Transcript 1. Lab1and PreLab 1 pipetting, 2013

1. Which tool would you use for
picking up 15 microliter? Which
tool for 25 ml? Why?
2. Why do the little test tubes
look this way?
3. Answer “Micropipette
technique”
Presenting
students:
Refer to this
handout, solve
it and explain
it.
Pre-lab 1 with practice plates
and
Lab 1 - Introduction to
Microvolumetrics and Pipetting
Hand out Practice Exercise
Student Guide pages 1.1 – 1.6
Purpose of Pre-lab 1 and Lab 1
• Become familiar with the small
volumes of solutions used in molecular
biology
• Introduce proper use of the adjustable
digital pipette
• Practice loading and running a gel
Decimals, prefixes, and units
microliter (l) = 1 / 1,000,000 = 10-6
0.000001 = one millionth of a liter
mililiter (ml) = 1 / 1,000
= 10-3 of
0.001 = one thousandth of a l
1 l = 0.001 ml
10 l = 0.01 ml
100 l = 0.1 ml
1000 l = 1.0 ml
Calculations: PreLab 1
Video – Micropipettes
(Open directly from folder;
about 10 minutes!)
OR – Go to SLIDE 11
Types of digital micropipettes
Top number = maximum allowed!
Types of digital micropipettes
How to set volume and use digital pipettes
Dispensing small volumes into a microfuge tube
Micropipetting “ Nevers”
Never - Use without a tip in place
Never - Lay it down with sample in the
tip
Never - Let the “plunger” button
snap back
Micropipette Tips
* Treat tips as “sterile”.
Always replace the top
cover of the tip box quickly.
* To add a tip, press the
end of the micropipette
firmly into the top of the
tip.
* To eject a tip, press the
eject button.
Get a feel for the
first and second
“stops”
* See the different
heights of the
plungers.
* Feel the different
distances the
plunger travels to
the first stop at the
different settings.
Set at 2.0 ul
20.0 ul
Loading and dispensing a sample
Loading:
Depress the plunger to the first stop.
(Not to the second stop!)
While still holding down the plunger,
place the tip in the sample and draw
fluid up by gradually allowing the
plunger to rise.
Withdraw the tip.
Examine the sample. It should extend
to the tip with no air bubble.
Dispensing:
Move pipette tip into the
desired tube near the
bottom.
Slowly press plunger to
the first stop to transfer
the sample, watch liquid
stick to wall.
Continue to press the
plunger to the second stop
to insure transfer of all the
sample.
Remove tip, then release
plunger.
Pipetting Technique
• Hold
micropipette
and microfuge
tubes at eye
level when
loading or
dispensing
samples
• With both elbows on
the table, use your
other hand to
stabilize the bottom
of the pipette.
• This student is
showing one
technique using a
practice gel.
Pipetting exercise 2 (p1.4)
Write group symbol and period
# on test tubes.
Keep mixtures in your box for
next step.
Journal 05.10.13:
No journal question yet. Get your
samples ready to be loaded onto a
gel.
-Complete mixtures A,B,C (Page 1.4)
-Spin down (if needed).
Loading Gels:
Insert pipette tip:
•Under buffer level
•Above gel well
Pipetting into a gel:
Cover the gel in the petri dish with
water.
Load 10 µ liter of loading dye in
each well.
(The glycerol in
the sample
should help it
sink in the well)
Loading errors
Tip should be
above, not in the
well.
Tip punched
through the gel
Dye spreading
under the well
K.
Schramm
loading of the
gel
Tip in buffer
Top surface of
gel
Sample in well
Bottom
surface of gel
K. Schramm
K. Schramm
Different pipetting techniques –
stablility is the key
Types of digital micropipettes
How to set volume and use digital pipettes
Dispensing small volumes into a microfuge tube
Using small volumes
Preparing an agarose gel
loading agarose gel
Teacherdomain folder (e.g. for Andres Berrio)