Authors : Katiana Simões Lopes Kittelson; Allana Cristina Faustino Martins; Gizele Celante; Arquimedes Gasparotto Júnior; Roberto Da Silva Gomes

Volume/Issue : Volume 10 - 2025, Issue 3 - March

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Google Scholar : https://tinyurl.com/5e46mw5s

Scribd : https://tinyurl.com/skveapj3

DOI : https://doi.org/10.38124/ijisrt/25mar736

Note : A published paper may take 4-5 working days from the publication date to appear in PlumX Metrics, Semantic Scholar, and ResearchGate.

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Abstract : Considering the number of available methods in molecular and cellular biology and the rapid development of new technologies, the need for an updated guide on what is currently applied and how to choose the best method for a specific goal has increased. Gold standard methods are known for their accuracy and reliable data under certain circumstances, allowing replicability. This perspective aims to characterize and define the current gold standard techniques from the last five years, enlighten their historical background, cite drawbacks and benefits, and delimit possible future aspects.

Keywords : Molecular Medicine; Multiomics; Methodological Techniques.

References :

1. Blanco, A.; Blanco, G. The genetic information (I). in Medical Biochemistry 501–534 (Elsevier, 2022). doi:10.1016/B978-0-323-91599-1.00017-1.
2. Ronai, I. How molecular techniques are developed from natural systems. Genetics 224, (2023).
3. Cardoso, J. R., Pereira, L. M., Iversen, M. D. & Ramos, A. L. What is gold standard and what is ground truth? Dental Press J Orthod 19, 27–30 (2014).
4. Morange, M. The Historiography of Molecular Biology. in 1–20 (2018). doi:10.1007/978-3-319-74456-8_11-1.
5. D’Adamo, G. L., Widdop, J. T. & Giles, E. M. The future is now? Clinical and translational aspects of “Omics” technologies. Immunol Cell Biol 99, 168–176 (2021).
6. Dai, X. & Shen, L. Advances and Trends in Omics Technology Development. Front Med (Lausanne) 9, (2022).
7. Vogeser, M. & Bendt, A. K. From research cohorts to the patient – a role for “omics” in diagnostics and laboratory medicine? Clinical Chemistry and Laboratory Medicine (CCLM) 61, 974–980 (2023).
8. Habibzadeh, F. On determining the sensitivity and specificity of a new diagnostic test through comparing its results against a non-gold-standard test. Biochem Med (Zagreb) 33, 5–9 (2023).
9. Parikh, R., Mathai, A., Parikh, S., Chandra Sekhar, G. & Thomas, R. Understanding and using sensitivity, specificity and predictive values. Indian J Ophthalmol 56, 45 (2008).
10. Morshedzadeh, F. et al. An Update on the Application of CRISPR Technology in Clinical Practice. Mol Biotechnol 66, 179–197 (2024).
11. Paul, B. & Montoya, G. CRISPR-Cas12a: Functional overview and applications. Biomed J 43, 8–17 (2020).
12. Ishino, Y., Shinagawa, H., Makino, K., Amemura, M. & Nakata, A. Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product. J Bacteriol 169, 5429–5433 (1987).
13. Ishino, Y., Krupovic, M. & Forterre, P. History of CRISPR-Cas from Encounter with a Mysterious Repeated Sequence to Genome Editing Technology. J Bacteriol 200, (2018).
14. Gostimskaya, I. CRISPR–Cas9: A History of Its Discovery and Ethical Considerations of Its Use in Genome Editing. Biochemistry (Moscow) 87, 777–788 (2022).
15. Liu, G., Lin, Q., Jin, S. & Gao, C. The CRISPR-Cas toolbox and gene editing technologies. Mol Cell 82, 333–347 (2022).
16. Hillary, V. E. & Ceasar, S. A. A Review on the Mechanism and Applications of CRISPR/Cas9/Cas12/Cas13/Cas14 Proteins Utilized for Genome Engineering. Mol Biotechnol 65, 311–325 (2023).
17. Khoshandam, M., Soltaninejad, H., Mousazadeh, M., Hamidieh, A. A. & Hosseinkhani, S. Clinical applications of the CRISPR/Cas9 genome-editing system: Delivery options and challenges in precision medicine. Genes Dis 11, 268–282 (2024).
18. Rahbaran, M. et al. Cloning and Embryo Splitting in Mammalians: Brief History, Methods, and Achievements. Stem Cells Int 2021, 1–11 (2021).
19. National Academy of Sciences (US), National Academy of Engineering (US), Institute of Medicine (US), National Research Council (US) & Committee on Science, E. and P. P. Scientific and Medical Aspects of Human Reproductive Cloning. (National Academies Press, Washington, D.C., 2002). doi:10.17226/10285.
20. McGrath, J. & Solter, D. Completion of mouse embryogenesis requires both the maternal and paternal genomes. Cell 37, 179–83 (1984).
21. Campbell, K. H. S., McWhir, J., Ritchie, W. A. & Wilmut, I. Sheep cloned by nuclear transfer from a cultured cell line. Nature 380, 64–66 (1996).
22. Bonetti, G. et al. Human Cloning: Biology, Ethics, and Social Implications. Clin Ter 174, 230–235 (2023).
