[CC LAB] L1: Pipetting

1. reconstituting controls and calibrators

2. preparing serum plasma dilutions

3. aliquoting specimens

what are pipettes used for (3)

Serologic pipettes

what pipette to use when the volume is 1-100mL

automated micropipettes or semi automated pipettes

what pipette to use when the volume is 1uL - 1mL

graduations

Lines marked on volume measuring devices that indicate volume.

meniscus

A curve formed by the surface of liquids confined in a narrow space, such as in a measuring device.

lower meniscus

which part of the meniscus will you read if clear & colorless fluid

upper meniscus

which part of the meniscus will you read if colored fluid

eye level

When reading the meniscus in the laboratory it should be ________ to avoid errors.

tolerance

How much error is allowed in the calibration or a measuring device.

accuracy

the _____ of the readings of the measurements is based on tolerance

lower tolerance limit (class A) because it reduce the range and more precised

which one is better? the higher tolerance limit or lower tolerance limit?

1. NIST - National Institute of Standards and Technology

2. ASTM - American Society for Testing and Materials

what are the organization groups responsible for the acceptance of pipettes

National Bureau of Standards (NBS)

NIST is also known as

ASTM E542

ASTM E969

Each pipette is calibrated in accordance with _______ and meets accuracy requirements of _________.

American Society for Testing and Technology

regardless of the design, most laboratory supplies must satisfy certain tolerances of

accuracy and fall into two classes of precision tolerance (Class A or Class B) as given by what organization?

National Institute of Standards and Technology

Including pipets, is manufactured and calibrated to deliver the most accurate volume of liquid.

• Defined by

College of American Pathologists (CAP)

According to what organization? - Volumetric pipets must be of certified accuracy (Class A)

swift delivery

S in the label of pipette stands for?

1. caulfield

2. spectroline

3. safety bulb

1. calibration Marks / Designs

2. Draining Characteristics

3. General Types / Categories

what are the 3 pipetting classifications?

1. To Contain (TC)

2. To Deliver (TD)

what are 2 types of calibration marks / designs?

1. Blow out

2. Self-Draining

what are 2 types of draining characteristics?

1. volumetric / transfer

2. measuring / graduated

what are 2 types of General types / categories

To Contain (TC)

hold or contains a particular volume but does not dispense the volume indicated

To Conta

Mercury

what is the calibrating medium of To Contain pipets

To Contain (TC)

micropipettes, long levy type, and sahli’s pipette are examples of what design

To Deliver (TD)

will dispense the volume indicated

gravity

To Deliver is designed to drain by ______

Distilled water

what is the calibrating medium of To Deliver (TD) pipet

To Deliver (TD)

Serological, Mohr pipette, Volumetric pipettes, Ostwald-Folin pipettes are examples of what pipet design?

blowout

Has 2 continuous rings located near the top of the

pipette or the area where we attach the pipette bulbs or

the safety bulbs for suction of the fluid.

blowout

The last drop of liquid should be expelled into the

receiving vessel through blowing out.

blowout

Serological and Ostwald-Folin pipettes are example of what draining characteristic

Self-draining

No ring markings located at the top of the pipettes. Draining is by gravity. what draining characteristic?

Self-Draining

Mohr pipette, Volumetric pipettes, Van Slyke pipettes are examples of what draining characteristic?

Volumetric / transfer pipets

Shaped like rolling pins with a large belly, one blunt end → the neck, one tapering end → the tip.

transfer pipette

what general type of pipet delivers only one exact volume, and they are used when a higher degree of accuracy is required

Transfer pipet

Used for preparation of standard solutions that are used (ex: in calibration)

measuring / graduated pipets

Long, cylindrical tubes drawn out to a tip and are calibrated in uniform fractional volume measurement.

measuring / graduated pipets

Mohr & Serologic Pipet are examples of what general type of pipet

measuring / graduated pipets

These pipettes can deliver multiple volumes, and are used when less accuracy of measurement is required

measuring / graduated pipets

Used for delivering of reagents or small amount of solvents, and however because of less accurate measurement for this pipet, this type of pipette should not be used for making standard solutions or quality control materials.

