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In situ flourometry to chlorophyll: calibration Mark Doubell & SARDI Oceanography team

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Page 1: µ } u Ç } Z o } } Z Ç o o W o ] ] } v - IMOSimos.org.au/fileadmin/user_upload/shared/ANMN/QC_Summit_2017/… · Title: Microsoft PowerPoint - Doubell_Fluorescence Calibration.pptx

In situ flourometry to chlorophyll: calibration

Mark Doubell & SARDI Oceanography team

Page 2: µ } u Ç } Z o } } Z Ç o o W o ] ] } v - IMOSimos.org.au/fileadmin/user_upload/shared/ANMN/QC_Summit_2017/… · Title: Microsoft PowerPoint - Doubell_Fluorescence Calibration.pptx

"Throughout the realm there shall be the same yard of the same size and it should be of iron chlorophyll a”

Assize of Measures (1196)Relevance:- Quality & standardization of data (product) collected by IMOS- Inferences for product uptake, delivery and use by wider community

Page 3: µ } u Ç } Z o } } Z Ç o o W o ] ] } v - IMOSimos.org.au/fileadmin/user_upload/shared/ANMN/QC_Summit_2017/… · Title: Microsoft PowerPoint - Doubell_Fluorescence Calibration.pptx

Chlorophyll a• Chl a provides a standard ‘proxy’ measure of phytoplankton

biomass (µg/L or mg/m3)

• Variety of applications;- ecology (e.g. spatial distribution patterns),- biophysical couplings, - primary production, - water quality and trophic status (e.g. ANZECC)

Seuront et al. 2007

Behrenfeld and Falkowski, 1997

ANZECC 2000

Page 4: µ } u Ç } Z o } } Z Ç o o W o ] ] } v - IMOSimos.org.au/fileadmin/user_upload/shared/ANMN/QC_Summit_2017/… · Title: Microsoft PowerPoint - Doubell_Fluorescence Calibration.pptx

In situ fluorometers• In situ fluorometers: many types on many platforms• Share common measurement principles:

- chl a is ubiquitous to phytoplankton and has fluorescent properties- intensity of emitted in vivo fluorescence is linearly proportional to [chl a] - detected intensity output is in raw analog (v) or digital (rfu) units

• Require calibration with a chl a standard to convert output into standard concentration units (µg/L or mg/m3)

Suggett et al. 2010

calibration

Page 5: µ } u Ç } Z o } } Z Ç o o W o ] ] } v - IMOSimos.org.au/fileadmin/user_upload/shared/ANMN/QC_Summit_2017/… · Title: Microsoft PowerPoint - Doubell_Fluorescence Calibration.pptx

(Simple) Calibration Procedure• Requires measurement of in vivo fluorescence and extracted [chl a]

for a dilution series of phytoplankton

- Chaetoecurus meulleri(diatom), log growth phase

- dark room @ 15oC- 2L triplicate samples ,

0.7 µm Whatman GFF, extracted in acetone

- [chl a] by flurometry, Synergy4 fluorometer, Sigma chl a standards

Kriston Bott @SARDI

Page 6: µ } u Ç } Z o } } Z Ç o o W o ] ] } v - IMOSimos.org.au/fileadmin/user_upload/shared/ANMN/QC_Summit_2017/… · Title: Microsoft PowerPoint - Doubell_Fluorescence Calibration.pptx

Calibration Results

Page 7: µ } u Ç } Z o } } Z Ç o o W o ] ] } v - IMOSimos.org.au/fileadmin/user_upload/shared/ANMN/QC_Summit_2017/… · Title: Microsoft PowerPoint - Doubell_Fluorescence Calibration.pptx

Calibration Results

Type SARDI CSIROCalibration standard Chl a Chl a Chl a Fluorescent dyeWetlab FLNTU 954 RFU Volts RFU VoltsDate of Cal 29/09/2016 10/09/2015DarkCount 56 0.071 81 0.070Scale Factor (SF) 0.012 10 0.011 41Type SARDI CSIROWetlab FLNTU 955 RFU Volts RFU VoltsDate of Cal 29/09/2016 10/09/2015DarkCount 55 0.065 73 0.066Scale Factor (SF) 0.012 10 0.011 45Type SARDI CSIROWetlab FLNTU 1173 RFU Volts RFU VoltsDate of Cal 29/09/2016 29/01/2014DarkCount 58 0.093 88 0.100Scale Factor (SF) 0.007 6 0.009 26Type SARDI CSIROWetlab FLNTU 1248 RFU Volts RFU VoltsDate of Cal 29/09/2016 6/11/2015DarkCount 49 0.072 67 0.073Scale Factor (SF) 0.012 10 0.009 38

25/07/2013

Factory

Factory

Factory

Factory

10/03/2008

10/03/2008

18/02/2010

Page 8: µ } u Ç } Z o } } Z Ç o o W o ] ] } v - IMOSimos.org.au/fileadmin/user_upload/shared/ANMN/QC_Summit_2017/… · Title: Microsoft PowerPoint - Doubell_Fluorescence Calibration.pptx

Recommendations• Objective: To calibrate in situ fluorometers

to provide a standard, traceable measure of [chl a].

1. Calibration of fluorometers with chl a from culture/s to convert raw output into standard concentration units (µg/L) - consensus on culture/s to be used (inter/national collaboration)

- facility with culture and analytical lab capabilities- calibration over ecologically/regionally relevant range of [chl a] (accuracy &

precision)

2. Establish relation between [chl a] from culture/s and fluorescent dye to optimize calibration procedure

3. Regional/local optimization of calibrated fluorometer measures against extracted [chl a] from water samples- consensus HPLC & or fluorometry based- further supported using pigments/microscopy/flow cytometry- will take time to build sufficient data set


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