Agilent Technologies PlateLoc User manual


PlateLoc Thermal
Microplate Sealer
Quick Reference Guide

Notices
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User Guide Part Number
G5402-90003
June 2009 rebranded edition
May 2008 quick reference guide edition
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Automation Solutions
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,

1
PlateLoc Thermal Microplate Sealer
Reference for Optimal Thermal Microplate Sealing

PlateLoc Thermal Microplate Sealer
Reference for Optimal Thermal Microplate Sealing
This reference contains the following topics:
•“Optimizing the sealing quality” on page 2
•“Optimizing seal parameters” on page 5
•“Requesting application support” on page 8
•“Frequently asked questions” on page 9

PlateLoc Thermal Microplate Sealer
About this guide
2Reference for Optimal Thermal Microplate Sealing
About this guide
This guide provides a quick- reference for using the PlateLoc Thermal
Microplate Sealer (PlateLoc Sealer). Use the following guidelines to identify
thermal- sealing- friendly labware and optimize microplate sealing using the
PlateLoc Sealer. You should use this guide in conjunction with the following:
•PlateLoc Thermal Microplate Sealer User Guide
•PlateLoc Seal Selection Guide
You can download the latest version of any PDF file or search the online
knowledge base from the Automation Solutions Knowledge Base page.
For more information about automation products, go to the Agilent
Technologies website at www.agilent.com/lifesciences/automation.
Optimizing the sealing quality
Introduction
The PlateLoc Sealer applies a sealing material on top of microplates to seal
individual wells. Sealing the wells protects the contents from evaporation,
condensation, oxidation, and cross- contamination during transport or storage.
The PlateLoc Sealer can be used as a standalone device or in a lab automation
system. The device accepts microplates made from a variety of materials.
The ideal sealing conditions can depend on a number of factors, especially the
labware. Using thermal seal- friendly microplates is key to protecting samples
and critical to successful sample sealing and storage applications.
Microplate sealing tips
The seal quality depends on many factors, including microplate design,
material, lot- to- lot consistency and features, the sealing parameters, whether
the microplates require an insert for support, and the type of seal used.
Microplate design
Automation- friendly and thermal- seal- friendly labware ensures successful
sealing.
• Use only microplates that comply with standards established by the Society
of Biomolecular Sciences (SBS).
• Raised chimneys around each well produce the best results.
• Avoid microplates that have a raised rim close to the perimeter of the
wells. The rim can prevent uniform contact between the hot plate and the
wells.
Figure 1 illustrates these features.

3
PlateLoc Thermal Microplate Sealer
Optimizing the sealing quality
Reference for Optimal Thermal Microplate Sealing
Figure 1 Microplate physical features and nomenclature
Figure 2 Left: Pneumatics drive the hot plate down to the microplate surface. The hot plate can
sense and automatically adjust for variations in microplate height. Seals are applied using
pressure and heat. Right: If the microplate has a raised rim that is higher than the chimney height
around its perimeter, the hot plate must fit within the rimmed area so that the hot plate can make
uniform contact with the seal and all sample wells simultaneously.
Microplate material
For the best sealing results:
• Use polypropylene microplates because they are the easiest to seal
effectively and reliably.
• Avoid acrylic microplates because they are not compatible with thermal
microplate sealing.
• If the microplates are flexible and tend to bend and move during the
sealing process, use adapter trays to stabilize the microplate position on
the plate stage. For example, place a flexible 96- well polypropylene
microplate on top of a 96- well polystyrene microplate to provide positional
support. Agilent Technologies also offers an adapter tray for PCR
microplate support.
Raised rim around
perimeter (if any) Sample well
Skirt
Plate
surface
Chimney: Top of well rises
beyond microplate surface
Insert
HOT PLATE
HOT PLATE
Microplate with
raised rim
Sealing film
Hot plate

