For expanded information on the AquaFluor, check out our user manual: Aquafluor Handheld Fluorometer
For a quick calibration video guideline on our older AquaFluor: https://www.youtube.com/watch?v=9MzKnrIM5Ow
The AquaFluor® can be calibrated using Primary standards. A Primary Standard contains the same fluorescent material that is present in your natural water samples. When the AquaFluor® is calibrated using primary standards of known concentration, readings will be displayed as actual quantitative concentrations. For example, if a 100-ppb standard of Dye X is used to calibrate the AquaFluor®, then any measured samples containing Dye X will be reported as Dye X ppb concentrations. For a list of calibration solutions that can be used for the different AquaFluor applications, use this article: Calibration Solutions for Various Instruments | Turner Designs Help Center
Standards and samples must be in the linear detection range of the instrument to get accurate quantitative results. Refer to Appendix B in the user manual for more details about the linear range and quenching of the samples.
For dye tracing applications the calibration is normally performed with a primary standard made from the same dye that is being used for the testing. The primary standard will either be made to a known concentration, typically in ppb (µg/L) units, or to a known dilution factor. Ideally the Primary Standard and Blank samples used for calibrating will be made with the same water the tests are being performed in. For more details on this and tracer dye use, please refer to the Turner Designs website under Application Notes for “A Practical Guide to Flow Measurement”.
Calibration Procedure
It is recommended for best accuracy, that you calibrate the AquaFluor® before performing your initial sample analysis. The AquaFluor® will save the calibration settings for each channel until a new calibration is performed.
Note: If your sampling conditions remain consistent, i.e., concentration range and temperature, it is not required to recalibrate the AquaFluor®.
If the temperature of your samples or the AquaFluor® changes significantly, the readings may show a small shift and, in this case, you should consider recalibrating. The solid secondary standard is a useful tool for checking the reading stability over time. Refer to Using a Solid Standard for Instruments | Turner Designs Help Center instructions for using the solid secondary standard.
The AquaFluor® is designed with “ambient light rejection”. The black sample compartment cover does NOT need to be closed when reading samples. This allows for the use of test tubes with different height dimensions, such as the 12x75 mm round bottom glass tubes. However, take care to avoid taking readings under direct sunlight. The orientation and cleanliness of the cuvettes or test tubes will have an impact on the accuracy of your results.
Calibration Procedure for Firmware prior to 1.00
Assign a Calibration Standard Value
This defines the numeric value that you want the standard to read. For example, if you calibrate with a primary standard that has a concentration of 50 µg/L, then you will set the value to 50 and the resulting unit of measure will be in µg/L.
Press <STD VAL>the button.
Use the ↑ or ↓ arrow buttons to set the standard value. Pressing the arrow button quickly will make a small incremental change to the value. Press and hold the arrow button to change the value using fast scrolling.
When finished, press the <ENT> button to accept the value and to return to the Home screen.
Perform the Calibration
Press the <CAL> button.
Press <ENT> to start the calibration.
Insert your blank sample and press <ENT>. The AquaFluor® will average the reading for 10 seconds and set the blanking zero point.
Insert the standard sample and press <ENT>. The reading is averaged for 10 seconds and then the Standard Calibration value is set.
Press <ENT> when the calibration is complete to accept the calibration. If <ENT> is not pressed within 10 seconds, you will be asked if you want to abort the calibration. Press the ↑ or ↓ arrow button to abort or accept the calibration respectively.
If at any time during steps 1 - 4 you want to stop the calibration, press <ESC>. This will return you to the Home screen and will default the instrument to the previous calibration
Calibration Check
Once the AquaFluor® calibration is accepted, press the <DIAG> button to select calibration details
View slope and fit for the linear regression of your calibration curve. The slope is used to calculate concentration estimates using fluorescence detected from measured samples. The fit provides information about the linearity of your calibration curve. For example, a fit value of 0.90 indicates that 90% of the points fall within the regression line. The closer to 1.00 (100%) the greater the correlation between your standards’ known concentration values and their respective responses
Calibration Procedure for Firmware 1.00
If at any time during steps 2a-2L you want to stop the calibration, press <ESC>. This will return you to the Home screen and will default the instrument to the previous calibration.
Assign a Standard Value
This value is used as a multiplier. If the value is set to 1 the measurements will not be scaled or adjusted and will reflect actual concentration estimates. If calibrating with primary standard that has concentration in ppb, set value to 1. If you calibrate with a primary standard, that has a concentration of 50 µg/L, and you want to use a multiplier, set the value to the desired multiplier and the resulting unit of measure is multiplied by the value providing an adjusted value in µg/L. NOTE: This action will not change the saved calibration; it simply multiplies the value displayed.
