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Manual Pipetting and Serial Dilution

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Tom Anthony Tonguia
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92 views27 pages

Manual Pipetting and Serial Dilution

Uploaded by

Tom Anthony Tonguia
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Manual Pipetting and Dilutions

TDCI MLS
BSMLS 3
Presented by Tom Anthony A. Tonguia, RMT,
MSMT ©
MANUAL PIPETTING
Pipette
• It is a laboratory instrument used to measure
out or transfer small quantities of liquid, in
volumes of milliliters (mL), microliters (μL).
• Pipetting
– It is the process of using a pipette, whether it's a
plastic pipette or a glass pipette, to measure or
transfer a small volume of a liquid sample.
– 100uL=0.1mL; 1000uL=1mL

References: Nichiryo (n.d) About Pipettes. Retrieved from: https://www.nichiryo.co.jp/en/productline/pipette.html


Scilogex (n.d) What is pipetting? Retrieved from: https://www.scilogex.com/blog/our-blog-1/post/what-is-pipetting-a-
simple-rundown-of-the-labs-most-fundamental-job-1
Pipette Characteristics
I. Design
a. To contain (TC)
• Ex. Sahli Hellige pipet, Long-Levy pipets
b. To deliver (TD)
• Ex. Mohr, Serologic and Volumetric pipets
Pipette Characteristics
II. Drainage characteristics
a. Blowout
• Ex. Sahli Hellige pipet, Long-Levy pipets
b. Self draining
• Ex. Mohr, Serologic and Volumetric pipets
Pipette Characteristics
III. Type
a. Measuring or Graduated
1. Mohr
2. Serological
3. Micropipet
• Air displacement pipet
• Positive displacement pipet
4. Bacteriologic
5. Ball, Kolmer or Kahn
Pipette Characteristics
III. Type
b. Transfer
1. Volumetric
2. Ostwald Folin
3. Pasteur pipettes
4. Automatic pipettes
Volumetric vs. Ostwald Folin
Automatic pipette
DILUTIONS
Dilutions
• It represents the ratio of concentrated or stock
material to the final volume of a solution and
consists of the volume or weight of the
concentrate plus the volume of the diluent, with
the concentration units remaining the same.
• The relationship of the dilution factor to
concentration is an inverse one; thus, the
dilution factor increases as the concentration
decreases.
Dilution factor
• Dilution factor= TV/Volume of solution to be
diluted
– *TV= Total Volume
Simple dilution
• Formula:
– Dilution=Volume of the solute/Volume of the
Solute + Volume of the solvent
• Sample Problem:
– Calculate the dilution if 5 milliliter of serum are
diluted with 15 mililiter of saline solution.
Simple dilution
• Diltuion=5/5+15=5/20
• 5/20=1/x
• 5x=20
• X=4
• The dilution is 1:4.
V1 x C1 = V2 x C2
• This involves changing a solution of known
volume and concentration (V1 x C1) to one
weaker concentration (V2 x C2).
• The three of the four values must be known to
solve the equation.
• Any units of volume and concentration can be
used as long as they are the same for both
sides of the equation.
V1 x C1 = V2 x C2
• How much 20% alcohol is required to make 1
liter of 10% alcohol?

V1 x 20% = 1 Liter x 10%


V1=10/20
V1=0.5 L
Proportion
• This is used when reagents are prepared by
adding together a specific amount of another
solution.
• V= C/A+B
• Where:
– C= total volume of final solution
– A= total parts of solution A
– B= total parts of solution B
– V= total volume of one part
Proportion
• Example: A procedure calls for sodium
hydroxide and water with the proportions of
being four parts of sodium hydroxide to six
parts of water. One hundred mililiters are
needed. How much sodium hydroxide and
how much water are required?
• V= 100mL/4+6= 100/10= 10
– Sodium Hydroxide= 4x10= 40mL
– Water= 6x10= 60mL
Serial Dilution
• It is defined as multiple progressive dilutions
ranging from more concentrated solutions to
less concentrated solutions.
• Formula:
– 1/Dilution Fold=volume transferred/total volume
Serial Dilution
• Example:
– What is the dilution fold of the following serial dilution
system consisting of five tubes? The following amounts
of diluents have been added to the tubes:
• 0.5 mL is added to tube 1
• 0.5 mL is added to tubes 2 to 5.
• Next, 0.5 mL of patient serum is added to tube 1 and 0.5 mL
is serially transferred through tube 5.
• Finally, 0.5 mL is discarded from tube 5
– What is the dilution of tube 3 in the preceding serial
system?
Serial Dilution
• Solution:
o 1/x=0.5/1.0
o X=2

o Solution of tube 1 = Dilution of x [1/dilution


fold]^(x-1)
o What is the dilution of tube 3 in the preceding
serial system?
o X=[1/2][1/2]^(3-1) = 1/8= 1:8

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