Abstract
Molecular biology is a widely used and widespread technique in research and as a laboratory
diagnostic tool, aiming to investigate targets of interest from the obtainment, identification,
and analysis of genetic material. In this context, methods, such as Polymerase Chain Reaction
(PCR), Reverse Transcription Polymerase Chain Reaction (RT-PCR), real-time PCR, loopmediated
isothermal amplification (LAMP), and loop-mediated isothermal amplification with reverse
transcription (RT-LAMP), can be cited. Such methods use enzymes, buffers, and thermosensitive
reagents, which require specific storage conditions. In an attempt to solve this problem,
the lyophilization procedure (dehydration process by sublimation) can be applied, aiming to preserve
and prolong the useful life of the reaction components in cases of temperature variation. In
this review, we present a synthesis of the lyophilization process, describing the events of each step
of the procedure and providing general information about the technique. Moreover, we selected
lyophilization protocols found in the literature, paying attention to the conditions chosen by the
authors for each step of the procedure, and structured the main data in tables, facilitating access to
information for researchers who need material to produce new functional protocols.
Keywords:
Lyophilization, polymerase chain reaction, loop-mediated isothermal amplification, protectants, freeze-dried, pointof- care, storage.
Graphical Abstract
[14]
Patapoff, T.W.; Overcashier, D.E. Importance of freezing on lyophilization development. BioPharm, 2002, 15(3), 16-21.
[26]
Nail, S.L.; Akers, M.J. Development and Manufacture of Protein Pharmaceuticals; Springer: New York, 2012.
[30]
Aradhya, S.; Ramlal, S.; Madalgi, R.; Murali, H.S.; Mondal, B.; Monda, A. l, G.S.; Kulkarni, R.D.; Batra, H.V. Simultaneous identification of diarrheagenic Escherichia coli using thermostabilized multiplex PCR formulation shruthi. Int. J. Sci. Res. Biol. Sci. Vol, 2020, 7(5)
[32]
Khazani, N. A.; Noor, N. Z. N. M.; Yean, C. Y.; Hasan, H.; Suraiya, S.; Mohamad, S. A A thermostabilized, one-step PCR assay for simultaneous detection of Klebsiella pneumoniae and Haemophilus influenzae. J. Trop. Med., 2017, 2017
[49]
Jadhav, T.R.; Moon, R.S. Review on lyophilization technique. World J. Pharm. Pharm. Sci., 2015, 4(5), 1906-1928.
[62]
Brivio, M.; Li, Y.; Ahlford, A.; Kjeldsen, B.G.; Reimers, J.L.; Bu, M.; Syvänen, A-C.; Bang, D.D.; Wolff, A. A simple and efficient method for on-chip storage of reagents: Towards lab-on-a-chip system for point-of-care DNA diagnostics. Micro Total Anal. Syst., 2007, 59-61.
[65]
de Campos-Stairiker, K.; Pidathala, A.S.; Gill, R.K.; Gelimson, I.; Varankovich, N.; Gill, S.S.; Slyadnev, M.; Kapur, S. Validation of microchip RT-PCR COVID-19 detection system. J. Biosci. Med., 2021, 9(9), 8-24.
[69]
Takekawa, J.Y.; Hill, N.J.; Schultz, A.K.; Iverson, S.A.; Cardona, C.J.; Boyce, W.M.; Dudley, J.P. Rapid diagnosis of avian influenza virus in wild birds: Use of a portable RRT-PCR and freeze-dried reagents in the field. J. Vis. Exp. JoVE, 2011, 54, 2829.