Press the <STD VAL> button.
Use the ↑ or ↓ arrow buttons to set the standard value. Pressing the arrow button quickly will make a small incremental change to the value. Press and hold the arrow button to change the value using fast scrolling.
When finished, press the <ENT> button to accept the value and to return to the Home screen.
Perform the Calibration
Press the <CAL> button.
Press <ENT> to start the calibration.
Insert your blank sample and press <ENT>. The AquaFluor® will average the reading for 10 seconds and set the blanking zero point.
The Relative Fluorescence Unit (RFU) for the blank will be displayed. Record this value for personal records and press <ENT>
Remove the blank.
Use the ↑ or ↓ arrow buttons to adjust the standard value. Pressing the arrow button quickly will make a small incremental change to the value. Press and hold the arrow button to change the value using fast scrolling.
Insert the cuvette with your standard and press <ENT>.
The reading is averaged for approximately 10 seconds.
The RFU for the standard measured is displayed. Record this value and press <ENT>.
Remove the standard.
Using the arrow keys, select to measure another standard or end the calibration and press <ENT>.
If Next Standard is selected, repeat steps f-k for your next standard.
If Cal Done is selected Press <ENT>, go to step L.
Press the ↑ or ↓ arrow buttons to toggle through the units available: RFU, ppb, ppm, µg/L, NTU, µM, cells/mL.
When desired units are displayed, press <ENT>. The Home screen will display the units selected indicating calibration process has completed.
Calibration Check for Firmware Version 1.00
Once the AquaFluor® calibration is accepted, press the button to see what your standard to blank ratio is on Firmware Version 1.00. The standard to blank ratio is defined as:
For most applications and users, a ratio between 5 and 30 is acceptable. The ratio needs to be greater than 5 so you have enough separation between your blank and standard.
If your %FS -STD:%FS-Blk ratio is greater than 30, and the Calibration Standard used is a concentration in your sample range, you can start measuring samples.
If your %FS -STD:%FS-Blk ratio is greater than 30, and the Calibration Standard used is not a concentration in your sample range dilute your Calibration Standard accordingly and recalibrate. Check the ratio again.
If your ratio is less than 5 you'll have to find a more concentrated standard for calibration and recalibrate.
If your ratio is greater than 5 and in a concentration range for your samples you can start measuring samples.
Best Practices with Sample Analysis
Handling Samples
Take care not to spill samples into the sample chamber. Wipe up any spills promptly.
The cuvette MUST BE DRY on the outside when taking readings. Any moisture or condensation on the outside of the cuvette can affect the reading.
Fill the cuvette with at least 3mL solution volume or at least 3/4 full. Significant error in the readings can result if the cuvette contains less than this minimum volume.
The AquaFluor® is very sensitive and even small amounts of material from a previous sample may contaminate the sample and result in errors. Use a clean cuvette for all readings. If you are using the same cuvette for your samples it is very important that you thoroughly rinse the cuvette between samples. A good way to confirm the cuvette cleanliness is to read a blank solution. If the reading is higher than the normal blank reading, the cuvette is not clean.
Any bubbles in the sample will affect the readings. Take care not to introduce bubbles into samples. Remove any bubbles by lightly tapping with your finger on the outside cuvette wall or cover the top of the cuvette and tilt the sample to help dissipate bubbles.
Positioning Samples
The orientation of the cuvette in the sample compartment can give slightly different readings especially for low concentration samples. This is due to variations in the walls of the cuvette that are not readily visible to the eye. We recommend that the cuvette be marked at the top on one side and positioned in the sample compartment the same way each time for best results.
Turbidity sample measurements are particularly sensitive to the quality and cleanliness of the cuvette. Small scratches or smudges on the cuvette will affect the accuracy of the readings. The Polystyrene cuvettes (P/N 7000-957) give the best Turbidity measurement results, due to better quality of the cuvette.
Data Quality
The AquaFluor® is only as accurate as the standards that are used to calibrate it. This is why it is important to take care when preparing standards, samples and blank. One should follow good laboratory practices when preparing all solutions and samples.
The best results are obtained if you determine the range you anticipate your samples are going fall in and calibrate to optimize resolution. If a low concentration calibration standard is used then the low end of the linear range will have better resolution. If a high concentration calibration standard is used the high end of the linear range will have better resolution.