Working With Micropipets: Practice These Conversions

WORKING WITH MICROPIPETS
INTRODUCTION
Laboratory work in molecular biology and biotechnology is usually done in minute quantities.
The unit of measure used for setting up reactions is the microliter (µl). One microliter is one
millionth (10-6) of a liter.
So: 1 L = 1,000,000 µl, and 1 ml = 1,000 µl
Practice these conversions:
1. Convert the following to ml:
100 µl
500 µl
3,000 µl
10 µl
2. Convert the following to µl:
5 ml
0.5 ml
0.004 ml
0.000001 ml
The micropipet is an instrument that allows us to
accurately measure µl volumes of reagents. Micropipets
are delicate, very expensive, and the cornerstone of our
work with DNA. In this lab, you will learn to properly use
and care for micropipets. A micropipet uses suction to
draw up specific amounts of liquid. Its parts allow you to
control how much liquid to suck up and dispense. It is
essentially a hollow barrel with an adjustable plunger
through it. On the left is a diagram of a micropipet and its
specific parts.
The control button, or plunger, allows the user to suck up
and dispense liquid.
The eject button allows ejection of micropipet tips after
use.
The volume knob allows the user to dial the amount of
liquid to be measured.
The number window shows the amount dialed.
The tip of the micropipet is where the micropipet tips are
placed. The entire white part is called the barrel.
MICROPIPETTING
Damaging these instruments can be avoided by following a few simple rules:
• Never rotate the volume knob beyond the upper or lower

range of the micropipet.
• Never use a micropipet without a tip in place.
• Never lay down a micropipet with a filled tip.
• Never allow plunger to snap back after ejecting fluid.
• Never immerse barrel of micropipet in fluid.
• Never flame micropipet tips.
Micropipets are designed to deliver a specified volume within a certain range, with the
appropriate tip in place. You have micropipets for the following ranges:
Name of micropipet Range of Volumes Delivered Tip To Use
P1000 200-1,000µl Blue
P200 20-200µl Yellow
P20 2-20µl* Yellow
*We use this micropipet to measure down to 1µl
Perhaps the most difficult part of using micropipets is setting them properly. On each of the
micropipets, you will find 3 numbers places in the number windows. However, the numbers
represent different volumes for P1000, P200, and P20:
Number P1000 P200 P20

1st X,000µl X00µl X0µl
2nd X00µl X0µl Xµl
3rd X0µl Xµl 0.Xµl
You will notice a red line on the P1000. This represents a decimal point in ml. The red line on
the P20 is the decimal point for µl.
Practice setting the following volumes:
1. P1000: 324, 1000, 546µl

2. P200: 24, 156, 87µl
3. P20: 2.4, 18.9, 6.0µl
2
MICROPIPETTING
USE OF MICROPIPETS
1. Observe the instructor's demonstration on the proper use of the
micropipet before beginning this exercise.
2. Obtain two 1.5 ml microfuge tubes and fill one with distilled
water. You will practice transferring liquid from one tube to the
other.
3. Choose a micropipet and set the dial to a desired volume. To
operate, your thumb should be at the top of the plunger, and your
fingers wrapped around the body. You may have the ejector
positioned under your thumb (see picture above) or facing out (I
prefer it facing out).
4. Place a tip onto the micropipet by pressing the tip of the
micropipet barrel firmly into a tip of the appropriate type (blue or yellow)
5. Depress the plunger to the first stop.
6. While holding the plunger down, place the tip into microfuge tube and into the liquid.
7. Slowly withdraw your thumb to suck liquid into tip. Watch that it goes up without air
bubbles. Do not snap back plunger!
8. Place the tip into the bottom of the receiving microfuge tube.
9. Press plunger to first stop to dispense liquid. Continue to press beyond to first stop to get out
all of the remaining liquid in the tip.
10. Pull tip out of liquid before relaxing the plunger back to original position.
11. Eject tip into waste container by pressing the ejector button.
USE OF MICROFUGES
1. Observe the instructor demonstrate the proper use of the microfuge and how to insert tubes in
a balanced configuration. This is extremely important, because spinning tubes in an
unbalanced position will damage the microfuge!
2. Be sure tubes you are spinning are in pairs and have approximately the same weight/volume
in them.
3. Open lid and remove rotor cover. Place tubes in pairs arrange so that they are at opposite
ends in the rotor.
4. Replace rotor cover and close lid.
5. Select appropriate time and push start. For short pulses, hold the pulse button for the desired
time.
6. Wait for rotor to stop completely before opening lid and removing your tubes.
3
MICROPIPETTING
SMALL VOLUME PIPETTING EXERCISE

