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Frequently Asked Questions

  1. What is a fluorescent dye?
  2. Why use fluorescent dyes?
  3. Which fluorescent dye(s) should I use?
  4. How much dye should I use?
  5. Can you detect more than one dye in a sample?
  6. Should I use charcoal receptors or water samples?
  7. How many sites should I monitor?
  8. Is dye purchased from one company equivalent to that from another company?/li>

1.What is a fluorescent dye?

Fluorescence is a physical property of certain compounds, including some dyes. In fluorescence, light photons of particular energy wavelengths are absorbed by the compound, then emitted at a lower energy level (longer wavelength) (Kass 1998, Rendell 1987).

We most commonly encounter fluorescence when looking at objects lit by a black light. Black lights give off energy in the 300-400nm range. Any material that absorbs light in this range will appear fluorescent. Optical Brighteners fluoresce under black light. They are commonly used in paper, fabrics, and detergents to make whites appear whiter because of this fluorescent property. Tinopal CBS-X optical brightener is one dye used by CHL that will fluoresce under black light. Most dyes used in groundwater tracing fluoresce optimally in light wavelengths above those given off by a black light.

Fluorescent dyes used by CHL absorb light between 300-600 nm and emit light at wavelengths about 15-20 nm longer than their absorbance maxima. CHL analyzes for fluorescent dyes using a Shimadzu RF 6000 or RF 5301 PC spectrofluorophotometer, which synchronously scans an excitation and emission range.

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2. Why Use Fluorescent Dyes?

Groundwater tracing using fluorescent dyes is the best method to determine the connectivity of recharge and discharge points and allows for determination of direction and velocity as well. Dye tracing is a vital tool in understanding groundwater flow and the resulting information is used in a variety of ways to protect groundwater and surface water quality.

Fluorescent dyes make excellent tracers for a variety of reasons, including:

  • They are water soluble.
  • They are highly detectable (some below 10 parts per trillion).
  • They have extremely low toxicity ratings.
  • They are inexpensive.
  • They are fairly stable in natural waters.
  • They have been used successfully for tracing for over one hundred years.


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3. Which fluorescent dye(s) should I use?

The answer to this question depends on:

  • Laminar or turbulent flow of water
  • Number and type of injection locations
  • Background fluorescence
  • Water quality parameters


We recommend background fluorescence analysis for every monitoring location, which must be done prior to the dye injection. Background monitoring helps to determine the types and amounts of dye that work best for your site by revealing the any background fluorescence of the water at your monitoring sites. Water will often have some level of fluorescence from natural or man-made sources. When multiple injections are planned, we choose dyes that have very different wavelengths of fluorescence. For example, fluorescein and sulphorhodamine B are good “companions” for dye trace investigations since their fluorescence wavelengths are 510 nm and 582 nm in water, respectively.


Crawford Hydrology Laboratory uses the following dyes:

  • Tinopal CBS-X (Optical Brightener)
  • D&C Green 8
  • Fluorescein (Uranine C, Acid Yellow 73)
  • Eosine (Acid Red 87)
  • FD&C Red 3
  • D&C Red 28 (Phloxine)
  • Rhodamine WT
  • Sulphorhodamine B (Acid Red 52)


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3.How much dye should I use?

There has been much debate on this subject and there are formulas to help determine the amount of dye needed. However, there is no easy answer, since the travel times and dilution factors are largely unknown at most locations. Never-the-less, 1-10 lbs of dye will suffice for most dye traces. We will make recommendations for each situation based on distance to monitoring sites, the site’s geologic and/or aquifer characteristics, expected travel times, type of injection site (well, sinkhole, sinking stream), and our past experience.
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5. Can you detect more than one dye in a sample?

Yes. Each dye we sell has a unique wavelength of fluorescence. We can uniquely identify each dye, and can identify multiple dyes in a sample. Dyes with similar wavelengths may have fluorescent peaks that overlap. In these cases, peakfitting software is necessary. We use this program conservatively to estimate the peak location and concentration of dyes with overlapping peaks. However, a high concentration of one dye can mask the peak of a dye with a very low concentration.
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6. Should I use charcoal dye receptors or water samples?

Using activated charcoal packets, installed in a well, stream, or spring is the best and the most cost-effective method for determining point-to-point connections of groundwater flow.  Fluorescent dyes are adsorbed onto the charcoal over time, and can be detected from eluted charcoal at lower concentrations than in water. However, charcoal can not quantitatively assess the amount of dye in the water.

Analysis of water samples for fluorescent dyes gives an accurate quantification of the dye concentration in the water at a given time. Using periodic sampling, especially with an automatic water sampler, provides precise information about the timing and concentration of the dye as it moves through the system in the form of a dye break-through curve. This information is valuable for detailed hydrologic studies, and is valuable in potentially contentious dye trace investigations.

You may wish to use a combination of charcoal dye receptors and water sampling for your dye trace investigation. Even if you plan to use only charcoal receptors, those purchased from CHL include a water sampling vial, which should be filled at the time of receptor collection as a back-up. The water sample can be held at the lab, and analyzed only if needed.
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7. How many sites should I monitor?

In order to not miss the dye, it is necessary to monitor every possible resurgence point between the injection location and a major discharge boundary, such as a river.  When conducting a dye trace, the dye must be detected at one or more locations to make any inferences to groundwater flow.  In the event that some or all of dye resurgence points are missed, or a dye is not recovered,  a repeat of the dye trace may be necessary.
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8. Is dye purchased from one company equivalent to that from another company?

Most fluorescent dyes are not made for laboratory or water tracing applications. Dye manufacturers produce products with varying purities and strengths. The specifications vary from manufacturer to manufacturer and distributor to distributor. Additionally, the same dye can have many names. One dye may be cheaper than another because it is actually less pure. For example, a “green dye” may look similar to fluorescein, but it may fluoresce at a different wavelength than the fluorescein CHL sells.

CHL has obtained the most reasonably priced quality bulk dyes with the highest purity and strength. We test each batch of dye received to ensure quality and check for cross-contamination problems. We carry the same dye products that we use to make our laboratory standard solutions. We know the fluorescence emission wavelengths of the products and their purities.

You can use dyes purchased from other companies. On a CHL lab analysis report sheet, the concentrations (parts per billion) of dye in a sample will be reported in units equivalent to the dyes we have in stock.

If you obtain powder dyes from CHL or another source, these should be mixed with water before arriving at the injection location,in most situations. CHL dyes are mixed to the solubility point and shipped to you for ease of use, clean-up and minimization of cross-contamination risk.

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Kass, Werner. 1998. Tracing Technique in Geohydrology. Stuttgart, Berlin. 581 pages.
Rendell, David. 1987. Fluorescence and Phosphorescence. Analytical Chemistry by Open Learning. Great Britain

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