PCR Plate Sealing: The Complete Practical Guide

Appropriately sealing a PCR plate will help protect your experiment by preventing evaporation of samples while in the thermocycler. As water escapes, it alters the efficiency of the reaction. Depending on the severity of evaporation across the PCR plate, the results could range from no amplification in affected wells to greater variability and reduced reproducibility overall.

This video demonstrates both the adhesive seal and capping methods:

Sealing plates for PCR is an essential step to prevent both contamination and evaporation during the amplification process. Here are the steps to take to help ensure a good PCR plate seal. 

1. Set up your PCR reactions in the wells of the plate in a clean room or workspace, taking care to avoid getting liquid outside the wells. It is important to maintain a clean environment throughout the setup and sealing process to avoid contamination with amplifiable material.
2. For sealing plates with adhesive film seals,
   1. Carefully peel the backing from the adhesive seal.
   2. Align the seal with the PCR plate then press the seal onto the plate.
   3. Using an applicator or other tool, firmly rub the seal along rows and columns so each well is sealed.
3. If sealing plates using PCR cap strips, 
   1. Make sure your PCR plate is secure in a rack.
   2. Align the cap strip(s) to the designated rows or wells of the plate
   3. Seal each cap strip to its set of wells using a PCR capping tool as shown in the above video.
4. Alternatively, press down well on each cap to insert it into its well. You should hear a snap when it is in place.
5. After sealing the PCR plate, visually inspect each well to ensure that there are no gaps or leaks. Caps should be at equal height across the plate. Any compromised seals should be addressed immediately by resealing or replacing the contents of the affected well.
6. Once properly sealed, proceed with amplification and analysis OR store the PCR plate in a suitable container to protect it from contamination and physical damage. Keep the plate in a controlled environment, such as a refrigerator or freezer if required for long-term storage.

Consider the following decision points and tips for picking the right sealing method for your PCR project.

A key decision is whether to use PCR caps or films for sealing plates. The primary benefit of caps is that they can be readily removed and replaced from the PCR plate as needed, whereas adhesive films tend to leave a residue upon removal and heat seals are not removeable. PCR caps may also be a better choice when using only a few rows or columns of the PCR plate.

When sealing plates with caps, PCR strip caps are most often used. It’s important to select caps that are compatible with the plate manufacturer to ensure optimal sealing performance and to choose either flat or domed caps depending on your thermal cycler. Since caps can be used on PCR plates as well as PCR tubes, labs can purchase just one kind of cap to match both.

Applying caps can be tedious for larger experiments where the entire plate is used or when multiple plates are required, though a PCR cap installing tool can make this easier. When re-capping wells in a PCR plate, take care to position the cap on the same well to prevent sample cross-contamination.

Film plate seals are more versatile since they can be used on any PCR plate, regardless of the manufacturer. This allows you to use film seals on any traditional 96-well PCR plate as well as other plate formats. Since the films can be cut to match the plate size or number of wells being used, you can tailor the film seal to your need. On the flip side, small strips of adhesive film require great care to seal properly. Forming a tight seal with films can be less obvious than caps, since the caps click into place when tightly sealed.

Your choice of plate seal type depends on your application. For qPCR experiments, which involve making high sensitivity fluorescence measurements directly in the plate, select optical seals to permit maximum fluorescence transmission from the wells. Not all clear seals have enough optical clarity for qPCR, so make sure that you are using optical seals to ensure effective data collection.

While not compatible with qPCR, aluminum foil films provide additional protection for light-sensitive samples. Furthermore, they are often better suited for colder temperature storage (≤–80°C). As a result, foil films are an excellent choice when long term storage is important. Foil seals can also be pierced, which means samples can be transferred from the plate using needles—either manually or with a robotic sampling platform.

If film seals are the best fit for your PCR workflow, it’s also important to determine whether you want adhesive or heat sealing. Generally, adhesive films are the most straight forward option for labs, since they only require a user to apply the sticky side to a plate surface and carefully establish tight seals, often using adhesive film applicator tools.

Heat sealing films generally require the use of a plate sealer, which binds a non-adhesive plastic film to the plate to form a semi-permanent seal. While heat sealing adds additional overhead cost due to purchasing a plate sealer, they are superior for long term storage since they provide durable seals that are more effective at preventing evaporation compared to adhesive seals.

 User bulletin: How to Use MicroAmp Reaction Plates, Tube Strips, and Tubes
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PCR Plate Seals and Accessories—Thermo Scientific