Pullulan Encapsulation of Labile Biomolecules to Give Stable Bioassay Tablets

Abstract

A simple and inexpensive method is reported for the long‐term stabilization of enzymes and other unstable reagents in premeasured quantities in water‐soluble tablets (cast, not compressed) made with pullulan, a nonionic polysaccharide that forms an oxygen impermeable solid upon drying. The pullulan tablets dissolve in aqueous solutions in seconds, thereby facilitating the easy execution of bioassays at remote sites with no need for special reagent handling and liquid pipetting. This approach is modular in nature, thus allowing the creation of individual tablets for enzymes and their substrates. Proof‐of‐principle demonstrations include a Taq polymerase tablet for DNA amplification through PCR and a pesticide assay kit consisting of separate tablets for acetylcholinesterase and its chromogenic substrate, indoxyl acetate, both of which are highly unstable. The encapsulated reagents remain stable at room temperature for months, thus enabling the room‐temperature shipping and storage of bioassay components.

Paper microfluidics for nucleic acid amplification testing (NAAT) of infectious diseases

“…enzymes are commercially available in large batches, but, once rehydrated, their stability becomes a key concern for the application of NAATs in resource-limited areas, since there is limited availability of cold storage facilities (32). We stored rehydrated LAMP mixtures under three different conditions (−20°C, 4°C, and at room temperature, i.e., ∼22 ± 2°C) for 72 h, and measured their activity every 12 h, using a target concentration of 10 5 IU/mL, by real-time PCR and by using the DNA detection strip used in the microfluidic device ( Fig.…”

Development of a point-of-care diagnostic for influenza detection with antiviral treatment effectiveness indication

“…Pullulan, from Aureobasidium pullulans , is another sugar-based preservative storage method used previously by another group for enzyme preservation 36 . In their study, they showed very good retention of TaqDP enzyme activity, greater than 90%, even after 50 days of dried storage in Pullulan capsules 36 .…”

“…Pullulan, from Aureobasidium pullulans , is another sugar-based preservative storage method used previously by another group for enzyme preservation 36 . In their study, they showed very good retention of TaqDP enzyme activity, greater than 90%, even after 50 days of dried storage in Pullulan capsules 36 . We tested the pullulan with the enzyme substrate and purified neuraminidase enzyme in a pre-dried storage assay to verify that there would not be any negative impact to assay performance (Figure 4).…”

Innovative technologies for point‐of‐care testing of viral hepatitis in low‐resource and decentralized settings

“…Freeze‐drying was used to store the Zika toehold switch biosensor on paper, as previously cited . Encapsulation of reagents in natural polysaccharide was also developed to store chemicals on chips . Storage of reagents in a liquid form, which was a challenge until now, was performed using micro pouches or bioinert glass ampoules.…”