AN41: Calculating Calibration Coefficients for WET Labs WETStar Fluorometer
This Application Note applies to the use of the WET Labs WETStar fluorometer with Sea-Bird CTDs. The WETStar fluorometer is factory configured to measure Chlorophyll-a, CDOM, Uranine, Rhodamine, or Phycoerythrin concentration, and provides a linear output. See the WET Labs documentation provided with your WETStar for your instrument range.
Note: The WETStar comes with a calibration sheet that lists values for Clean Water Offset and Scale Factor. Each of these values is supplied in terms of both voltage and counts.
- Use the voltage values in the Sea-Bird software if using a voltage output WETStar.
- Use the counts values in the Sea-Bird software if using an RS-232 output WETStar (compatible only with
SBE 16plus V2, 16plus-IM V2, or 19plus V2 CTDs).
Setting up Configuration (.xmlcon) File
- 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 WETStar: Select Fluorometer – WET Labs WETStar for one of the external voltage channels on the CTD.
- For the RS-232 (digital) output WETStar (SBE 16plus V2, 16plus-IM V2, or 19plus V2 CTDs only): Select Fluorometer – WET Labs WETStar for one of the RS-232 channels on the CTD.
- The software prompts for Blank Output and Scale Factor, and calculates:
Concentration (μg/l or ppb, as applicable) = (Output – Blank Output) * Scale Factor
- Output (volts for analog output sensor; counts for digital output sensor) = in situ output of the fluorometer
- Blank Output (volts for analog output sensor; counts for digital output sensor) = measured output for a seawater blank (pure, de-ionized water); called Clean Water Offset on WETStar calibration sheet
- Scale Factor (μg/l – volt, μg/l – count, ppb/volt, or ppb/count, as applicable) = multiplier
Note: The configuration file can only be saved as an .xmlcon file (not a .con file) if the RS-232 output WETStar is one of the sensors.
Example Concentration Calculation in Sea-Bird Software:
Blank Output = 0.05 volts and Scale Factor = 14.10 µg/l-volts (from calibration sheet)
Measured voltage from WETStar= Vsample = 4.65 volts
Concentration = (Output – Blank Output) * Scale Factor = (4.65 - 0.05) * 14.10 = 64.9 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. It is only through the use of these calibrations that a meaningful and accurate measure of chlorophyll-a can be obtained. 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 indicates chlorophyll-a concentration is 50 µg/l when fluorometer reads 3.2 volts; measured signal for a sea water blank is 0.05 volts.
concentration = (Output – Blank Output) * Scale Factor
50 = (3.2 - 0.05) * Scale Factor
Solving for Scale Factor:
Scale Factor = (50) / (3.2 - 0.05) = 15.87 µg/l
Enter the new Scale Factor in the configuration (.xmlcon) file.
Application Note Revision History
|March 2003||Reflect new calibration procedure used by WET Labs, which now provides Vblank and Scale Factor on calibration sheet.|
|May 2007||Incorporate Seasave V7.|
|February 2010||- Change Seasoft-Win32 to Seasoft V2.
- Add information on .xmlcon file.
- Update address.
|May 2011||SBE Data Processing and Seasave 7.21d software revision: WET Labs RS-232 output sensors, including Triplet, are compatible with RS-232 sensor channel on SBE 16plus V2, 16plus-IM V2, and 19plus V2.|