FASTtracka Fluorimeter, Fast Repetition Rate Fluorimeter



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FAST^tracka System Datasheet (pdf)		Environmental Monitoring (pdf) Algae Monitoring using FRRF (pdf)
FAST^actDatasheet

Fast Repetition Rate fluorometry (FRRf) is a sophisticated, non-intrusive method for probing oxygenic photosynthesis in algae, through detailed analysis of chlorophyll a fluorescence.

We have developed a very sensitive, cost-effective and fully automated FRRf-based system for the detection of toxicants within domestic water supplies, which employs a multi-parameter model to analyse FRRf data from algae (the 'transducer' within the system). This system provides much greater sensitivity, to the presence of toxicants, than can be realised with alternative chlorophyll-a fluorometers, plus limited 'mode of action' information.

Laboratory data, which clearly demonstrate the ability of this system to detect a number of toxicants at sub-lethal concentrations, have been used to develop a 'Red, Amber, Green' (RAG) algorithm, which can be applied in real time. With this algorithm, an

Amber event is triggered if one or more key parameters changes beyond a preset limit, in the right direction and on a time scale that is too rapid to be attributable to natural variation. A Red event is triggered if this change persists beyond a set number of sequential measurements.The first variant of the RAG system uses natural flora as the target material. This makes it suitable for use with unprocessed water, such as reservoirs. Field data from a number of UK and US sites have shown the RAG system to be very resistant to the triggering of false positives, by diurnal cycles and other natural phenomena, whilst being sensitive enough to detect a real life contamination event.

A second variant of the RAG system, primarily being developed for use with treated water supplies, utilises immobilised cells of Chlorella sp. at the target material. This system should provide substantially enhanced detection capabilities and even greater resistance to false positives.
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The Fastracka II in profiling mode being deployed off the RV Polarstern during the LOHAFEX cruise in early 2009.

The FASTtracka II in profiling mode being deployed off the RV Polarstern during the LOHAFEX cruise in early 2009.

Applications

-	Monitoring photosynthetic performance in phytoplankton (oceanography, limnology)
-	Primary production studies (oceanography)
-	Toxicity monitoring
-	Water quality monitoring (reservoirs, rivers and streams)
-	Bloom detection
-	Environmental monitoring of phytoplankton populations (water supplies, fish farming)

Features

-	Advanced programmability, with the facility to concatenate up to 10 protocols with up to 5 blocks of flashlets in each protocol
-	Straightforward implementation of both Single Turnover (ST) and Multiple Turnover (MT) modes
-	Suitable for profiling and in towed vehicle systems, where the ST mode is essential
-	Facility to incrementally increase the interval between flashlets, between 0 and 100%, for improved relaxation kinetics in both ST and MT modes
-	Normalisation of data throughout the full dynamic range (data acquired at different gain levels) is possible, thanks to a newly developed calibration method

-	Options for both PAR and depth sensors
-	Monitors both LED output and photomultiplier sensitivity at each flashlet, thereby minimising measurement artefacts
-	Near surface operation possible, even under high ambient light conditions
-	High rejection of ambient light
-	Accepts supply voltage of between 9 and 72 V
-	Large dynamic range (0 to 600 µg l^-1)
-	Fast data download (up to 900 kb) to an RS232 or USB serial port

