Laboratory Practices
Laboratory Practices
5940022LabPrac.fm Page 21
term reagent blank refers to that portion of the test result contributed solely by
the reagent. This produces a positive error in the test results.
Every effort is made to produce reagents with the lowest possible blank; and, for
most reagents, it is less than 0.009 absorbance units. It is sometimes impossible
or impractical to produce reagents with such a low blank. When using such
reagents, it is best to determine the reagent blank by performing the procedure
using high-quality water (deionized, distilled, etc.) to zero the instrument.The
resulting value is then expressed in the concentration units of the test and is
subtracted from each sample determination that uses the same reagent lot. The
DR/2400 Spectrophotometer software allows the reagent blank value to be
stored and subtracted automatically from each sample value (see Running a
Blank in the DR/2400 instrument manual). The reagent blank needs to be
determined only at first use, when a new lot of reagent has been opened, or if
you suspect contamination has occurred.
In most tests, the reagent blank is so small the instrument may be zeroed on
either an untreated portion of the original water sample or on deionized water.
This will not result in a significant loss of accuracy unless the test is for very low
levels of a chemical. It is then best to use a reagent blank prepared as above.
2.7 Sample Dilution
Most Hach colorimetric tests use volumes of 10 and 25 mL. However, in some
tests, the color developed in the sample may be too intense to be measured due to
high levels of analyte, or unexpected colors may develop due to an interference.
In either case, dilute the sample to produce a measurable endpoint or to
determine if interfering substances are present.
To dilute the sample easily, pipet the chosen sample portion into a clean
graduated cylinder (or volumetric flask for more accurate work). Fill the
cylinder (or flask) to the desired volume with deionized water. Mix well. Use the
diluted sample when running the test.
Table 4 shows relative quantities and the multiplication factors to use with a
graduated cylinder. The concentration of the sample is equal to the diluted
sample result multiplied by the multiplication factor.
Table 4 Sample Dilution Volumes
Sample
Volume (mL)
mL Deionized Water
Used to Bring the Volume
to 25 mL
Multiplication Factor
25.0 0.0 1
12.5 12.5 2
10.0
1
15.0 2.5
5.0
*
20.0 5
2.5
*
22.5 10
1.0
*
24.0 25
0.250
*
24.75 100
1
For sample sizes of 10 mL or less, use a pipet to measure the sample into the graduated cylinder or
volumetric flask.
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