Why Glycerol-free Reagents Matter in Molecular Diagnostics

Precision and reproducibility are paramount in molecular testing. Even minor components in reagent formulations can significantly impact diagnostic assay development and testing outcomes. Glycerol, a common cryoprotectant in many enzyme formulations, is valued for its protective properties when storing liquid reagents in a freezer, as it prevents ice crystal formation. However, the limitations of glycerol—such as its potential to complicate lyophilization (freeze-drying) workflows and challenge automated processes—have become increasingly apparent. To navigate these challenges, our eBook "Navigating Amplification Workflows for Robust Diagnostic Assay Development" offers in-depth insights and practical solutions.

Transitioning to glycerol-free reagents offers a way to overcome these obstacles, paving the way for more accessible and accurate diagnostic workflows. Read on to learn how glycerol-free alternatives are transforming the landscape of molecular biology.

 

Benefits of glycerol-free reagents for molecular diagnostics

Demand for glycerol-free reagents is increasing based on their benefits for advancing molecular diagnostics. By shifting to glycerol-free reagents, the accuracy, efficiency, and portability of diagnostic assays can be enhanced. However, it may be important to adopt alternative strategies to maintain the protective effects of glycerol, such as using other stabilizers, to ensure enzyme stability and functionality are preserved without the drawbacks of glycerol.

(1) Lyophilization compatibility, long-term stability, and global reach

Glycerol is incompatible with the lyophilization process, which is crucial for creating stable, dry reagents that do not require refrigeration. The drying process in lyophilization requires the sublimation of water (converting solid to vapor without passing through the liquid phase). The presence of glycerol can hinder this process because it retains moisture, making complete drying challenging. Consequently, formulations intended for lyophilization should be glycerol-free to ensure successful drying and long-term stability.

Lyophilized formulations can be stored at room temperature, eliminating the need for cold-chain shipping and cold storage. This is particularly beneficial for point-of-care or field diagnostics, especially in regions with limited access to cold-chain logistics.

By simplifying distribution and ensuring reliable performance across diverse climates and conditions, glycerol-free, lyophilized reagents in molecular diagnostic assays support global health equity in regions that typically lack sufficient healthcare resources.

 

(2) Automation accuracy

Glycerol's high viscosity can pose challenges in automated systems commonly used in diagnostic laboratories. These systems rely on precise and reproducible pipetting, and the viscous nature of glycerol-containing solutions leads to slower flow rates during pipetting. While several best practices are typically used when pipetting viscous liquids (e.g., reverse pipetting, liquid class development), glycerol can still cause inaccuracies in volume dispensing, clogging of pipettes, and difficulty achieving reproducible results. The slower aspiration and dispensing rates necessitate adjustments to protocol parameters to minimize liquid loss, ensure accurate pipetting, and provide adequate mixing, all of which complicate the automation workflow.

However, glycerol-free reagents effectively overcome these pitfalls by reducing solution viscosity, allowing for faster, more precise pipetting. This improvement enhances the accuracy and reliability of automated high-throughput systems while shortening the overall turnaround time. This makes glycerol-free formulations helpful for maintaining the reproducibility required in molecular diagnostics.

 

Making an assay reagent glycerol-free

Making a product glycerol-free involves several key steps to ensure the reagent remains stable, functional, and compatible with various applications, particularly in molecular diagnostics. The process begins with evaluating and selecting alternative stabilizers that can provide the necessary protective effects without the drawbacks of glycerol. Following this, the product is carefully reformulated, and concentrations and conditions are fine-tuned to ensure the glycerol-free version performs as effectively as the original. Rigorous testing, including shelf-life assessments and freeze-thaw cycle stability, is conducted to confirm the product's robustness. Finally, any necessary adjustments to the manufacturing process are made to accommodate the new formulation, ensuring consistent quality, even at larger production scales.

 

Offering comprehensive support for glycerol-free products

At New England Biolabs (NEB), we offer extensive support for scientists transitioning to glycerol-free reagents, including those developing molecular diagnostic assays. We provide a range of glycerol-free enzyme formulations specifically designed to meet the demands of advanced diagnostic assays, including PCR and isothermal amplification.

Our glycerol-free products are available in various concentrations and formats, tailored to fit your workflow. For those requiring further customization, our Customized Solutions Team is ready to assist in developing glycerol-free reagents to meet your unique requirements. These glycerol-free options are especially beneficial for those looking to improve the compatibility of their reagents with lyophilization processes, enhance automation in high-throughput systems, and ensure reliable performance across diverse conditions.

We also offer technical support and consultation to help labs validate and optimize their protocols when switching to glycerol-free formulations, ensuring a smooth and effective transition.

 

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