PIPETTING TECHNIQUES  Time-consuming for 30 minutes, or after 4 hours reinactivated by

Pipettes – for the quantitative transfer of reagents and  Can be unreliable
the preparations of serial dilutions of specimens such as
serum.
CHECK YOUR SPECIMEN BEFORE USING THE
PIPETTE
 Lipemia, hemolysis, and bacterial
contamination can make the specimen
unacceptable
 Icteric (presence of excess bilirubin in the blood
stream) or turbid serum may give valid results or
may interfere
 Contamination with alkali or acid must be
avoided
COLORS TO REMEMBER AND WHAT TO EXPECT
 Icteric sample - dark to very dark yellow; brownish
like; expect patients to have a very high bilirubin or
ALT /SGPT
 Lipemic sample – turbid to milky consistency of the
sera; expect patients to have a very high cholesterol
and triglycerides
 Hemolyzed sample – red; destruction of red blood
cells --- REPEAT COLLECTION
MANUAL PIPETTING
 Hand pipetting using fixed volume is fast in small
applications and only requires the hand of a
practiced lab tech instead of extra hardware
 Repetitive actions can lead to injury
 5-10 samples per hour
PIPETTING WITH MANUAL PIPETTES
1. Check the pipette to ascertain its correct size, being
careful also to check for broken delivery or suction
tips.
2. Wearing protective gloves, hold the pipette lightly
between the thumb and the last three fingers,
leaving the index finger free
3. Place the tip of the pipette well below the surface of
the liquid to be pipetted
4. Using mechanical suction or an aspirator bulb,
carefully draw the liquid up into the pipette until the
level of liquid is well above the calibration mark
5. Quickly cover the suction opening at the top of the
pipette with the index finger
6. Wipe the outside of the pipette dry with a piece of
Kim-Wipe tissue to remove excess fluid
7. Hold the pipette in a vertical position with the
delivery tip against the inside of the original vessel.
Carefully allow the liquid in the pipette to drain by
gravity until the bottom of the meniscus is exactly at
the calibration mark. To do this, do not entirely
remove the index finger from the suction hole end of
the pipette; rather, by rolling the finger slightly over
the opening, allow slow drainage to take place
8. While still holding the pipette in a vertical position,
touch the tip of the pipette to the inside wall of the
receiving vessel. Remove the index finger from the
top of the pipette in a vertical position for correct
drainage. In TD (to deliver) pipettes, a small amount
of fluid will remain in the delivery tip
9. To be certain that the drainage is as complete as
possible, touch the delivery tip of the pipette to
another area on the inside wall of the receiving
vessel
ALWAYS BE MINDFUL OF THE MENISCUS
 The meniscus is the curvature in the top surface of
a liquid
 All readings must be made with the eye at the level
of the meniscus
The greatest potential hazard is when MOUTH
PIPETTING is done instead of mechanical suction.
Mouth pipetting is never acceptable in the clinical
laboratory.
SEMI-AUTOMATIC PIPETTES
 Offers a way for labs to incrementally scale up
production and increase reproducibility
 Only requiring the technician to move a hand probe
from vessel to vessel
 11-100 samples per hour
AUTOMATION
 Most valuable in high-throughput applications that
benefit from completely removing human
movements
 Can process hundreds of samples at a time and
follow highly complex methods without deviation
 Protection from hazardous/infectious samples
KEEP IN MIND!!!
 Bubbles and viscous solutions can cause problems
with the measurement and delivery of samples and
solutions
 BUBBLES = WRONG PROCEDURE HANDLING
 If some procedures require inactivated serum,
complement can be activated by heating to 56o C
heating for 10 minutes
TYPES OF PIPETTE
Transfer pipette – disposable plastic pipettes used to
transfer volume small volumes of liquids
Volumetric pipette – “to deliver” (TD) types that
have the bulb closer to the center and
accurately deliver a fixed volume of aqueous
solution
Ostwald Folin pipette – TD types that have the
bulb closer to the delivery tip because they
deliver viscous fluids. These pipettes deliver
an accurate volume by being “blown out”
using a pipetting bulb
Graduated pipette – is a pipette with its volume, in
increments, marked along the tube. It is used to
accurately measure and transfer a volume of liquid from
one container to another; tapered end and graduation
marks on the stem
Serological pipette – the orifice or tip opening
is larger in the serologic pipette than in other
pipettes. The rate of fall of liquid is much too fast
for great accuracy or precision; calibrated to the
tip and must be “blown out” to deliver entire
volume. The need to blow out is indicated by the
etched rings at the top of the pipette

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 Mohr pipette – calibrated between marks; PROPER PIPETTING TECHNIQUES
cannot be blown out TO MAXIMIZE PRECISION AND MINIMIZE
CONTAMINATION

1. Adjust pipette to the correct volume

2. Check if you are using the correct pipette tip
3. Make sure no bubbles are produced
4. Always use the pipette in a vertical position
5. No reusing of pipette tips
COMMON PIPETTING ERRORS
1. Loose pipette tip
Micropipette - allow rapid repetitive measurements
2. Tilting the pipettor
and delivery of predetermined volumes of reagents and
3. Plunger quick release (will cause air bubbles)
specimens; piston-operated devices that allow
4. Second stop draw
repeated, accurate, reproducible delivery of specimens,
reagents, and other liquids requiring measurement in
small amounts

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