are averaged under "Avg Intensity" and the PTDH-free blank is subtracted under "Adj Intensity", and the standard deviation is shown under "St Dev". The propagated error and LOD of the sample standard addition curve are also shown here.

Variables:

• Fluorescence [A.U.] - fluorescence values are reported by the instrument under the column and row representing the plate's well location. In the averaged data, the "nmol Phi added" column is the amount of Phi spiked in each sample, to help identify what wells correspond to what samples.