Xylem 6-Series Multiparameter - Manual (Page 272)

Principles of Operation Section 5
YSI Incorporated Environmental Monitoring Systems Manual 5-34
2. Accurately weigh 0.500 g of the 20 % Rhodamine concentrate, quantitatively transfer the viscous liquid
to a 1000-mL volumetric flask and fill the flask to the top graduation. Mix well. This solution contains
100 mg of Rhodamine WT per 1000 mL of water.
3. Accurately transfer 1.0 mL of the solution prepared in the above step to a 1000 mL volumetric and then
fill the flask to the top graduation with purified water. Mix well to obtain a solution that is 100 ug/L
(0.10 mg/L) in water (a 1000:1 dilution of the concentrated solution).
4. Store the concentrated standard solution in a darkened glass bottle in a refrigerator to retard
decomposition. The dilute standard prepared in the previous step should be used within 5 days of its
preparation.
When Rhodamine WT standards are required in the future, perform another dilution of the concentrated dye
solution after warming it to ambient temperature. Our experience has indicated that the concentrated solution
that has been kept at cold temperatures is much more stable than the dilute solution stored at room
temperature.
It is well known that the intensity of the fluorescence of many dyes shows an inverse relationship with
temperature. The effect must also be accounted for when “calibrating” the YSI PC sensor with Rhodamine
WT. Enter the calibration value from the table below corresponding to the temperature of the standard.
WARNING: The “PC tempco” factor in the Advanced|Sensor menu MUST BE SET TO ZERO,
when calibrating with Rhodamine WT.
Table. Approximate PC-containing equivalent of 100 ug/L Rhodamine WT as a function of temperature.
T, C Cells/mL to Enter T,
C
Cells/mL to Enter
30 44940 18 76580
28 49700 16 83580
26 54600 14 91420
24 58940 12 98140
22 64120 10 107940
20 70000 8 113540
REMEMBER: The use of Rhodamine WT for “calibration” of the Phycocyanin Probe is only an
approximation. To assure the maximum accuracy for the 6131 sensor, the user must relate the field
fluorescence readings to data from actual BGA samples as described above. YSI does not provide an
accuracy specification for the sensor due to the limitations described above and below.
EFFECT OF TURBIDITY ON BGA-PC READINGS
As described above, the filters in front of the photodiode in the YSI 6131 Phycocyanin Probe prevent most
of the orange light which is used to excite the Rhodamine molecules from reaching the detector after being
backscattered off of non-fluorescent particles (turbidity) in environmental water. However, the filter
system is not perfect and a minor interference on PC-containing BGA readings from suspended solids will
result. Laboratory experiments indicate that a YSI 6131 sensor will have a turbidity interference
characterized by a factor of about 21 cells/mL of PC-containing BGA per NTU of turbidity. For example,
at a turbidity of 100 NTU, a PC-containing BGA reading of 2100 cell/mL will be observed over and above
the reading actually due to the presence of BGA. Users may wish to use the independently-determined
turbidity value and the above compensation factor to correct measured PC-containing BGA values using,
for example, a spreadsheet.