PlateLoc Thermal Microplate Sealer
Optimizing the sealing quality
4Reference for Optimal Thermal Microplate Sealing
Figure 3 Left: Microplate and PCR Plate Support. Right: Microplate in PCR Plate Support.
Microplate make and model
• Be aware that similar microplates made from the same material but from
different manufacturers may perform differently.
• Evaluate every microplate make and model to ensure consistent results.
Microplate features
Be aware of design features on the top surface of the microplate that can
significantly affect the quality of the seal.
• The microplate lacks raised chimneys and it has molded branding or
legends, such as the manufacturer’s name or logo, or the sample locating
legends (for example A, B, C, . . . and 1, 2, 3 . . .) close to the tops of the
wells (close enough so that seal touches or covers the imprint).
• Tooling marks or manufacturing aids, such as circular depressions
(sometimes with branching) caused by ejection pins or part of the molding
process. Gaps or gutters may not only prevent binding of the seal, but may
actually facilitate cross- contamination of liquid samples.
Sealing parameters
The following parameters can be adjusted to optimize the sealing results:
•Temperature
•Time
Perform optimization tests to determine the best values to use. For details, see
“Optimizing the sealing parameters (temperature versus time)” on page 10.
Insert thickness
Choose the right insert to support the microplate and ensure uniform sealing.
A set of four inserts are presently shipping with every PlateLoc Sealer, and a
special insert is available for Labcyte microplates.

5
PlateLoc Thermal Microplate Sealer
Optimizing seal parameters
Reference for Optimal Thermal Microplate Sealing
Figure 4 Microplate without and with insert
Figure 5 Standard inserts. The number on the inserts indicates the thickness of the metal pad
(for example, 180 means 0.180 in thick). Note: The 90 insert has a 0.90-in metal pad with foam
padding on the microplate-facing side and is used with microplates that require flexible support.
Seal material
Different seal materials have different properties and can produce different
results. Perform optimization tests to determine the best seal to use.
Optimizing seal parameters
About this section
This section explains how to inspect the seal quality and, if necessary, how to
adjust the sealing parameters to improve the seal. Before proceeding, download
a copy of the PlateLoc Seal Selection Guide from the Agilent Technologies
website at http://www.chem.agilent.com/en-us/products/instruments/automation/
microplatehandling/plateloc/pages/default.aspx.
Apply a seal using the time and temperature starting points suggested in the
PlateLoc Sealer Selection Guide, for your chosen seal and microplate material.
Examine the well impressions made on the seal. The pattern and quality of
these impressions is a good indicator of seal quality.
Evaluating the seal
After the seal cycle is finished, wait for the microplate to cool down enough so
that it is safe to handle (10 to 30 seconds). Do not handle the microplate
immediately after the seal cycle is finished. The sealed microplate might be
hot.
Note: Pierceable seals might permanently adhere to the microplate. Do not wait
too long or the seals will be difficult to remove.
Remove the microplate from the PlateLoc Sealer. Carefully peel off the seal
material by lifting one of the corners of the seal as shown in Figure 6.
Bend direction
Microplate insert
Wells

PlateLoc Thermal Microplate Sealer
Optimizing seal parameters
6Reference for Optimal Thermal Microplate Sealing
Figure 6 Removing the seal.
Inspect the underside of the removed seal material. If there are unbroken
impressions of each well on the underside of the seal material, the microplate
was properly sealed.
Figure 7 Example of a good seal.
Use a magnifying glass or inspection microscope, if necessary, to see the area
surrounding the sample well rim.
Look closely at a single sample well. A good seal might look like the
illustrations on the left side of Figure 8. The well impression should match the
thickness of the chimney wall, as shown in the left side of Figure 9. A poor
seal might look like the illustrations on the center or right side of Figure 8,
indicating there may not have been enough heat applied to form a complete
impression of the sample well rim.
Figure 8 Evaluating the seals of sample wells. L: Good impression, with defined edge of
chimney/rim. C and R: Poor impression, with missing or very faint segments.
If there are faint or broken impressions in the seal material, as shown in
Figure 8, increase either the sealing temperature or time (duration).
00201 PlateLoc sealed plate
00201 PlateLoc sealed plate

