Thermolab Sartorius MC5 Manual de usuario
Liquid Handling
Application Notebook
Tips on how to pipette
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3
Contents
Pipetting terms ........................................................................................................ 4
Types of pipettes ..................................................................................................... 4
General guidelines and pipetting techniques .......................................................... 7
Recommendations for pipetting different compounds ......................................... 10
Pipetting guidelines for selected compounds ....................................................... 11
Pre enting cross-contamination ............................................................................ 14
Finntip Filter tests .................................................................................................. 15
The plastics of arious Finntips ............................................................................. 15
Troubleshooting .................................................................................................... 16
Calibrating your pipettes ....................................................................................... 17
Formulas for calculating results ............................................................................ 21
Con ersion tables .................................................................................................. 23
Ensuring optimum performance ........................................................................... 24
Factors affecting the accuracy of air displacement pipettes ................................. 26
Maintenance of your Finnpipette .......................................................................... 27
Autocla ing ........................................................................................................... 27
UV resistance ........................................................................................................ 27
General guidelines for decontaminating Pipettes
when Working with Different Liquids .................................................................... 28
Chemical compatibility of plastics ......................................................................... 29
Frequently asked questions .................................................................................. 31
Making your lab work lighter ................................................................................. 34
Finnpipette Warranty Policy ................................................................................... 35
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Pipetting terms
Aspirate - to draw up the sample
Dispense - to deli er the sample
Blow-out - to empty the tip completely
Calibration check - checking the difference between the dispensed olume
and the selected olume
Ad ustment - altering the pipette so that the dispensed olume is within the
specifications
Types of pipettes
Although Thermo Labsystems supplies pipettes for all application needs, most lab-
oratories are equipped with two types of pipettes.
Air displacement pipettes are meant for general use with aqueous solutions.
Positi e displacement pipettes are used for high iscosity and olatile liquids.
Both types of pipettes ha e a piston that mo es in a cylinder or in a capillary. In
air displacement pipettes, a certain olume of air remains between the piston and
the liquid. In positi e displacement pipetting, the piston is in direct contact with
the liquid.
Air displacement pipetting
Air displacement pipetting, used for standard pipetting applications, is highly accu-
rate. Howe er, conditions such as atmospheric pressure as well as the specific
gra ity and iscosity of the solution may ha e an effect on the performance of air
displacement pipettes.
• Finnpipettes for air displacement pipetting:
Finnpipette Digital, Finnpipette Colour, Finnpipette Fixed Volume,
Finnpipette BioControl, Finnpipette Multistepper, Finnpipette Varichannel
• Finntips for air displacement pipetting:
Finntip (standard), Finntip Filter, Finntip BioCon, Finntip Multisteppper,
Finntip Band 4, Finntip Wide
5
How does an air displacement pipette work?
1. The piston mo es to the appropriate position when the olume is set.
2. When the operating button is pressed to the first stop, the piston expels the
same olume of air as indicated on the olume setting.
3. After immersing the tip into the liquid, the operating button is released.
This creates a partial acuum and the specified olume of liquid is aspirated
into the tip.
4. When the operating button is pressed to the first stop again, the air dispenses
the liquid. To empty the tip completely the operating button is pressed to the
second stop (blow out).
2.
4.
1.
3.
6
Positive displacement pipetting
Positi e displacement pipetting is used for applications like PCR and other DNA
amplification techniques. The microsyringe tips used in positi e displacement pi-
pettes are disposable. This helps to a oid sample-to-sample cross-contamination
(also known as sample carry-o er), and contamination due to the aerosol effect.
• Finnpipettes for positive displacement pipetting:
Finnpipette Stepper, Finnpipette PDP
• Finntips for positive displacement pipetting:
Finntip Stepper, Finntip PDP
How does a positive displacement pipette work?
1. The piston mo es to the appropriate position when the olume is set.
2. When the operating button is pressed to the first stop, the piston descends
to the tip opening.
3. After the tip is immersed into the liquid, the operating button is released.
The plunger is then raised and a partial acuum is created. This causes
the liquid to enter the tip.
4. When the operating button is pressed again, the piston descends, expelling
the liquid from the tip.
1. 2. 3. 4.
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General guidelines
and pipetting techniques
General guidelines
- Check your pipette at the beginning of your working day for dust and dirt on
the outside. If needed, wipe with 70% ethanol.
- Set the olume within the range specified for the pipette.
- Hold the pipette so the ‘grippy finger rest’ rests on your index finger.
- To maximise accuracy, the pipette, tip and liquid should be at the same
temperature.
- Check that you are using tips recommended by the manufacturer. To ensure
accuracy, use only high-quality tips made from contamination-free
polypropylene.
