CLINICAL LAB SCIENEC

CHAPTER 16: IMMUNOLOGY AND SEROLOGY 405
level of antibodies against a specific antigen may be required. Normal saline, which
approximates the amount of sodium chloride found in the plasma, is commonly used
as a fluid for diluting the samples. The higher the dilution, the higher is the amount
of antibodies or antigens that are being tested for because larger amounts of the
diluting fluid are being used. As the dilutional ratio becomes higher, the amount of
the original sample becomes smaller and the amount of diluent becomes larger. This
concept is sometimes difficult for the beginning laboratory technician or technologist
to understand. The following procedure should serve to inform the student of the
purpose and theory behind a serial dilution titer.
When a serial dilution is performed on a patient’s serum, it is necessary to
observe the pattern of positive test results obtained. A phenomenon called a prozone
reaction may cause results to be negative in early, less dilute specimens, while
increasingly stronger reactions are seen as the dilutions become more dilute. This
situation arises when the level of antibodies is extremely elevated and the optimum
level of antibody to antigen is exceeded. To find the titer (antibody level) of a certain
antibody, clinical laboratory workers prepare serial dilutions of a serum sample,
usually in a two-fold dilution pattern, where each subsequent dilution halves the
level of antibodies in the diluted serum.
An antibody titer that consists of serial dilutions of serum is a semi-quantitative
serological procedure. It is used to determine the stage of infection or the patient’s
response to a medical condition. In setting up a serial dilution, the patient’s sample
is diluted the number of times that will yield a significant titer (level of antibodies).
Samples are set up by consistently doubling the dilution by a factor of 2 for each
tube: 1:2, 1:4, 1:8, etc. (Figure 16-1). This is called a two-fold dilutional titer and is the
most common ratio used for antibody titers.
However, dilutions are not always by a factor of 2; sometimes, factors of 10 may
be used, such as with dilutions of 1:10, 1:20, etc. This also results in a doubling of
the ratio for each successive tube. Sometimes it is necessary to make dilutions for
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 16-1 Setting up a two-fold dilution series.
Source: Delmar/Cengage Learning.
1. Set up 9 tubes, each containing 1 mL of diluent.
2. Transfer 1 mL of patient serum to tube 1.
3. Mix the serum and diluent, and transfer 1 mL of the mixture to tube 2.
4. Repeat the procedure, transferring 1 mL each time after mixing with diluent.
5. Discard 1 mL from the last tube.
6. When dilution series is complete, each of the 9 tubes should contain 1 mL.
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