7
PlateLoc Thermal Microplate Sealer
Optimizing seal parameters
Reference for Optimal Thermal Microplate Sealing
If the well impressions on the seal are significantly thicker than the chimney
wall (as shown in Figure 9), decrease the sealing temperature or duration.
Thick impressions may indicate excessive melting of the seal, causing the seal
to be difficult to remove and the resealing capacity reduced.
Figure 9 Evaluating rim width of seal. L: Good impression, with thin, defined edge of chimney/
rim. R: Poor impression, with increased width.
Issues related to inserts
The hot plate is a heated metal surface that presses the seal onto the
microplate. For optimal sealing, the microplate needs to be firmly supported so
that it does not bend during the sealing process, often requiring use of an
insert to keep it flat. Figure 10 illustrates the effect of insert height on
microplate sealing. Left: if the insert is too thin, the center of the microplate
will bend downward during sealing, leading to good sealing around the edges,
but poor sealing in the middle of the microplate. Center: if the insert is the
correct thickness, the entire microplate surface will be sealed properly. Right:
if the insert is too thick, the edges of the microplate will bend downward
during sealing, resulting in good sealing in the middle of the microplate, but
poor sealing around the edges.
Note: Thicker inserts are preferred over inserts that are too thin.
Figure 10 The effect of using inserts on microplate sealing. L: Insert too thin; C: Insert correct
height; R: Insert too thick.
Approximately same
thickness as chimney
wall
Thicker than chimney wall

PlateLoc Thermal Microplate Sealer
Requesting application support
8Reference for Optimal Thermal Microplate Sealing
Requesting application support
To help expedite your support request, you can complete and submit the
Request for PlateLoc Sealer Application Support.
For other questions, you can contact Automation Solutions Technical Support
at one of the following:
Europe
Phone: [phone]
email: [e-mail]
US and rest of world
Phone: [phone] (US only) or +1.408.345.8011
email: [email protected]

9
PlateLoc Thermal Microplate Sealer
Frequently asked questions
Reference for Optimal Thermal Microplate Sealing
Frequently asked questions
Consumables—seal
Q
Is the PlateLoc Sealer compatible with other third- party sealing materials?
A
Agilent Technologies does not verify, validate, test, or support the use of third-
party sealing material on the PlateLoc Sealer. There is also some risk that non-
Agilent Technologies sealing material may melt onto internal (non- user-
serviceable) components, necessitating service.
Hot plate temperature verification and validation
Q
During instrument calibration, does Agilent Technologies check that the hot
plate is providing the temperature indicated on the LCD screen of the PlateLoc
Sealer?
A
We don’t. The temperature reading is the average of two thermocouple
readings, which can vary slightly from thermocouple to thermocouple. The
thermocouples aren’t calibrated because we have found little variation in their
measurements.
If the thermocouple readings differ significantly, an error message is generated.
The precision of the heating element is more important than its accuracy.
Being able to precisely adjust the temperature is more crucial to optimizing
sealing parameters than setting an accurate temperature. The same microplate
and seal can require different sealing parameters depending on the amount
and type of liquid in the microplate, the temperature of the microplate, and
the desired seal strength, so absolute temperature readings are not critical to
good sealing.
Hot plate cleaning
Q
I accidentally installed the thermal seal roll upside down (bonding layer up). I
now have seal melted on the hot plate. Are there approved cleaning
procedures? Is there a way to prevent this from happening again?
A
If Agilent Technologies installed your PlateLoc Sealer, ask for training on
cleaning the hot plate. Through routine use, the hot plate will eventually
become dirty from excess seal build- up, melted pieces of microplate, and
general dirt and debris. The PlateLoc Thermal Microplate Sealer User Guide,
which can be downloaded from the Agilent Technologies Automation Solutions
Knowledge Base, describes routine maintenance, including cleaning the hot
plate and touch screen.