- Tips are designed for single use. They should not be cleaned for reuse as their
metrological characteristics will no longer be reliable.
- Pre-rinsing (three - fi e times) the tip with the liquid to be pipetted impro es
accuracy, especially when using positi e displacement tips.
- A oid turning the pipette on its side when there is liquid in the tip. Liquid might
go to the interior of the pipette and contaminate the pipette.
- A oid contamination to or from fingers by using the tip ejector.
- Always store pipettes in an upright position when not in use. Finnpipette
stands are ideal for this purpose.
- Check calibration regularly, depending on the frequency of use and on the
application, but at least once a year. If used daily, a three-month inter al is
recommended. Follow the instructions for recalibration in the instruction
manual (see also page 17).
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Forward pipetting
1. Press the operating button to the first stop.
2. Dip the tip into the solution to a depth of 1 cm, and slowly release the
operating button. Wait 1-2 seconds and withdraw the tip from the liquid,
touching it against the edge of the reser oir to remo e excess liquid.
3. Dispense the liquid into the recei ing essel by gently pressing the operating
button to the first stop. After one second, press the operating button to the
second stop. This action will empty the tip. Remo e the tip from the essel,
sliding it up the wall of the essel.
4. Release the operating button to the ready position.
Reverse pipetting
The re erse technique is used for pipetting solutions with a high iscosity or a ten-
dency to foam. This method is also recommended for dispensing small olumes.
Re erse pipetting is only possible with air displacement pipettes.
1. Press the operating button to the second stop.
2. Dip the tip into the solution to a depth of 1 cm, and slowly release the
operating button. This action will fill the tip with a olume that is larger than
the set olume. Wait 1-2 seconds and withdraw the tip from the liquid, touching
it against the edge of the reser oir to remo e excess liquid.
3. Dispense the liquid into the recei ing essel by pressing the operating button
gently and steadily to the first stop. This olume is equal to the set olume.
Hold the button in this position. Some liquid will remain in the tip, and this
should not be dispensed.
4. The liquid remaining in the tip can be pipetted back into the original solution or
disposed together with the tip.
5. Release the operating button to the ready position.
Ready position 1 2 3 4
First stop
Second stop
Ready position 1 2 3 4 5
First stop
Second stop
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Repetitive pipetting
This technique is intended for repeated pipetting of the same olume.
1. Press the operating button to the second stop.
2. Dip the tip into the solution to a depth of 1 cm, and slowly release the
operating button. Withdraw the tip from the liquid, touching it against the edge
of the reser oir to remo e excess liquid.
3. Dispense the liquid into the recei ing essel by gently pressing the operating
button to the first stop. Hold the button in this position. Some liquid will remain
in the tip, and this should not be dispensed.
4. Continue pipetting by repeating steps 2 and 3.
Pipetting whole blood
Use forward technique steps 1 and 2 to fill the tip with blood (do not prerinse the
tip). Wipe the tip carefully with a dry clean cloth.
1. Dip the tip into the blood and press the operating button to the first stop.
Make sure the tip is sufficiently below the surface.
2. Release the operating button slowly to the ready position. This action will fill
the tip with blood. Do not remo e the tip from the solution.
3. Press the operating button to the first stop and release slowly. Repeat this
process until the interior wall of the tip is clear.
4. Press the operating button to the second stop and completely empty the tip.
Remo e the tip by sliding it along the wall of the essel.
5. Release the operating button to the ready position.
Ready position 1 2 3 4
First stop
Second stop
Ready position 1 2 3 4 5 6
First stop
Second stop
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Recommendations for pipetting
different compounds
Solution/compound
Aqueous solution
Viscous solution
Volatile compounds
Nucleotide solutions
Radioactive compounds
Acid/alkalis
Toxic samples
Examples
Buffers, diluted salt solutions
Protein and nucleic acid solutions,
glycerol, T een 20/40/60/80
Methanol, hexane
Genomic DNA, PCR products
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Carbonate,
3
H-thymidine
H
2
SO
4
, HCl, NaOH
Pipette
Air displacement
Air displacement
Positive displacement
Positive displacement
Air displacement
Air displacement
Positive displacement
Air displacement
Positive displacement
Air displacement
Positive displacement
Tip
Standard
Standard or ide orifice
Positive displacement
Positive displacement
Filter
Filter or ide orifice
Positive displacement
Filter
Positive displacement
Filter
Positive displacement
Pipetting
technique
For ard
Reverse
For ard
For ard
For ard
For ard
Comments
Pipette slo ly to avoid bubble formation.
Pipette rapidly to avoid evaporation. Carbon filter tips
prevent vapor going into the pipette very effectively
For genomic DNA ide orifice tips can be used to
eliminate mechanical shearing.
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