Effective Cryopreservation of a Bioluminescent Auxotrophic Escherichia coli-Based Amino Acid Array to Enable Long-Term Ready-to-Use Applications
- Authors
- Ahn, Hee Tae; Jang, In Seung; Dang, Thinh Viet; Kim, Yi Hyang; Lee, Dong Hoon; Choi, Hyeun Seok; Yu, Byung Jo; Kim, Moon Il
- Issue Date
- Aug-2021
- Publisher
- MDPI
- Keywords
- Amino acid array; Cryoprotectant; Escherichia coli auxotroph; Long-term preservation; Trehalose
- Citation
- BIOSENSORS-BASEL, v.11, no.8
- Journal Title
- BIOSENSORS-BASEL
- Volume
- 11
- Number
- 8
- URI
- https://scholarworks.bwise.kr/gachon/handle/2020.sw.gachon/82034
- DOI
- 10.3390/bios11080252
- ISSN
- 2079-6374
- Abstract
- Amino acid arrays comprising bioluminescent amino acid auxotrophic Escherichia coli are effective systems to quantitatively determine multiple amino acids. However, there is a need to develop a method for convenient long-term preservation of the array to enable its practical applications. Here, we reported a potential strategy to efficiently maintain cell viability within the portable array. The method involves immobilization of cells within agarose gel supplemented with an appropriate cryoprotectant in individual wells of a 96-well plate, followed by storage under freezing conditions. Six cryoprotectants, namely dimethyl sulfoxide, glycerol, ethylene glycol, polyethylene glycol, sucrose, and trehalose, were tested in the methionine (Met) auxotroph-based array. Carbohydrate-type cryoprotectants (glycerol, sucrose, and trehalose) efficiently preserved the linearity of determination of Met concentration. In particular, the array with 5% trehalose exhibited the best performance. The Met array with 5% trehalose could determine Met concentration with high linearity (R2 value = approximately 0.99) even after storage at −20◦C for up to 3 months. The clinical utilities of the Met and Leu array, preserved at −20◦C for 3 months, were also verified by successfully quantifying Met and Leu in spiked blood serum samples for the diagnosis of the corresponding metabolic diseases. This long-term preservation protocol enables the development of a ready-to-use bioluminescent E. coli-based amino acid array to quantify multiple amino acids and can replace the currently used laborious analytical methods. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.
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