PlateLoc Thermal Microplate Sealer
Frequently asked questions
10 Reference for Optimal Thermal Microplate Sealing
To eliminate the chance of mis- loading the seal roll, use the seal- loading card
that comes with every roll of Agilent Technologies seal. This card features
pictures and instructions for loading new rolls of seal. Detailed information
can be found in the PlateLoc Thermal Microplate Sealer User Guide.
Maximum sealing temperature
Q
What is the highest temperature and longest duration I can set on my PlateLoc
Sealer?
A
The maximum temperature is 235 °C. The duration can be adjusted from 0.5 to
12 seconds.
Optimizing the sealing parameters (temperature versus time)
Q
I am not getting an effective seal because the sealing material is not adhering
very well. Can I keep boosting the temperature or is it better to just increase
the time?
A
The most common mistake new users make is to start at too high a
temperature or continue increasing the temperature unnecessarily. Excessive
heat can damage the microplate and prevent future resealing by prematurely
melting the chimney surrounding the top of the sample wells. Refer to the
PlateLoc Seal Selection Guide that can be downloaded from the Agilent
Technologies Automation Solutions website. The selection guide recommends
starting temperatures based on seal type. In general, we recommend sealing at
the lowest temperature possible for the shortest time.
Resealing the same microplate
Q
We are evaluating one of Agilent Technologies peelable seals. How many times
can we seal and reseal a single microplate?
A
If you minimize the temperature and time required to seal the microplate, the
well chimney should usually last a minimum of five sealing cycles. Many more
successful cycles may be possible, depending on the seal material, microplate
material, sealing parameters, etc.
Seal cooling time
Q
We’re trying to optimize sealing parameters, so we seal a microplate, wait for
it to cool, then peel off the seal by hand and examine the well impressions on
the back of the seal. How long should we wait before peeling the seal off?

11
PlateLoc Thermal Microplate Sealer
Frequently asked questions
Reference for Optimal Thermal Microplate Sealing
A
We suggest waiting a minimum of 10 seconds or as long as it takes for the
sealing film on the microplate to return to room temperature.
Removing seal from cold storage microplates
Q
We’d like to seal our pharmaceutical microplate library, and store the sealed
microplates at –20 °C. Do we need to allow the microplates to return to room
temperature prior to peeling off the seal?
A
Yes. Sealed microplates that have been stored in a refrigerator or freezer
should be returned to room temperature before attempting to remove the seal.
It is good laboratory practice to thoroughly test your methods prior to sealing
an entire compound library. Be sure to choose a seal material that is designed
for low- temperature storage and is removable (peelable). Note that some seals
are designed to not be peeled off, but rather pierced mechanically for sample
access.
Minimizing or eliminating cross-contamination
Q
How careful do we need to be when we manually peel off the seal by hand?
What if there is liquid under the seal? Can droplets from one well contaminate
another?
A
There is always the chance that condensate or liquid from a sample well may
be transferred to the underside of the sealing film. One option is to centrifuge
the microplate before removing the seal. Another option is to not remove the
seal—use a pierceable seal. Whether you choose peelable or pierceable seal, it’s
best to centrifuge the microplate before removing samples.
Gas-purging option
Q
The samples we have are in DMSO and we plan to use the PlateLoc Sealer to
seal our microplates with Agilent Technologies Peelable Aluminium Seal PN
06643.001 prior to putting our microplates into storage at –20 °C. How might
we further reduce hydration and oxidation while sealing these microplates?
A
The Agilent Technologies gas- purging PlateLoc Sealer was developed for
pharmaceutical customers to replace the air (containing moisture and oxygen)
in their microplate wells with a dry, inert gas such as argon before the
microplates are sealed. Because of the permeability of microplate materials
such as polypropylene and polystyrene, the gas- purging effects typically last up
to 24 hours when the microplates are stored at room temperature, perhaps
longer if stored at lower temperatures.

