Identifying Common Bottlenecks in PCR Workflows

Polymerase chain reaction (PCR) is a technology that has advanced tremendously since its inception—Real-Time PCR, multiplexing, and automation have made the platform faster and more accessible, allowing PCR to become a popular tool in modern laboratories However, these innovations have also set a high bar for efficiency, and now, continuous optimization of PCR workflows isn’t just an option—it’s an expectation.

For all its advancements, labs continue to grapple with a number of inefficiencies common in PCR workflows that compromise their ability to achieve reliable, reproducible results.
Let’s take a closer look at some of these persistent challenges so we can navigate potential solutions.

Personnel Are Occupied with Time-Consuming Assay Prep

Inefficiencies begin even before your PCR workflow does—labs must first prepare their assays before being able to use them. This is a manual and tedious process for lab personnel with many steps, typically including:

• Gathering materials and reagents
• Calculating volumes
• Combining reagents
• Aliquoting master mix
• Adding primers and probes
• Adding template (when indicated)
• Mixing and preparing for PCR

While necessary, assay prep ties up staff with rote work which may not be reflective of their level of training, skill, and experience. This can lead to frustration on the part of the staff member, and might also mean a poor allocation of expertise within your lab.

Manual Assay Prep Can Be Error-Prone

Assay preparation in PCR workflows is typically manual, which introduces inherent challenges. Volumes may be inappropriately calculated, expired products may accidentally get used, and accuracy depends heavily on the technician’s skill and precision. These inconsistencies can lead to:

• Variability in results: Small pipetting errors can have outsized impacts on downstream data.
• Inconsistent pipetting volumes: Human error and individual performance can make it
  difficult to maintain precise reagent amounts.
• Cross-contamination risks: Manual assay preparation means reagents are frequently
  handled, which raises the likelihood of contamination and thus potentially jeopardizes
  sensitive experiments.

Whether your lab is clinical or pre-clinical, PCR is a technology with a low tolerance for error, which means minimizing opportunities for mistakes should be high on your list of priorities.

Assay Preparation Carries Hidden Costs

In addition to introducing opportunities for error, assay preparation carries costs outside of the simple use of materials and personnel time. Labs often struggle to predict the exact amount of reagents needed due to variables like pipetting loss, unexpected sample volumes, or changes in experimental design. Because of this, labs will typically factor in overage and prepare surplus reagents to ensure consistency, but these are practices which have their own set of challenges:

• Material waste: Primers, probes, and enzymes are often costly, and wasting them
  can add up over time.
• Inventory complexity: Managing a wide array of reagents with varying storage
  conditions and expiration dates requires meticulous oversight.
  In many cases, tracking of these variables may not be perfect and more waste will
  ensue.

• More strain on personnel time: It should also be considered that inventory oversight
  requires its own allocation of staff time and cognitive load.

For many labs, these hidden costs are not insignificant and can often impact both the lab’s budget and its overall operational efficiency.

Assay Diversity Can Lead to Challenges in Reproducibility

One of the strengths of PCR is its adaptability to different experimental needs, but this flexibility also introduces complexity. Each assay can require its own unique annealing temperatures, enzymes, or cycling conditions. This can lead to:

• Misapplied protocols: If careful attention is not given, personnel may use conditions
  optimized for one assay on another, leading to unreliable or incorrect results.
• Time lost to troubleshooting:  Assay condition errors can be difficult to identify, and
  requires retracing your steps in order to identify where the issue originated.
  Afterward, rerunning samples and recalibrating methods can further delay projects and
  strain resources.

Inconsistent assay conditions contribute to the types of reproducibility issues that can undermine the credibility of experimental findings.

A Path Forward: Introducing Breakaway, Pre-Plated Panels

Despite these persistent challenges, labs are not without recourse to be able to address them. Some are choosing to adopt the use of breakaway, pre-plated panels as a way to improve their PCR workflows. By outsourcing assay preparation, labs can eliminate many of the inefficiencies described above and discover:

• Reduced waste: Panels that can break away allow you to use just what you need and save remaining assays for future use. You’ll also experience less waste from complex reagent inventory management and factored-in overages.
• Reduced variability: Pre-plated panels minimize human error and ensure consistent reagent volumes so your PCR workflows can reliably depend on precise master mixes.
• Lower contamination risk: Less handling means a cleaner workflow and fewer
  opportunities for contamination.
• Time savings: Ready-to-use panels free up technicians to focus on high-value tasks like
  data analysis and assay development, rather than assay prep or inventory management.
• Improved reproducibility: Pre-optimized conditions for multiple assays reduce the risk of
  variability and errors.

Pre-plated, breakaway panels might just be the tool your lab needs to make steps in reducing inefficiencies and improving outcomes without compromising quality. The future of PCR lies in solutions that empower labs to deliver reliable, reproducible results—every time.

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