by Ninad Pendse, Lead Product Manager for Molecular Diagnostic Products, Asuragen
When it comes to making smart and careful buying decisions for the lab, we tend to focus on the big-ticket items: next-generation sequencers, high-throughput diagnostic testing platforms, even thermal cyclers can require hours of research and conversations with peers. But often it’s the smaller items that really make a difference in how well day-to-day workflows perform. When there are problems with reagents, consumables, or assay kits, they can be enough to throw the lab into temporary chaos.
That’s why it is worth spending as much time choosing the humble assay kit as it is a new instrument platform. Ideally, these workhorse kits should allow lab teams to run important workflows reliably, rigorously, and cost-effectively. They should be made by a manufacturer that can be considered a trusted partner — a vendor as committed to quality control and robust results as the lab staff that will be generating the data.
This buyer’s guide includes a variety of useful factors to consider when selecting an assay kit for use in translational research or a clinical laboratory. Some of these will be obvious, such as cost or performance, while others may be more unexpected. Certain factors worth considering are specific to key applications, so those will be reviewed as well.
Factors to consider in choosing an assay kit
All-in-one kit. Some manufacturers include all of the materials needed for a workflow, while others require users to source important components. In general, the assay kits that are simplest to integrate into the lab are ones that do not force users to source products from a large number of different vendors. Look for assays that include all — or at least nearly all — necessary components in one easy-to-order package.
Calibrators and controls. Certain elements that are essential for running an assay reliably should always be included in the kit. Calibrators and controls are intrinsic to the ability to run a high-quality workflow. If assay kits do not include them, or detailed instructions for which calibrators and controls to use and how to acquire them, that’s a real red flag.
Clear, detailed instructions. Speaking of instructions, the entire assay kit should come with well-written, step-by-step documentation for how to use it. Instructions should cover all key elements, such as how to run the assay, how to set up the master mixes, how to set up the thermal cycling, and so on.
Company reputation. It takes years to build a strong reputation, which is why this factor is important. Whenever possible, choose assays from companies with a long track record of demonstrated success. If you know a vendor has cracked a difficult nut with one type of assay, it’s a safer bet that the vendor can do the same thing in another area. For assay developers, longevity isn’t an accident; staying power means that they’re doing something right. On the flip side, even an exciting new assay from a startup company may not be worthwhile; users can’t be sure of how a young company will support the assay over time or whether it will be around for as long as the lab wants to use the kit.
Data analysis solutions. For very simple assays, this is not necessary. But with more complex assays, users should expect data analysis solutions to come with the kit to help make sense of the results. That might come in the form of software or, alternatively, access to a secure, cloud-based analysis pipeline. However they’re delivered, analysis tools should come from the manufacturer and generate clear, easy-to-interpret results in a format that can be ported to downstream analytical pipelines when needed.
Real cost. This is probably the first thing any buyer thinks about, but it’s easy to see the price tag and miss the bigger picture. In addition to the price a vendor will charge for the assay kit, it’s important to factor in the costs of all components needed to run the workflow, including staff time. Assays that require more hands-on time may have lower up-front prices, but all that extra manual effort can add substantial costs to the process. In addition, assays with higher failure or rerun rates can increase costs as well.
Regulatory requirements. Many labs don’t have to worry about this, but for clinical labs it’s a real concern. Choosing an FDA-cleared assay instead of developing their own test can ease some of the burden for clinical labs. Unfortunately, because regulatory clearance is so challenging to achieve, tests that go through the process are often locked in their approved state; newly identified genetic mutations, for instance, can’t easily be added after a test has gone through review.
Right platform. Generally speaking, it’s safe to assume that the developer of an assay kit has selected the underlying instrumentation for which it will work best — PCR, capillary electrophoresis, next-generation sequencing, etc. But whether the technology required to run the assay kit matches the instrumentation available in the lab is another story. If the top-performing assay kit has to be run on a specialty device the lab doesn’t have and would require considerable capital expense to acquire, the lab might be better served going with the next best kit if it runs on more widely available instruments.
Technical support. Never underestimate the value of vendor support. Even the best assay kits can have problems, and it’s imperative that users have a reliable contact for troubleshooting. Ideally, vendors will also be willing to provide training and other kinds of ongoing support to ensure smooth assay operations in the lab.
Technology compatibility. When a specialty or proprietary platform is required to run an assay, it’s worth checking to see if the same vendor also offers other assays designed for that same instrument. When a reliable vendor can provide multiple assays on a certain platform, it’s a better argument for making the investment in new technology.
Verification and validation. These processes are performed by the vendor and can often be opaque to outsiders — but they’re definitely worth checking. Assay developers can rush a new kit to market, especially when it’s not for clinical use, or they can take the time needed for rigorous internal testing. Extensive testing might include checking a number of different variables with the kit, such as how it performs with various extraction methods, thermal cyclers, and so on. This process ensures that stated sensitivity, specificity, and other performance metrics hold up across these variables. Ultimately, the best-tested assays should work reliably in any lab with any operator. In order to trust performance claims, look for rigorous verification and validation processes.
Application-specific considerations
Assays used for certain key applications in translational research or clinical laboratory use can come with additional layers of complexity that should also be considered in the buying process. Many of the challenges that arise relate to genes or genetic regions that are difficult to amplify, sequence, or analyze.
For example, assays used for neurological disorders or rare disorders — especially panel assays that test for a number of different conditions at once — often must be able to detect pathogenic repeats. Repeat expansions, such as the CGG repeat in the FMR1 gene that is responsible for fragile X syndrome, can be quite large and challenging to resolve due to the accuracy and sensitivity required to quantify copy numbers and detect small interruptions. Short-read sequencers tend to struggle with these regions, which frequently consist of extremely GC-rich content. A more effective approach for repeat disorders typically involves long-read sequencing, which represents these regions more accurately.
For carrier screening, the American College of Medical Genetics and Genomics now recommends analyzing 113 genes for nearly all pregnant patients and people planning to become pregnant. About a dozen of the genes included in that list are very difficult to sequence, particularly with conventional short-read sequencing platforms. Many clinical labs have had to set up a number of different workflows: one for all of the easy genes, and separate ones for the problematic genes, which are often addressed with several different technologies. This has become a serious burden for clinical laboratories. A better alternative would be a single assay covering all of the tough-to-sequence genes recommended for carrier screening, which would allow labs to streamline their operations with just two workflows needed to cover all 113 genes.
In cancer research or oncology, exceptional sensitivity is often necessary to find the relevant signal amid a noisy background. This might involve detecting rare transcripts or genetic mutations. For cancer-related assays, a robust process that leads to both high specificity and sensitivity is essential. This applies to assays designed to detect gene fusions to match patients to the right treatment, to monitor treatment response among individuals undergoing certain types of therapy, and much more.
Looking ahead
This list of factors to consider might seem overwhelming at first, but using it as a guide in the assay selection process can lead to much better laboratory results in the long run. When in doubt, look for a trusted partner to deliver high-quality products; reliable manufacturers will often include a lot of the information above on their websites to make it easier for researchers to evaluate their options.
About the author
Ninad Pendse serves as Lead Product Manager for molecular diagnostic products at Asuragen, a Bio-Techne brand.