PlateLoc Thermal Microplate Sealer
Frequently asked questions
12 Reference for Optimal Thermal Microplate Sealing
Microplate-sealing technology and automation
Q
We are overwhelmed by the technology choices currently available for sealing
microplates (thermal, adhesive, laser, cap mats, etc.). Is any one actually better
than the other?
A
The best choice is application- dependent. To determine whether a manual,
semi- automated, or automated approach would be cost- effective for your lab,
evaluate how many microplates you are sealing today and how this might
change over the next 12 months.
There are benefits to all sealing approaches. Each approach has strengths and
weaknesses that must be weighed against the cost and benefit to your specific
samples.
Thermal microplate sealing is a proven approach and is generally believed to
pose the least risk of chemical incompatibility (samples and seal material)
when compared to adhesive sealing. Thermal sealing subjects the top of the
microplates to a few degrees of heat transfer, but the heat is typically not
enough to affect most samples in life science applications. Thermal seals also
typically cost less than adhesive seals.
Adhesive microplate sealing employs pressure- sensitive adhesive films (PSAs)
and is most commonly used for thermally sensitive samples. Protein
crystallography and some cell biology applications are instances in which
customers most often use adhesive sealers.
Heat-sensitive samples
Q
We want to seal our microplates to safely store our samples, but our samples
are heat sensitive. A colleague suggested we consider adhesive sealing
technology. Exactly how much heat will our samples be exposed to if we use a
PlateLoc Sealer?
A
Most commonly, the PlateLoc Sealer is set to 130–170 °C for a duration of 1–2
seconds. Under these conditions, the temperature of the top of the microplate,
and possibly the sample, could increase a few degrees. However, this can vary
based on microplate material, microplate height, sample volume, etc. Agilent
Technologies recommends that you verify that the PlateLoc Sealer does not
adversely affect your samples.
Continuous (24/7) operation
Q
We intend to share our PlateLoc Sealer in a core lab. Is it OK to operate it
24/7?
A
The PlateLoc Sealer is robust and designed for challenging, high- throughput,
continuous- duty, production applications. We encourage you to use it for your
most demanding applications. We also encourage you to talk to other PlateLoc
Sealer customers, or consider an extended warranty.

13
PlateLoc Thermal Microplate Sealer
Frequently asked questions
Reference for Optimal Thermal Microplate Sealing
Evaporation verification and validation
Q
Do you have an established protocol we can use to test the PlateLoc seal for
permeability and evaporation?
A
A simple way to test for evaporation uses an experimental microplate and a
control such as a 100- g weight. Add an equal amount of water to all wells.
Weigh both the experimental microplate and the control. Seal the microplate.
Weigh both again. Place the experimental microplate into an oven to accelerate
the evaporation process (we suggest 65 °C for 3 hours or longer). Carefully
remove the microplate from the oven. Let it cool down to room temperature,
then weigh it again. Compare the initial and final weights of the experimental
microplate. Finally, weigh the control one more time to ensure the scale is
performing consistently.
Seal-peeling options and ergonomics
Q
We are presently hand- peeling seals from our microplates. We are now peeling
hundreds of microplates per day, and we are concerned about potential
repetitive motion injuries. What are our options?
A
Some labs choose to use pierceable seals which can be pierced using
automated piercing devices such as the Agilent Technologies Microplate Seal
Piercer. Another option is to use a microplate peeler device. These standalone
devices can be automated, and can be integrated with robotic microplate
handlers such as the Agilent Technologies BenchCel Microplate Handling
Workstation.
Short sample rolls
Q
Do you have free sample rolls of seal we can test with our microplates prior to
purchasing an entire roll of seal?
A
Contact Automation Solutions Technical Support for a wide variety of short,
sample rolls of seal for this purpose.
Low-profile microplate support
Q
We’d like to seal Aurora® 1536 low profile microplates. Is there a special
microplate support available to facilitate a good seal?
A
Yes. Contact Automation Solutions Technical Support and ask for PN 17708.001

PlateLoc Thermal Microplate Sealer
Frequently asked questions
14 Reference for Optimal Thermal Microplate Sealing
Microplate height
Q
Can low- profile high- density microplates (for example, 1536- well microplates)
or tall deep- well microplates be sealed with the PlateLoc Sealer?
A
Yes. Low- profile microplates tend to bend, so you must match the microplate
with the appropriate microplate support to prevent this from happening. The
sample well surface and the sample well bottoms should be rigid and parallel.
Inserts (see “Insert thickness” on page 4) are designed to assist in these
circumstances. The hot plate is mounted in the top of the sealing chamber and
descends to the appropriate height for sealing, automatically adjusting to the
microplate height. Tall labware, such as deep- well microplates as tall as 2.5 in
(63.5 mm), can be accommodated. High- volume storage and collection
microplates exceeding 2.5 in are not compatible with the PlateLoc Sealer (for
example, Porvair Sciences 48- well microplates of 7 and 10 mL sample volume,
etc.).
How does thermal sealing work?
Q
How does thermal microplate seal bond to the microplate? Will the glue
contaminate my samples? Will it work with DMSO or organic solvents?
A
Hot melt adhesives (HMAs, also referred to as thermoplastic adhesives) are
solid at room temperature, become molten when heated, and adhesion develops
as the melt solidifies while it cools. Most hot melt adhesives achieve 50% of the
bond strength after one minute, 75% after one hour, and 100% after one day.
Chemical resistance will vary based on the formulation and additives. For
example, for ethylene vinyl acetate copolymer (EVA), chemical resistance varies
from poor (for aromatic hydrocarbons such as benzene or halogenated
hydrocarbons), to good (for oils and greases), to very good (for dilute acids,
alkalis, aliphatic hydrocarbons, and alcohols). Confirm that any seal you use is
compatible with your samples (see next question).
Figure 11 Hypothetical thermal microplate sealing film structure.
Optional Top Coating
(e.g. Primer for Printing)
Backing / Support Layer
(e.g. Aluminum or Clear Polymer Laminates)
HMA (Hot Melt Adhesive) Layer
(e.g. EVA with additives, similar to “Hot Melt Glue Stick”

