In PCR and qPCR workflows, much of the focus is placed on reagents, primers, plates, and instrumentation. Yet the sealing film placed over the plate can also play an important role in protecting reaction conditions throughout thermal cycling.
With repeated heating and cooling, an effective seal helps minimize evaporation, maintain consistent reaction volumes, and reduce the risk of cross-contamination between wells. Choosing the right film becomes particularly important with low-volume reactions, where even small amounts of evaporation can change reagent concentrations and potentially affect assay consistency.
PCR or qPCR? The application matters
While conventional PCR and qPCR share many sealing requirements, there is one important difference: optical detection.
For conventional PCR, the primary goal is typically to maintain a secure seal throughout thermal cycling. Temperature resistance, evaporation control, plate compatibility, and reliable adhesion are therefore key considerations.
For qPCR, these characteristics remain important, but optical performance must also be considered. Many real-time PCR instruments measure fluorescence through the top of the plate, making the sealing film part of the optical pathway.
What should you consider when selecting a film?
Evaporation control: A strong vapor barrier helps maintain reaction volume during repeated thermal cycling. This becomes increasingly important as reaction volumes decrease.
Temperature resistance: The film and adhesive must tolerate repeated changes in temperature without lifting, shrinking, or losing adhesion.
Optical clarity: For qPCR, consider whether the film is designed specifically for fluorescence-based detection rather than simply being visually clear.
Plate compatibility: Plate material, well geometry, and sealing surface can all influence adhesion. A film that performs well on one plate design may perform differently on another.
Adhesive properties: More adhesive is not necessarily better. The goal is a secure seal that withstands thermal cycling while meeting the needs of the downstream workflow.
Post-PCR access: Consider what happens after amplification. If samples need to be recovered, ease of removal or the ability to pierce the seal may influence the best choice.
Don’t overlook sealing technique
Film selection is only part of achieving a reliable seal. Pressure-sensitive films require firm, even application across the plate, including the perimeter and areas between wells. Inconsistent pressure can result in incomplete adhesion and contribute to well-to-well variability.
For reproducible PCR workflows, standardizing how the film is applied can be just as important as standardizing the film itself.
Think beyond simply covering the plate
There is no single sealing film that is ideal for every PCR application. Conventional PCR, qPCR, low-volume assays, automation, and post-amplification workflows can each place different demands on the seal.
The best approach is to consider the plate, sealing film, instrument, and assay as a complete system.
Choosing a film based on the requirements of the entire workflow can help minimize evaporation, protect samples, reduce preventable variability, and support more consistent PCR and qPCR results.
Selecting an Excel Scientific sealing solution
Excel Scientific offers several sealing options designed for different PCR and qPCR workflow requirements:
For qPCR – ThermalSeal® RT2™
An optically clear sealing film designed specifically for qPCR. Its highly conformable adhesive creates a strong seal to help minimize evaporation and cross-contamination while providing the optical clarity needed for fluorescence-based detection. ThermalSeal RT2 is particularly suited to flat-top and hard-shell PCR plates.
For qPCR requiring strong evaporation protection – ThermalSeal® RTS™
An optically clear film with low autofluorescence and a silicone adhesive designed to provide a strong seal. ThermalSeal RTS is an option when both optical performance and evaporation control are important.
For conventional PCR – eXTReme™ Seal
Designed for PCR applications where minimizing evaporation is a priority. Its thick, strong adhesive provides a secure seal during thermal cycling while allowing clean removal after use. Different film sizes are available to accommodate raised-rim, non-skirted, and flat-top PCR plates.
For PCR requiring sample access – FoilSeal™ PCR
A pierceable aluminum foil option that provides a strong barrier against evaporation and cross-contamination. The foil can be pierced with a pipette tip or robotic probe, making it useful when samples need to be accessed after sealing.
The appropriate choice ultimately depends on the assay, plate geometry, instrument, reaction volume, and downstream workflow. Matching these requirements to the characteristics of the sealing material helps ensure that the seal supports the experiment rather than becoming another source of variability.