1. To deliver

2. Blowout

3. Measuring / graduated

serologic pipet

1. design -

2. draining characteristic -

3. general type -

1. To Deliver

2. Self-draining

3. Measuring / Graduated

Mohr

1. design -

2. draining characteristic -

3. general type -

measuring / graduated

bacteriologic

general type -

measuring / graduated

ball, Kolmer, or Kahn

general type -

measuring / graduated

micropipette

general type -

1. To Deliver

2. Self-draining

3. Transfer / volumetric

volumetric

1. design -

2. draining characteristic -

3. general type -

1. To Deliver

2. Blowout

3. Transfer / volumetric

Ostwald-Folin

1. design -

2. draining characteristic -

3. general type -

Transfer / volumetric

Van Slyke Pipette

general type -

Transfer / volumetric

Sahli’s pipette

general type -

serologic pipette

Mohr

Micropipette

Micropipette

can only hold from 0.1 microliter to 1000 microliter (1mL = 1000 microliter)

Micropipette

o Sahli-Hellige pipette

o Lang-Levey pipette

o RBC and WBC pipettes

o Kirk and Overflow pipette

these are examples of ___ pipette

volumetric

used for non-viscous fluids

Volumetric

Have been used to add the diluent to a lyophilized control (freeze dried/powdered control) or to measure standards and controls.

Ostwald-Folin

what transfer pipette - has the belly located at the center

Ostwald-Folin

the bulb is closer to the delivery tip, reducing the surface area in contact with the liquid

Ostwald-Folin

used for viscous fluids (whole blood, serum)

Van Slyke Pipette

Thick-walled capillary tubing with bulb in the center.

Sahli’s pipette

Can deliver 20 ul (microliters) of the aliquot.

Used in hematology, hemoglobin measurements and determination.

Volumetric pipette

Ostwald-Folin Pipette

Van Slyke Pipette

Sahli’s pipette

Mohr’s pipette

Volumetric pipette

Serologic pipette

Ostwald-Folin Pipette

Automated / Semi-automated micropipette

Lambda pipette / Dropping pipette

left - single channel

right - multichannel

what type of semi-automated pipette?

left -

right -

0.1uL to 1000uL

what volumes are used for semi-automated pipette?

semi-automated pipettes

offer more convenience and efficiency to pipetting. no pipetting bulb is required nor do pipettes have to be washed. only need to replace the pipetting tips every time you aspirate fluids.

propylene

what plastic tips are used in semi-automated pipette

semi-automated pipette

Advantages:

o Safety

o Stability

o Ease of use

o Increased precision

o Ability to save time

o Less cleaning required.

what pipette?

1. plunger

2. tip ejector button

3. friction ring

4. body

5. connecting nut

6. tip ejector

7. tip holder

8. tip cone

9. pipette tip

plunger

Press and release to aspirate, decant, or dispense the fluid.

tip ejector button

Used, by clicking or pushing, to discard the tip.

tip ejector

Once pushed, it will push the tip to discard.

tip cone

Where you attach the pipette tips.

variable volume micropipette

● Change or adjust the volumes.

● From 100 to 1000 microliters (based in the picture.).

● Single channel micropipette

Fixed volume micropipette

● Can only give a fixed volume of micropipette.

● If it has 1000 microliters, it can only dispense 1000 microliters or 1 mL.

● There are also other types like 200 microliters, 20 microliters, 10 microliters, so on and so forth.

● Single channel micropipette

Multichannel micropipette

● Has a lot of tips.

● It can accommodate more than one pipet tips.

1. air displacement pipette

2. dispenser / dilutor pipette

3. positive displacement pipette

3 examples of mechanical or automatic pipettes

air displacement pipette

● Relies on the piston for suction creation (with the use of air) to draw the sample into a disposable tip.

● The most common principle applies.

dispenser / dilutor pipette

● Obtain the liquid from a common reservoir and dispense it repeatedly.

Positive displacement pipette

● Operates by mobbing the piston in the pipette tip or barrel, much like a hypodermic needle.

○ It suctions the fluid with the use of its piston in the pipette tip or barrel.

● Does not require a different tip for each use.

● Recommended for saline, water, and phosphate buffers.

● Not suitable for very dense liquids.