1. Obtain a P20 micropipet, yellow tips and 2 microfuge tubes.
2. Label the tubes A and B. Use the matrix below as a guide for adding appropriate volumes of
the colored solutions to each
tube. Tube Red Blue Yellow Water Color Observed
3. First add the appropriate A - 2 µl 2 µl 11 µl
amount of water to the B 3 µl 2 µl - 10 µl
bottom of each tube. Be sure all of the liquid comes out and forms a small bubble of liquid at
the bottom of the tube.
4. Next add the colored solutions one at a time. Dispense the solution directly into the small
bubble of liquid at the bottom of the tube.
5. After you have added all of your solutions into each tube, practice mixing the contents with a
micropipet. Set your micropipet to 15 µl and slowly pipet the mixture up and down until well
mixed.
6. Place all the tubes in the microfuge and apply a short (1-2 seconds) pulse. Make sure the
tubes are placed in a balanced configuration!
7. Record the final color of the solution in each tube in the table above.
8. A total of 15 µl was pipetted into each tube. Check that your measurements were accurate by
setting the P20 to 15 µl and withdrawing the contents of each tube.
LARGE VOLUME PIPETTING EXERCISE

1. Obtain a P200 and P1000 micropipet, yellow tips and blue tips and 4 microfuge tubes.
2. Label the tubes C, D, E, & F
3. Use the matrix below as a guide for adding appropriate volumes of the colored solutions to
each tube.
4. Mix well (by finger Tube Red Blue Yellow Water Color
vortexing or micropipetting) Observed
and place all the tubes in the C 40 µl - 40 µl 920 µl
microfuge and apply a short
D - 20 µl 260 µl 720 µl
(1-2 seconds) pulse. Make
sure the tubes are placed E 120 µl 20 µl 860 µl
in a balanced F 60 µl 30 µl 30 µl 880 µl
configuration!
5. Record the final color of the solution in each tube in the table above.
6. A total of 1,000 µl (1ml) was pipetted into each tube. Check that your measurements were
accurate by setting the P1000 to 1,000 µl and withdrawing the contents of each tube.
4
MICROPIPETTING
ANALYSIS
Today, you will take a quiz and demonstrate your micropipetting skills. Be sure you know how
to:
1. Convert from liters to milliliters to microliters.
2. Identify micropipet parts and their purposes.
3. Choose and set the correct micropipet for the job.
4. Properly use the micropipet.
5. Properly use a microfuge.

Working With Micropipets: Practice These Conversions

Uploaded by

Copyright:

Available Formats

Working With Micropipets: Practice These Conversions

Uploaded by

Document Information

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Working With Micropipets: Practice These Conversions

Uploaded by

Copyright:

Available Formats

WORKING WITH MICROPIPETS

Damaging these instruments can be avoided by following a few simple rules:

• Never rotate the volume knob beyond the upper or lower

Number P1000 P200 P20

1. P1000: 324, 1000, 546µl

SMALL VOLUME PIPETTING EXERCISE

LARGE VOLUME PIPETTING EXERCISE

You might also like

Pfad - The Proxy pFad of © 2024 Garber Painting. All rights reserved.