CLINICAL LAB SCIENEC

CHAPTER 8: PIPETTING AND USE OF GLASSWARE 201
• When aspirating a liquid, the tip of the pipette must be submerged
3 to 4 mm below the surface of the liquid, at approximately a
90-degree angle.
• When dispensing a sample, a pipette should be held at a 45-degree angle
with the tip placed against the side of the receiving vessel. Glass vessels
are preferable as the surface tension of the glass enhances the complete
emptying of the tip, avoiding some errors of measurement and resulting
in greater accuracy of testing.
• The tip must never be wiped off or blotted in any way, even from the
exterior, while liquid is in the tip. These actions tend to attract and
thus bleed off some of the liquid, resulting in decreased accuracy and
reproducibility.
• A dry tip should be moistened for each aspiration by drawing up and
dispensing the chosen volume a minimum of three times. This action
reduces the surface tension on the inside walls of the tip and also
provides the proper level of intertip humidity, which reduces evaporation
of the sample liquid.
• Most pipettes are calibrated “to deliver” (TD) and not “to contain”
(TC). If the pipettes are TD pipettes, they should not be rinsed following
the delivery of their contents. If the pipette is labeled TC, it should
be rinsed to obtain the correct amount of material. If the fluid to be
measured is quite viscous or sticky (such as glycerol solutions), the
pipette must be calibrated, and in this case the outside of the tip must
be carefully wiped with lint-free tissue to remove the liquid adhering to
the tip. Care must be exercised not to touch the opening of the pipette
tip, a process that may require some practice. Accuracy in delivering
liquids with high or low viscosity may require a “positive displacement”
or piston-driver type pipetter, which is quite distinct from an air
displacement pipetter.
• To achieve maximum accuracy, and especially when calibrating
the pipette, relative humidity in the room environment should be
maintained between 50% and 75%, and in no case should the humidity
be allowed to drop below 50%. Maintaining the correct humidity
limits the rate of sample evaporation, which can cause significant
errors, especially at lower volumes. Sample evaporation is a problem
in dry climate areas and in winter months when humidity is lower
than at other seasons. Humidifiers may be necessary to provide for an
environment conducive to an acceptable accuracy level for calibrating
sensitive pipetters.
Factors Leading to Errors in Pipetting Procedures
The importance of fastidious operator skill cannot be overstated. A high-quality,
well-calibrated pipette in the hand of an uninterested or untrained operator is
an unreliable instrument. Four basic factors can negatively affect the accuracy
and reproducibility of even highly trained laboratory workers, and these factors
must be allowed for or minimized to achieve optimal accuracy.
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