15
PlateLoc Thermal Microplate Sealer
Frequently asked questions
Reference for Optimal Thermal Microplate Sealing
Chemical compatibility verification and validation
Q
We’d like to confirm the thermal microplate seal we have selected is chemically
compatible with our samples for long- term storage. Do you have a suggested
method?
A
Please see the Association for Laboratory Automation (ALA), Final Conference
Program, January 27–31, 2001. Page 276 of the PDF contains the abstract from
a poster paper generated by 3M with useful protocols for evaluating adhesive-
based microplate seals.
www.labautomation.org/conference/pdfs/LA2001Book.pdf
Another useful reference is Comparison of Microplate Sealing Tapes Using
Standardized Test Protocols [T- 41]. Terry W. Lewis, Maurice H. Kuypers,
Mialena M. Walker Medical Specialties Department 3M Health Care, St. Paul,
Minnesota.
http://lab- robotics.org/Presentations/3M/3M_comparison4.PDF
Sterile and RNase-/DNase-free microplate seal
Q
Do you sell microplate seals that are certified to be sterile as well as RNase- /
DNase- free?
A
No, not at this time. The majority of PlateLoc Sealer customers have been able
to use the existing materials. Agilent Technologies can develop seal materials
with these features, provided the customer is willing to invest the time and
substantial cost required to do so.
There are also costly technical challenges to offering materials such as
aluminium seals in a sterile format. Since e- beam sterilization cannot penetrate
aluminum very well, gamma radiation is the only practical solution. However,
the cost is substantial. Contact Agilent Technologies for more information.
Water bath thermal cycling
Q
Are Agilent Technologies thermal microplate seals compatible with water bath
thermal cycling?
A
The aluminium seal can be used for a limited time in a water bath, and the
clear seal for longer periods. However, the hot melt adhesive layer may be
damaged by prolonged exposure to water. Also, it is very important that no air
bubbles migrate into the gap between the seal surface and the top of the
microplate (the gap is due to the raised rim or chimney of the sample well).
For example, if two microplates are held vertically together (seal surface to
seal surface) and air bubbles rise into the gap between the seal and the
microplate surface itself, the expanding bubbles may apply force to the seal
which may loosen the seal. One workaround is to place a piece of conforming
foam between the two sealed microplates, held with sufficient force to prevent
the air bubbles from entering the gap between the seal and microplate top
surface.

PlateLoc Thermal Microplate Sealer
Frequently asked questions
16 Reference for Optimal Thermal Microplate Sealing
Edge effects
Q
We are trying to seal microplates for an ELISA assay and are concerned about
potential edge effects. Any suggestions on how to reduce or eliminate the
problem?
A
Edge effects, such as variability in the sample data due to well- to- well
differences in evaporation, can be a challenge. Rather than optimize the
microplate or sealing parameters, some labs don’t use the sample wells in the
outer rows of the microplate, although this impacts both cost and throughput.
Microplate manufacturers are incorporating new microplate design features to
help reduce or eliminate the problem. Experimenting with these new designs
may improve your results and throughput, providing they meet the key sealing
criteria previously discussed—for example, having raised sample- well chimneys.
Application Support
Q
We have additional questions and need help with our applications. How can we
request help?
A
To help expedite your support request, you can complete and submit the
Request for PlateLoc Sealer Application Support.


Quick Reference Guide
G5402-90003