APPLICATION NOTE NO. 62
Calculating Calibration Coefficients for WET Labs ECO-AFL and ECO-FL Fluorometer, ECO-NTU Turbidity Meter, and ECO-FL-NTU Fluorometer/Turbidity Meter (voltage or RS-232 output sensor)
Revised May 2011

Print version
(Note: This application note is for use with Seasave and SBE Data Processing version 7.21d and later. An earlier version of the application note is compatible with earlier software versions.)

This Application Note applies to the following WET Labs fluorometers and turbidity meters:

• ECO-AFL fluorometer -- older model, not in current production by WET Labs
• ECO-FL fluorometer -- FL, FLD, FL(RT), FL(RT)D, FLS, FLB, or FLSB
• ECO-NTU turbidity meter -- NTU(RT), NTU(RT)D, or NTUS
• ECO-FL-NTU -- combines a fluorometer and turbidity meter, and requires two channels in your CTD

Note: The ECO-FL is also available as part of the WET Labs Triplet, an RS-232 output instrument that incorporates three WET Labs ECO sensors. The Triplet can include up to three ECO-FL or ECO-NTU sensors (or one of those sensors in combination with other ECO sensors). Set up each RS-232 output channel with the coefficients from the calibration sheet for the sensor on that channel.

The fluorometer has a response that is linear over the measurement range provided.

• ECO-AFL (no longer current production) -- Measurement range is approximately 0.02 - 100.0 µg/l or 0.04 - 200 µg/l.
• ECO-FL -- Measurement range for the analog output version of this single channel sensor can be adjusted with the analog scaling value. To change the range, connect the ECO-FL directly to the computer and use WET Labs' ECOView Host software to enter a new analog scaling value.

Analog Scaling Value / Range (enter in ECOView Host)	1	2	4 (factory default)
Chlorophyll a: Nominal Range (µg/l)	0 - 30	0 - 50	0 - 125
Rhodamine: Nominal Range (ppb)	0 - 55	0 - 110	0 - 230
Phycocyanin: Nominal Range (ppb)	0 - 100	0 - 200	0 - 400
Phycoerythrin: Nominal Range (ppb)	0 - 55	0 - 110	0 - 230
CDOM: Nominal Range (ppb)	0 - 125	0 - 250	0 - 500
Uranine: Nominal Range (ppb)	0 - 100	0 - 200	0 - 400

Note: Measurement ranges specified by WET Labs have varied over the years; consult the documentation that was provided with your sensor for the appropriate ranges, or contact WET Labs.

• ECO-FL-NTU -- Measurement ranges for this combination fluorometer and turbidity meter is factory-configured, and cannot be adjusted in the field. For the fluorometer channel, the range is approximately 0 to 30, 50, 75, or 125 µg/l (see Turbidity Meter Calibration Coefficients below for the turbidity meter channel).
• ECO-FL as part of the Triplet ‑ When the ECO-FL is part of the Triplet, the measurement range is factory-configured and cannot be adjusted in the field. For the fluorometer channel, the ranges are:

Chlorophyll a: Nominal Range (µg/l)	0 - 50
Rhodamine: Nominal Range (ppb)	0 - 175
Phycocyanin: Nominal Range (ppb)	0 - 175
Phycoerythrin: Nominal Range (ppb)	0 -175
CDOM: Nominal Range (ppb)	0 - 375
Uranine: Nominal Range (ppb)	0 - 300

Setting Up Configuration (.xmlcon) File for Fluorometer

1. Use the Configure Inputs menu in Seasave V7 (real-time data acquisition software), or the Configure menu in SBE Data Processing (post-processing software), to create / modify the .xmlcon file (see software Help files).

• For the voltage (analog) output ECO FL: Select Fluorometer - WET Labs CDOM for the CDOM fluorometer or Fluorometer - WET Labs ECO-AFL/FL for all other fluorometers for one of the external voltage channels on the CTD.
• For the RS-232 (digital) output ECO FL (SBE 16plus V2, 16plus-IM V2, or 19plus V2 CTDs only): Select Fluorometer - WET Labs CDOM for the CDOM fluorometer or Fluorometer - WET Labs ECO-AFL/FL for all other fluorometers for one of the RS-232 channels on the CTD.

2. The software prompts for Dark Output and Scale Factor, and calculates:

concentration (µg/l or ppb, as applicable) = (Output - Dark Output) * Scale Factor     (see Note)

where:

• Output (volts for analog output sensor; counts for digital output sensor) = in-situ output of the fluorometer
• Dark Output (volts for analog output sensor; counts for digital output sensor) = measured output for a seawater blank (pure, de-ionized water) with black tape over detector     (see Note)
• Scale Factor (µg/l-volts, µg/l-counts, ppb/volts, or ppb/counts, as applicable) = multiplier
• The fluorometer comes with a calibration sheet that lists values for Vblank and Scale Factor (see Note). If you changed the analog scaling value (ECO-FL series only), change the Scale Factor to correspond.

The fluorometer comes with a calibration sheet that lists values for Dark Output and Scale Factor (see Note). Each of these values is supplied in terms of both voltage and counts. If you changed the analog scaling value (ECO-FL series only), change the Scale Factor to correspond.

Notes:

• The configuration file can only be saved as an .xmlcon file (not a .con file) if the digital output version of the ECO-FL is one of the sensors.
• Calibration sheets from WET Labs for older fluorometers may list Vblank instead of Dark Output. Use the Vblank value in place of Dark Output when setting up the configuration file.

