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Improving Pipetting Techniques For better accuracy and performance. Pipetting 360 °. Part of the Good Pipetting Series of Seminars. Improving Pipetting Techniques. Definitions Minimizing errors Hands-on Workshop (optional). Air-displacement Pipette. Piston. Shaft. Air space. Disposable

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improving pipetting techniques for better accuracy and performance

Improving Pipetting TechniquesFor better accuracy and performance

Pipetting 360°

Part of the Good Pipetting Series of Seminars

improving pipetting techniques
Improving Pipetting Techniques
  • Definitions
  • Minimizing errors
  • Hands-on Workshop (optional)
air displacement pipette
Air-displacement Pipette

Piston

Shaft

Air space

Disposable

tip

Sample

air displacement pipetting cycle
Air-displacement pipetting cycle

Depress

Hold

Tip Ejection

Aspirate

Dispense through Blowout

pipette performance specifications
Pipette performance specifications
  • Accuracy +/- 1%
  • Precision 0.25% to 0.33%
    • Ultramicro pipette specifications are wider

Precise but not accurate

Not precise or accurate

Preciseandaccurate

basic techniques for minimizing errors
Basic techniques for minimizing errors
  • Optimizing volume range
  • Setting the micrometer
  • Tip immersion angle
  • Tip immersion depth
  • Tip immersion time
  • Pre-rinsing pipette tips
  • Aspiration rate
  • Dispensing techniques
  • Hand-warming effects

Errors from poor technique can range from 0.1% - 5% or more

impact of errors in technique
Impact of errors in technique
  • Errors less than 0.5%
    • Calibrators need to be aware as all errors can impact results
  • Errors 0.6% - 1%
    • Normal users need to decide the importance of each error versus the extra time and effort required
  • Errors greater than 1%
    • Everyone needs to be aware of these errors
optimizing volume range
Optimizing volume range
  • Normal Range
    • 10% - 100% of volume
      • Operating at 10% range requires good technique
  • Optimized Range
    • typically 35% - 100% of volume
      • Less technique dependent
      • Assures accuracy and precision

Optimizing volumerangetypically improves accuracy up to 1%

setting the micrometer
Setting the micrometer
  • Approach each volume in the same direction each time
  • Turn micrometer 1/3 revolution above desired

volume

  • Dial down to volume setting

Correctly setting the micrometer improves accuracy up to 0.5%

tip immersion angle
Tip immersion angle

Incorrectimmersionangle

Correctimmersionangle

Aspirating with the pipetteperpendicular improvesaccuracy up to 0.5%,ultramicroup to 2.5%

tip immersion depth

Depth

Tip immersion depth

Recommended immersion depth improves accuracy up to 1%, ultramicro up to 5%

tip immersion time for macrovolume pipettes
Tip immersion time for macrovolume pipettes
  • Maintain tip immersion time of 1 second (minimum) after aspiration
  • Withdraw tip slowly, smoothly from liquid source
  • Important for large volume samples and viscous liquid samples
effect of pre rinsing tips

100.2

100.1

10 model

Normalized Volume

(%)

200 model

100.0

1000 model

99.9

99.8

0

1

2

Number ofPre-rinses

Effect of pre-rinsing tips
pre rinsing pipette tips
Pre-rinsing pipette tips
  • Pre-rinse tip with same liquid that is being dispensed
  • Aspirate sample into tip, and then dispense back into reservoir or to waste
  • Pre-rinsing provides identicalcontactsurfaces for all aliquots

Two pre-rinses provides up to 0.2% greater accuracy

when used with aqueous liquids

aspiration rate effects
Aspiration rate effects
  • Use consistent
    • Pipetting rhythm
    • Pressure on plunger
    • Speed and smoothness

Inconsistentaspirationcan affect accuracy up to 1%

aspiration rate effects1
Aspiration rate effects
  • Use consistent
    • Pipetting rhythm
    • Pressure on plunger
    • Speed and smoothness
  • Aspiration too quickly
  • Liquid splash-up into shaft damaging piston and seal
  • Introduces aerosols and sample cross-contamination

Tooquickaspirationcanaffectaccuracyupto5%ormore

dispensing techniques
Dispensing techniques
  • Thin-wall, FinePointTMtips provide maximum droplet dispensing
  • Threetechniques
    • Alongside-wall
dispensing techniques1
Dispensing techniques
  • Thin-wall, FinePointTMtips provide maximum droplet dispensing
  • Three techniques
    • Along side-wall

2) Above vessel / liquid surface

dispensing techniques2
Dispensing techniques
  • Thin-wall, FinePointTMtips provide maximum droplet dispensing
  • Three techniques

1) Along side-wall

2) Above vessel/ liquid surface

3)Directlyintoliquid

Correctdispensingtechniqueimprovesaccuracyupto1%

hand warming effects
Hand-warming effects

Prolongedhand-warmingintroduceserrorsofupto0.2%for a high-quality pipette, 0.5% for others

improving pipetting techniques summary
Improving pipetting techniques summary
  • Optimizing volume range
  • Setting the micrometer
  • Tip immersion angle
  • Tip immersion depth
  • Tip immersion time
  • Pre-rinsing pipette tips
  • Aspiration rate effects
  • Dispensing techniques
  • Hand-warming effects
errors in pipetting
Errors in pipetting

Small errors are important to calibration technicians, everyone should care about large errors.

slide23

Good Pipetting Technique =

Minimal Errors =

Good Performance

Rainin Instrument, LLC

a METTLER TOLEDO Company

  • 800-472-4646
  • www.rainin.com
  • pipets@rainin.com