CLINICAL LAB SCIENEC

CHAPTER 9: LABORATORY MATHEMATICS 225
1. Set up 9 tubes, each containing 1.0 mL of diluent.
2. Transfer 1.0 mL of patient serum to tube 1.
3. Mix the serum and diluent, and transfer 1.0 mL of the mixture to tube 2.
4. Repeat the procedure, transferring 1.0 mL each time after mixing with diluent.
5. Discard 1.0 mL from the last tube.
6. When dilution series is complete, each of the nine tubes should contain 1.0 mL.
1 mL
serum
1 mL 1 mL 1 mL 1 mL 1 mL 1 mL 1 mL 1 mL 1 mL
discard
Serum
Dilution 1:2
1:4
1:8
1:16
1:32
1:64
1:128
1:256
1:512
Tube
#1 #2
#3
#4
#5
#6
#7
#8
#9
FIGURE 9-12 Setting up and performing a two-fold serial dilution.
Source: Delmar/Cengage Learning.
3. The last tube will contain 2.0 mL, so 1.0 mL must be discarded from this tube,
since the addition of any indicator cells, etc., would change the dilution of the
last tube (Figure 9-12).
Interpretation
Each sample after the first tube will have 1.0 mL added to an equal amount of
diluent. This will effectively cut by one-half the strength of the preceding tube
from tubes 2 through 9. If a reaction occurred in tubes 1 through 5, the result
would be reported as positive at a 1:32 dilution.
Beer’s Law
Beer’s law is used when a spectrophotometer measures the amount of light absorbed
when the beam is directed through a solution that may be colored or where a reaction
may occur at a wavelength not visible to the naked human eye. These values are
measured with an ultraviolet wavelength and are often used for enzymatic reactions
where an enzyme reacts with a substrate contained in the reagent. Many reactions
follow Beer’s law (that the absorption coefficient is directly proportional to the concentration
of a solution) when a given wavelength is established under conditions
of temperature, pH, length of the light wave, and the depth of the chamber in which
the reacting substances are placed (this is often standardized at 1 cm).
Automated instruments and manual spectrophotometers use this law for
many common analyses of constituents in the human blood and other body
fluids. When a manual assay is being performed, the amount of concentration
in a patient’s sample is compared with a known standard. A simple Beer’s law
graph (Figure 9-13) indicates absorbance in nanometers (nm). A spectrophotometer
is used to measure the absorbance of the standard and the absorbance
Copyright 2010 Cengage Learning. All Rights Reserved. May not be copied, scanned, or duplicated, in whole or in part. Due to electronic rights, some third party content may be suppressed from the eBook and/or eChapter(s).
Editorial review has deemed that any suppressed content does not materially affect the overall learning experience. Cengage Learning reserves the right to remove additional content at any time if subsequent rights restrictions require it.