Example Chlorophyll a Concentration Calculation in Sea-Bird Software:

Dark Output = 0.05 volts and Scale Factor = 12.35 µg/l-volts (from calibration sheet)

Measured voltage from fluorometer = Output = 4.65 volts

Calculated concentration (µg/l) = (Output - Dark Output) * Scale Factor
                                             = (4.65 - 0.05) * 12.35 = 56.8 µg/l

While the factory-supplied Scale Factor can be used to obtain approximate values, field calibration is highly recommended.

• For example, the relationship between fluorescence and chlorophyll a is highly variable, and is not easy to determine in the laboratory. Species distribution, ambient light level, and health of the stock are just some of the factors that affect the relationship. To accurately measure chlorophyll a concentration with a fluorometer, perform calibrations on seawater samples with concentrations of plankton populations that are similar to what is expected in-situ. Determine chlorophyll-a concentrations independently, and use those concentrations, as well as readings from the fluorometer, to determine the correct Scale Factor. The Scale Factor is correct as long as the condition of the plankton population does not change; the condition does change with season and geographic location.

Example Calculation of Scale Factor from field calibration:

Seawater sample analysis shows chlorophyll a concentration is 50 µg/l when fluorometer reads 3.2 volts; measured signal for seawater blank is 0.05 volts.

concentration (µg/l) = (Output - Dark Output) * Scale Factor
50 = (3.2 - 0.05) * Scale Factor

Solving: Scale Factor = (50) / (3.2 - 0.05) = 15.87 µg/l

Enter new Scale Factor in configuration (.xmlcon) file.

 

The turbidity meter has a response that is linear over the measurement range provided.

• ECO-NTU -- Measurement range for the analog output version of this single-channel sensor can be adjusted with the analog scaling value. To change the range, connect the ECO-NTU directly to the computer and use WET Labs' ECOView Host software to enter a new analog scaling value.

Analog Scaling Value / Range (enter in ECOView Host)	1	2	4 (factory default)
Sea-Bird PN 24345, 24346, and 24348: Nominal Range (NTU)	0 - 30	0 - 60	0 - 125
Sea-Bird PN 24367: Nominal Range (NTU)	0 - 250	0 - 500	0 - 1000

• ECO-FL-NTU -- Measurement ranges for this combination fluorometer and turbidity meter is factory-configured, and cannot be adjusted in the field. For the turbidity meter channel, the range is approximately 0 to 10, 25, 100, 200, or 1000 NTU (see Fluorometer Calibration Coefficients above for the fluorometer channel).
• ECO-NTU as part of the Triplet -- When the ECO-NTU is part of the Triplet, the measurement range is factory-configured and cannot be adjusted in the field. For the turbidity meter channel, the range is 0 to 10, 25, 300, or 1000 NTU.

Note: If you require m ^-1 sr ^-1 output, WET Labs can provide a secondary calibration for these instruments in m ^-1 sr ^-1 at an additional cost. Alternatively, you can purchase the ECO-BB, which comes calibrated to m ^-1 sr ^-1. See Application Note 87 for details on how to use m ^-1 sr ^-1 output with Sea-Bird CTDs and software.

Setting Up Configuration (.xmlcon) File for Turbidity Meter

1. Use the Configure Inputs menu in Seasave V7 (real-time data acquisition software), or the Configure menu in SBE Data Processing (post-processing software), to create / modify the .xmlcon file (see software Help files).

• For the voltage (analog) output ECO-NTU: Select OBS/Nephelometer/Turbidity – Turbidity, WET Labs, ECO-NTU for one of the external voltage channels on the CTD.
• For the RS-232 (digital) output ECO-NTU (SBE 16plus V2, 16plus-IM V2, or 19plus V2 CTDs only): Select OBS/Nephelometer/Turbidity – Turbidity, WET Labs, ECO-NTU for one of the RS-232 channels on the CTD.

2. The software prompts for Dark Output and Scale Factor, and calculates:

Turbidity (NTU) = (Output - Dark Output) * Scale Factor     (see Note)

where:

• Output (volts for analog output sensor; counts for digital output sensor) = in-situ output of the turbidity meter
• Dark Output (volts for analog output sensor; counts for digital output sensor) = measured output for a seawater blank (pure, de-ionized water) with black tape over detector
• Scale Factor (NTU/volts or NTU/counts, as applicable) = multiplier

The turbidity meter comes with a calibration sheet that lists values for Dark Output and Scale Factor. Each of these values is supplied in terms of both voltage and counts. If you changed the analog scaling value (single-channel ECO-NTU only), change the Scale Factor to correspond.

Note: The configuration file can only be saved as an .xmlcon file (not a .con file) if the digital output version of the ECO-NTU is one of the sensors.

While the factory-supplied Scale Factor can be used to obtain approximate values, field calibration is highly recommended. The relationship between turbidity and NTU is highly variable, and is not easy to determine in the laboratory. Particle shape and size are some of the factors that affect the relationship. To accurately measure NTU with a turbidity meter, perform calibrations on seawater samples with distributions of particles that are similar to what is expected in-situ. Determine NTU independently, and use those values, as well as readings from the turbidity meter, to determine the correct Scale Factor. The Scale Factor is correct as long as the distribution of particle sizes and shapes does not change; the condition does change with season and geographic location.

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