Hardware

The RAG system is centred around a FASTtracka II, FRRf sensor. With this sensor, an array of ten, blue LEDs provides very intense, 1 µs 'flashlets' to a volume of approximately 800 mm3. The red chlorophyll a fluorescence, stimulated by the flashlets from the LEDs, is collected through the central part of the front lens and transferred to a photomultiplier tube. A series of filters and apertures block light from the LEDs reaching the photomultiplier tube. Top left shows the front lens of the sensor, which has a 15 mm focal length in water. The 2^nd image provides a cross-sectional view of the 'front end' of the sensor. The 3^rd image models water flow through the sample chamber and the 4^th shows an installed RAG system.
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FRRf and the RAG system
FRRf provides detailed information about photosystem II (PS II) function. Because PS II sits at the start of photosynthetic electron transport, PS II function is sensitive to a range of toxicants with 'downstream' sites of action. FRRf uses the 'single turnover' method, transiently to drive all PS II centres from an 'open' state into a 'closed' state during a 200 µs saturation phase of 100, 1 µs flashlets. This results in an increase in the yield of chlorophyll a fluorescence from PS II, to a maximum level. During a subsequent 2.5 ms relaxation phase, more widely spaced 1 µs flashlets allow closed centres to re-open, resulting in a decrease in fluorescence yield. .	Click image to enlarge
The blue line in the following traces was generated by an iterative curve-fitting process, which operates in real time. This curve fit generates the four RAG parameters used to detect potential contamination events – Fv/Fm and p from the saturation phase, plus tf and Ra from the relaxation phase, as shown.
Click image to enlarge	Click image to enlarge
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Lab-based measurements
The RAG algorithm initially was developed using data from lab-based tests with a range of toxicants, conducted by DSTL. These data led to identification of the four standard RAG parameters: Fv/Fm and p from the saturation phase, plus tf and Ra from the relaxation phase. Differences in the combined affect on the four RAG parameters were seen with different toxicants, allowing for some discrimination among toxicant types.
Click image to enlarge -
Additional lab-based tests were conducted, to verify the robustness of the four RAG parameters at different measurement LED intensities, as a proxy for variable levels of turbidity within natural samples.
Click image to enlarge
. Current laboratory work is directed towards development of a new RAG system variant, which uses immobilized Chlorella sp. at the target material.

. Field Data - including 'an event'
Real-time data analysis and presentation are provided through a Windows-based Graphic User Interface, specifically developed for this application. In the screen-shot, below, the RAG system has been running a short time. The small dialog at centre provides the current status of all four RAG parameters. The graph at bottom provides a continuous record of events. .
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The RAG system uses data from earlier measurements to predict the value of each of the four RAG parameters. If the predicted value is significantly different from the actual value, an Amber event is triggered. If the different persists, a Red event is triggered. The RAG	algorithm successfully handles diurnal fluctuations, whilst maintaining a high degree of sensitivity to potential contamination events. The traces, below, are from an 'uneventful' week, which saw clear diurnal cycling within all four RAG parameters, a few Amber events, but no Red events. ..
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The data presented below show a 'Red' contamination event at a UK site (see inset). The event was detected for the Fv/Fm parameter only, which effectively rules out a number of possible contaminants, including cyanide. Independent analysis confirmed the appearance of Triclosan at a concentration of 22.6 mg m-3, at the time of detection. ..	Half way through the data plot, there is a debris-induced transient change in all four RAG parameters. It is notable that this transient did not trigger a Red event. The dramatic changes towards the end of the data plot are due to a pump failure, which did trigger a Red event.
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Specifications

PHYSICAL
Sensor alone	339.5 mm (excluding connector)
With profiling cage	550.0 mm (excluding connector)
Sensor diameter Ti (Ac)*	112.5mm (130mm)
With profiling kit	170.5mm
Pressure housing	Titanium or Acetal C
Dark chamber	Acetal C
Connector kits	Acetal C + Stainless Steel
Weight in air / water	60 N / 30 N approx. (6kg/3kg)
*Ti = titanium pressure housing, Ac = Acetal C pressure housing
OPERATIONAL
Range (standard)	0 - 600 µg l^-1
Excitation	470 nm peak, ± 20 nm
Detection	685 nm peak, ± 10 nm
Sample volume	1 cm^-3
Internal storage	4 GBytes
Power in	9 to 72 VDC
Operating temperature	-10°C to +40°C
Storage temperature	-10°C to +50°C
Max. operating depth Ti (Ac)*	500 m (200 m)
Basic Instrument:	The FAST^tracka II is provided with a 3 m cable, Handbook and Calibration Certificate and fitting software
Available options:	Deck unit, profiling cage, dark chamber, profiling adaptor kit and pumped adaptor kit. PAR sensor and depth sensor

GAP VIII Workshop on Aquatic Primary Productivity, Eilat, Israel

FASTtracka II sensor was tested in the oligotrophic waters around Eilat and exceeded expectations

Images from the GAP VIII Workshop on Aquatic Primary Productivity (Israel). During this time CTG's Dr Kevin Oxborough worked with the open ocean group to trial the new FAST^tracka II sensor in oligotrophic waters and to discuss the potential of fast repetition rate fluorometry (FRR). In terms of performance, the FAST^tracka II exceeded expectations.

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