Absolute precision

The relatively new field of molecular diagnostics offers much – but accurate and precise liquid handling is vital to this burgeoning area 

MOLECULAR diagnostics is a relatively new territory that offers much promise for early disease detection and personalised patient care. When analysing samples at the molecular level, there are serious consequences of error and tests must be highly accurate and precise. For example, molecular diagnostics may be used to detect the presence of DNA in samples collected at a crime scene, or to determine whether parents carry a genetic mutation that causes inherited diseases, like cystic fibrosis.

While quality molecular test results are essential, standardised quality controls for molecular diagnostic tests were rare until recently, when Maine Molecular Quality Control (MMQCI) was founded to fill this market need. The company manufactures and sells the first FDA-approved quality control for genetic testing and is on a mission to help companies using or manufacturing molecular diagnostics enhance data integrity.  

Quality is the company’s mantra and they practice what they preach. As pipetting is one of the most ubiquitous and important processes in the development and manufacture of its quality control materials, MMQCI has a stringent liquid handling quality assurance program.

To ensure that its pipetting is accurate to the sub-ml level, MMQCI uses the ARTEL PCS pipette calibration system. The result has been greater control over operations, enhanced efficiency, and confidence in laboratory operations.

Advances are continually made in the molecular diagnostic field, leading to more widespread use in numerous applications. Molecular diagnostics currently offer a wealth of benefits such as rapid analysis, sensitive responses, and the provision of detailed information for diagnosis and personalised treatment.

As the popularity of molecular diagnostics continued to rise, Dr Clark Rundell and Joan Gordon, the founders of MMQCI, recognised that the lack of standardised quality control materials could be detrimental to the growth of this promising industry. Both have extensive experience in the heavily regulatory clinical laboratory market, where quality controls are part of the daily routine. Yet at the onset of molecular testing, quality control samples were often derived from patient samples or other biological materials, which can vary in consistency.

To address the quality issues in the molecular diagnostic industry, MMQCI invented a process to stabilise target DNA constructs to be used as controls for molecular-based diagnostic assays. Protocols were then developed to manufacture an FDA-cleared quality control for molecular genetics test platforms. Now, the company markets and sells standardised INTROL quality control products that laboratories can use to monitor molecular test processes, including extraction, amplification, and detection steps.

From R&D to product manufacture, testing and package filling, liquid handling quality control processes are in place throughout MMQCI’s facilities. Pipetting accuracy and precision are most essential in the manufacture of the company’s INTROL controls. Pipettes are used for the most critical manufacturing task – the addition of small volumes of DNA. Quantities must be exact for reproducibility. 

“Pipettes are the single most used tool in our laboratories,” said Joan Gordon, President, MMQCI. “We manufacture our products using highly controlled processes so that each lot contains the identical concentration. There is no room for error.”

Reproducible quality controls provide the ability to identify variation in a laboratory test system. For example, trending over time in the signal strength can indicate a failing probe or system malfunction that could lead to failure to detect a genetic mutation. 

Consider the case of MMQCI’s INTROL cystic fibrosis panel I control. A laboratory monitoring the signal strength of its assay using these controls would expect a new lot of controls to produce signal strength comparable to the previous lot. It is critical that MMQCI’s product lots be consistent batch-to-batch so that any variation is due to the system, and not due to variation in the control.

In addition, the small volumes typically handled in MMQCI’s facilities make liquid handling quality assurance even more important. The smaller the volume being pipetted, the larger the risk for error, because small volume pipettes are more prone to error. 

“We’re often pipetting five microliters or two microliters,” said Paula Pinette, Customer Service, MMQCI. “We try to scale up the volumes handled in the most critical steps to use as large a volume as possible, but this could still be only 15ml.”

When amplifying DNA using PCR, which is a common task at MMQCI, pipettes are used to add essential components to the master mix, including enzyme, buffer, and water. Here, where small batches are frequently run, inaccuracy of even one ml can make a difference.

At MMQCI, pipetting is also important for product testing, whether for an intermediate product or for a finished product prior to market release. Typical quality control testing includes extraction, PCR, and detection, which all require pipetting. Pipetting error could lead to the release of an under-performing control or the withholding of a performing product.

Pipettes are also used to fill product packages prior to shipment to customers.  While inaccurate volumes will not affect the performance of the control, it can lead to customer satisfaction issues. “We do not want to shortchange a customer who ordered two milliliters of product due to faulty pipetting,” said Pinette.

While pipette calibration is a given at MMQCI, like most regulated laboratories,

“At the onset of molecular testing, quality control samples were often derived from patient samples or other biological materials, which can vary in consistency.” “We manufacture our products using highly controlled processes so that each lot contains the identical concentration. There is no room for error.”

there are several available options. One option is to outsource pipette calibration to an external service provider, which was the method employed by MMQCI when the company was in its infancy. However, as the company grew and operations expanded, the company sought stronger control over its quality control processes.

The company chose to implement the ARTEL PCS in its own laboratory quality program. The PCS can be used in the laboratory to rapidly verify the accuracy and precision of pipetted volumes. The PCS also provides NIST-traceable measurements for standardiSed assessment of dispensed volumes. Now MMQCI conducts its annual pipette calibrations on-site and is also able to calibrate its critical pipettes on a more frequent quarterly schedule. These pipettes are used for the most volume-dependent, mission critical operations, such as the addition of DNA. More frequent checks provide greater assurance of pipette performance and product quality. 

The PCS also has pipette tracker software, which automatically alerts laboratory managers when pipettes are due for calibration and stores performance data for each pipette. This information is essential for FDA audits and quality assurance as well as to meet ISO and CE Mark standards, which is important as MMQCI pursues business opportunities in Europe.

“We are an FDA-registered facility and follow cGMP guidelines. We need records for all of our critical processes, and pipetting is clearly one of them. The PCS helps us demonstrate our stringent liquid handling quality control processes,” said Gordon.

While previously working in other laboratories, Gordon and Pinette used a variety of techniques to calibrate pipettes, including gravimetric weighing. However, this technology is time-consuming and prone to variability caused by environmental conditions, such as static electricity and evaporation, and must be conducted in humidity- and temperature-controlled facilities.

In addition to helping MMQCI prevent liquid handling errors caused by pipette failure, the PCS also is used to improve operator pipetting technique. When handling the minute volumes typical of molecular diagnostics, accuracy and precision are critical and operator technique can contribute to errors.

“The PCS allows us to immediately see how small tweaks to pipetting technique can have a drastic effect on pipetted volume,” said Pinette. MMQCI now requires all scientists to pass annual pipetting proficiency tests and review proper technique as part of the company quality plan.

MMQCI has also leveraged its relationship with ARTEL to develop SOPs on proper pipetting technique, relying on ARTEL for guidance on topics from insertion depth to pipetting angles. 

“The PCS is fast, easy-to-use, and provides traceable results,” said Gordon. “ARTEL also has the right credentials, which takes the onus off of us in terms of proving the credibility of our pipette calibration method for cGMP approval.”

After switching to the PCS, MMQCI now has control over its liquid handling operations and can rest assured that its pipetting is accurate and precise, ensuring that its products always meet the demanding quality requirements of its customers.