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Nowadays, almost every biology lab has a PCR instrument — from portable, battery-operated machines, to “PCR-by-water baths”, do-it-yourself PCR, or familiar vendor packages, including those with real-time quantification or droplet digital capabilities, DNA amplification depends on a robust thermal cycler.
If you are looking to upgrade, add to, or even replace the lab’s current PCR equipment, there are more factors to consider than just the purchasing cost. Here is a rundown of our top features to look out for when you are shopping for a new PCR instrument:
1. Sample Capacity
Versatility to accommodate individual 0.2 mL tubes, strips of tubes and multiwell plates is a good place to start with any PCR instrument. While you may not need to run PCR on 96- or 384-well plates today, the lab might depend on that capacity in the future.
PCR instrument hinges on the use of “skirted” or “unskirted” plates. If your lab routinely uses one style of multiwell plate (and not the other), and they are PCR-compatible, it might be cost-effective to select a thermal cycler that can accommodate them.
2. Reaction Volumes, Interchangeable Sample-Blocks
Some permanent sample block thermal cyclers support large reaction sizes, such as 0.5 mL volume tubes, but compared to standard 96-well blocks, running large reaction volumes will reduce the total number of samples you can run at once. To keep your options open, look for a PCR instrument that has multiple or interchangeable blocks. This is a good way to save on bench space – you can simply order another block when a new need arises, rather than buying another machine!
This can also be an effective way to reduce sample cross-contamination between different types of experimental material, by using one block at a time for each type of experiment. Having the ability to restrict cross-contamination might be useful in a lab that routinely analyzes both mammalian and bacterial gene expression.
3. Temperature Range and Ramp Rates
The majority of PCR reactions run well under standard protocols. It is often the non-standard or complex protocols (e.g., those with highly customized or modified primers or reagents) that require extra optimization. Having the possibility to run PCR over a wide temperature range (e.g., 0-100°C) ensures that your scope for optimization is not constrained by hardware limits.
Controllable ramp rates, or how fast the cycler will heat or cool to a given temperature, is a minor feature but is worth mentioning because slower rates ( in °C per second) may carry a higher risk of non-specific primer binding. So, opt for cyclers with fastest ramp rates for more PCR success each day!
4. Temperature Accuracy and Uniformity
Do keep in mind that there may be a trade off between the speed at which a cycle ramps up to temperature and the temperature accuracy during that run. High speed may also introduce higher variability, at least in qPCR.1 You want a PCR instrument that heats every sample in the block the same way, hits the target temperature on the dot (plus or minus a few tenths of a degree), and maintains it consistently. Here is where technical specifications are everyone’s friend: select equipment that has the lowest +/- range for operating margins when it comes to both temperature accuracy and uniformity.
5. Heated Lids
During PCR, sample tubes or plates are heated by direct contact with the sample block. At the top of each tube there is usually a small part that is not in direct contact with the sample block at all. Without a heated lid you run the risk of creating a hot-to-cold temperature gradient where the liquid in the bottom of the tube can evaporate and subsequently condense at the top of the tube. With such small reaction volumes this can have a significant impact on the concentration of the PCR reaction components, skewing qPCR results, and potentially inhibiting an otherwise straightforward PCR run.
In a nutshell, heated lids are a must-have feature to help maintain temperature uniformity in every PCR reaction.
6. Auto-Restart Function
Another must-have feature incorporates smart technology that allows a run to pick up where it left off after an interruption caused by a power outage, and continue all the way to the end. This is an automatic feature on the thermal cyclers that possess it.
7. Zonal Heating
Having the ability to set up a number of distinct temperature zones on a sample block helps you find the perfect annealing temperatures for your primers more quickly, sometimes in as little as one run. This technique is commonly referred to as gradient PCR and is a go-to option when troubleshooting a reaction with no amplification.
8. Adaptability and Scalability
Any new PCR instrument needs to be flexible enough to grow with the lab because after all, this is a significant purchase that will serve the lab for many years to come. It will increase the capacity at which you can run reactions. It will add support for new and advancing biotechnologies. And whether the lab runs each cycler independently, or controls banks of them by keyboard and mouse, it encourages researchers to collaborate by storing custom settings, programs and amplification data, all in one place.
What other favorite features do you look for in a PCR instrument? Feel free to share them with us by writing in the comments section.
- Hilscher C, Vahrson W, Dittmer DP. (2005). Faster quantitative real-time PCR protocols may lose sensitivity and show increased variability. Nucleic Acids Research, 33(21):e182.