The Correct Preparation of

Serum Dilution
 Definition of terms
1. Titer
– End point of dilution
2. Serial Dilution
– The stepwise dilution of a substance in a solution
3. Solute
– Material being diluted
4. Solvent (Diluent)
– The medium making up the rest of the solution
 Dilution
• It is a process to reduce strength of
concentrated solution, or to make specimens
being analyzed to be weaker solution from
stronger solutions in various laboratory
procedures to yield great accuracy.
• There are two things to consider in dilution:
– Solute
– Diluent
• The relationship between these two is expressed as a
fraction.
 Why is there a need in dilution?
1. Preparing standards
2. Preparing quality control materials
3. Preparing patient sample to be read/analyzed by
machine
4. Test result cannot be read because of too high or low
levels of analyte
5. Less specimen available to test
6. Looking for least amount of antibody that will react to
the antigen
7. Aliquot
– A portion of a larger whole, especially a sample taken for
chemical analysis or other treatment
• Dilution can be done singly, or in a random
manner following this formula:
– RD=TV/Volume of solute
– Whereby: RD is the reciprocal of dilution
• Total Volume (TV)= volume of solute + volume of
solvent
• Volume of Solute is the amount of the sample or
(analyte)
 Serial Dilution
• The commonly used serial dilution is the Two-
fold.
 Serial Dilution Scheme
• Each tube contains 0.2 mL reached of diluent.
• Patient serum (0.2mL) is withdrawn and added
to tube two.
• The process is continued until the last tube is
reached.
• The sample is mixed, and 0.2 mL is discarded.
• Note that in this two-fold dilution, the amount
of antibody is slashed in half in each successive
tube.
• Dilution= Volume of solute/ Total volume
• (T1)1/2 x (T2)1/2 x (T3)1/2 x (T4)1/2 x (T5)1/2
– T1= 1:2
– T2= 1:4
– T3= 1:8
– T4= 1:16
– T5= 1:32
– T6= 1:64
• Then when 0.2 mL of the 1:2 dilution is added
to 0.2 mL of diluent, a 1:4 dilution is obtained.
• The final dilution is obtained by counting the
number of tubes and setting up a
multiplication series in which the original
dilution factor is raised to a power equal to
the number of tubes.
• In this example, if the first tube contains a 1:2
dilution, the dilution in tube number six is
1:64.
 Procedure A:
1. Prepare test tubes in a rack
2. Label the tubes as follows: TT1 to TT5
3. Place the following:
a. Place 0.60 mL NSS+0.40 mL serum to TT1 and mix properly.
b. Place 0.80 mL NSS+0.20 mL serum to TT2 and mix properly.
c. Place 0.85 mL NSS+0.15 mL serum to TT3 and mix properly.
d. Place 0.90 mL NSS+0.10 mL serum to TT4 and mix properly.
e. Place 0.92 mL NSS+0.08 mL serum to TT5 and mix properly.
4. Tabulate procedure 3 using and indicate the total
volume and dilution.
5. Show your computation of the total volume and
dilution
 Procedure B:
1. Prepare test tubes in a rack
2. Label the tubes as follows: TT1 to TT5
3. Place the following:
a. Place 0.80 mL NSS+0.20 mL serum to TT1 and mix properly.
b. Place 0.80 mL NSS+0.20 mL serum from TT1 to TT2 and mix
properly.
c. Place 0.80 mL NSS+0.20 mL serum from TT2 to TT3 and mix
properly.
d. Place 0.80 mL NSS+0.20 mL serum from TT3 to TT4 and mix
properly.
e. Place 0.80 mL NSS+0.20 mL serum from TT4 to TT5 and mix
properly.
4. Tabulate procedure 3 using and indicate the total volume
and dilution.