a sample for delivery. Pre-wetting the tip influences

accuracy by increasing the humidity within the

tip, minimizing evaporation of the solution. This is

Serology/Immunology 593

particularly important for solvents with high vapor

pressure.

3. Work at room temperature: Allow liquids to ambient

temperature. Make sure the tips and solution are at

the same temperature.

4. Use consistent plunger pressure and speed: Depress

and release the plunger smoothly and consistently for

each sample. The fatigue and strain caused due to this

is minimized in new generation pipettes due to their

ergonomic design.

5. Use the correct pipette tip: Securely attach a tip

designed for use with the pipette. Pick out fresh and

uncontaminated tips only. Do not reuse pipette tips. If

the shape of the pipette is disfigured, discard and use a

fresh one.

6. Storage and maintenance: Always store pipettes in an

upright position when not in use. Pipette stands are

ideal for this purpose. Check calibrations regularly,

and follow instructions of the manufacturer for recalibration. Proper care and maintenance of the pipette

will ensure a longer life.

Criteria for Choosing the Right Pipette

In order to maximize the accuracy and reproducibility

of volume delivery using micropipette, it is critical to

evaluate all of the components comprising the volume

delivery system. The choice of pipette, the selection of the

most appropriate instrument of the application, should

be based on prioritizing various criteria characterizing

instrument performance. The choice of pipette tip is also

critical to performance. The following is a list of criteria

that can be used in choosing a pipette.

Accuracy and Reproducibility

The following are problems to watch out for when

reviewing pipette performance:

¾ Pipettes with advertised ranges that exceed the

performance tolerance provided by the manufacturer

¾ Pipettes whose tolerances are too tightly specified;

these will have trouble meeting the manufacturer’s

specified tolerance limits

¾ Mulltichannel pipettes with tolerances that are

interchannel statistics as opposed to intrachannel

statistics.

Durability

A pipettes’ durability is primarily a function of the

sturdiness of its components. In general, the thicker the

plastic the more durable the pipette.

Ergonomics

Whatever pipette an end-user chooses, it is critical that

the end-user feels comfortable pipetting for the extended

periods of time typical of much of lab use. If a pipette is

too large for the end-users hand it is extremely likely that

it would cause repeat motion related injury to the hand.

In addition, it would be more difficult for the end-user

to develop proper technique that would deliver accurate

results with that pipette.

Specific Applications

There are several types of pipettes designed for specific

applications. For example–autoclavibility. It is important

to check for the following information:

¾ Is the entire pipette autoclavable, or are only some

parts autoclavable?

¾ What are the recommended conditions for autoclaving?

¾ Can the plastic used for the pipette’s body, shaft and tip

cones can withstand exposure to UV light?

¾ What are the chemical compatibilities and

incompatibilities of the pipette?

Quality of Product Support

It is important to know:

¾ How supportive the manufacturer and or the distributor

of the pipette are?

¾ How responsive is customer service on warranty issues?

¾ How knowledgeable is the technical staff in terms of the

mechanics and technical specifications of the pipette?

¾ How accessible is the manufacturer for visits?

Use of Multiple Brands of Pipettes

There are two major issues:

1. The need to train technical staff on each type of pipette

separately. Different brands may use different designs

for the pipetting mechanisms requiring differences

in pipetting technique. These pipettes may require

the application of different amounts of force while

pipetting which is a skill that requires training and

repetition to acquire.

594 Concise Book of Medical Laboratory Technology: Methods and Interpretations 2. Stocking of variety of pipettes. Many labs try to stock

a single tip for all brands. Unfortunately the choice of

single tip ends up in a compromise given the variety

of shapes and plastic compositions of tips.

STREPTAVIDIN-BIOTIN SYSTEMS

Streptavidin-Biotin Systems, Better than Traditional

Antibody Capture Systems

Streptavidin-Biotin Based IEMA Systems use a biotinylated

antibody (biotin-labeled 1st antibody/capture). This is

because biotin can be attached to the FC portion of an

antibody in relatively high proportion without loss of

immunoreactivity.

The binding ratio of Avidin to Biotin is 4:1. One molecule

of Streptavidin, which is a tetramer can bind with four

molecules of Biotin/Biotinylated 1st Antibody.

In a traditional enzyme immunoassay, a limited space

is normally available for coating the Capture/1st Antibody

in the bottom of the microwell/plastic tube.

Ideally, if one can increase the number of Capture/1st

Antibodies coated on the microwell, the assay sensitivity

goes up because more number of Antigen binding sites

are available in case of low concentration of analytes

(Antigens) present in the sample.

Streptavidin–biotin based systems coat streptavidin

on the microwell/plastic tubes instead of directly coating

the capture/1st antibody. Capitalizing the tetrameric

valency of streptavidin to biotin, each molecule of coated

streptavidin binds with four molecules of biotinylated

capture/1st antibody thus providing an excess of binding

sites to the system, which ensures four fold higher

sensitivity of the IEMA system.

In other words, the streptavidin-biotin system helps to

increase the number of binding sites and thus increasing

the chances and probability of binding an antigen to an

antibody by four fold.

Streptavidin possess greater electrostatic attraction for

the microwell/plastic tubes.

Streptavidin/avidin is more inert in assay systems.

Why Streptavidin-Biotin Based Lema Systems are a

Better Choice for Tropical Laboratories?

Stability: The binding of avidin and biotin is not disturbed

by extremes of salt, pH or temperature.

Specificity and sensitivity: Avidin has a very high binding

affinity for biotin and so the system avidin-biotin is highly

specific moreover the rate constant for the avidin-biotin

association is also fast.