23. Tadesse, T., Muluneh, T. & Abraham, S. Agricultural and Biomedical Application of Animal Cloning: Review. Journal of Natural Sciences Research 8, (2018).
24. Alkorta, I., Beriain, I. M. & Rodríguez-Arias, D. Cloning and the Oviedo Convention: The Socio-cultural Construction of Regulation. in The Global Dynamics of Regenerative Medicine 150–168 (Palgrave Macmillan UK, London, 2013). doi:10.1057/9781137026552_6.
25. Kashim, M. I. A. M. et al. Animal cloning and consumption of its by-products: A scientific and Islamic perspectives. Saudi J Biol Sci 28, 2995–3000 (2021).
26. Woodfint, R. M., Hamlin, E. & Lee, K. Avian Bioreactor Systems: A Review. Mol Biotechnol 60, 975–983 (2018).
27. Lin, Z., Nielsen, J. & Liu, Z. Bioprospecting Through Cloning of Whole Natural Product Biosynthetic Gene Clusters. Front Bioeng Biotechnol 8, (2020).
28. Behjati, S. & Tarpey, P. S. What is next generation sequencing? Arch Dis Child Educ Pract Ed 98, 236–238 (2013).
29. McCombie, W. R., McPherson, J. D. & Mardis, E. R. Next-Generation Sequencing Technologies. Cold Spring Harb Perspect Med 9, a036798 (2019).
30. Larson, N. B., Oberg, A. L., Adjei, A. A. & Wang, L. A Clinician’s Guide to Bioinformatics for Next-Generation Sequencing. Journal of Thoracic Oncology 18, 143–157 (2023).
31. Sanger, F., Nicklen, S. & Coulson, A. R. DNA sequencing with chain-terminating inhibitors. Proceedings of the National Academy of Sciences 74, 5463–5467 (1977).
32. Maxam, A. M. & Gilbert, W. A new method for sequencing DNA. Proceedings of the National Academy of Sciences 74, 560–564 (1977).
33. Gupta, N. & Verma, V. K. Next-Generation Sequencing and Its Application: Empowering in Public Health Beyond Reality. in 313–341 (2019). doi:10.1007/978-981-13-8844-6_15.
34. Mandlik, J. S., Patil, A. S. & Singh, S. Next-Generation Sequencing (NGS): Platforms and Applications. J Pharm Bioallied Sci 16, S41–S45 (2024).
35. Hilt, E. E. & Ferrieri, P. Next Generation and Other Sequencing Technologies in Diagnostic Microbiology and Infectious Diseases. Genes (Basel) 13, 1566 (2022).
36. Haque, A., Engel, J., Teichmann, S. A. & Lönnberg, T. A practical guide to single-cell RNA-sequencing for biomedical research and clinical applications. Genome Med 9, 75 (2017).
37. Tang, F. et al. mRNA-Seq whole-transcriptome analysis of a single cell. Nat Methods 6, 377–382 (2009).
38. Jovic, D. et al. Single‐cell RNA sequencing technologies and applications: A brief overview. Clin Transl Med 12, (2022).
39. Khozyainova, A. A. et al. Complex Analysis of Single-Cell RNA Sequencing Data. Biochemistry (Moscow) 88, 231–252 (2023).
40. Zhang, Y. et al. Single‐cell RNA sequencing in cancer research. Journal of Experimental & Clinical Cancer Research 40, 81 (2021).
41. Van de Sande, B. et al. Applications of single-cell RNA sequencing in drug discovery and development. Nat Rev Drug Discov 22, 496–520 (2023).
42. Ziegenhain, C., Hendriks, G.-J., Hagemann-Jensen, M. & Sandberg, R. Molecular spikes: a gold standard for single-cell RNA counting. Nat Methods 19, 560–566 (2022).
43. Khehra, N., Padda, I. S. & Swift, C. J. Polymerase Chain Reaction (PCR). (2024).
44. Artika, I. M., Dewi, Y. P., Nainggolan, I. M., Siregar, J. E. & Antonjaya, U. Real-Time Polymerase Chain Reaction: Current Techniques, Applications, and Role in COVID-19 Diagnosis. Genes (Basel) 13, 2387 (2022).
45. Zeron-Medina, J., Ochoa de Olza, M., Braña, I. & Rodon, J. The Personalization of Therapy: Molecular Profiling Technologies and Their Application. Semin Oncol 42, 775–787 (2015).
46. Zhu, H. et al. PCR Past, Present and Future. Biotechniques 69, 317–325 (2020).
47. Harshitha, R. & Arunraj, D. R. Real‐time quantitative PCR : A tool for absolute and relative quantification. Biochemistry and Molecular Biology Education 49, 800–812 (2021).
48. Kadja, T., Liu, C., Sun, Y. & Chodavarapu, V. P. Low-Cost, Real-Time Polymerase Chain Reaction System for Point-of-Care Medical Diagnosis. Sensors 22, 2320 (2022).
49. Hu, B. et al. Therapeutic siRNA: state of the art. Signal Transduct Target Ther 5, 101 (2020).
50. Traber, G. M. & Yu, A.-M. RNAi-Based Therapeutics and Novel RNA Bioengineering Technologies. Journal of Pharmacology and Experimental Therapeutics 384, 133–154 (2023).
51. Friedrich, M. & Aigner, A. Therapeutic siRNA: State-of-the-Art and Future Perspectives. BioDrugs 36, 549–571 (2022).
52. Zhang, C. & Zhang, B. RNA therapeutics: updates and future potential. Sci China Life Sci 66, 12–30 (2023).
53. Koeppe, S., Kawchuk, L. & Kalischuk, M. RNA Interference Past and Future Applications in Plants. Int J Mol Sci 24, 9755 (2023).
54. Ho, P. T. B., Clark, I. M. & Le, L. T. T. MicroRNA-Based Diagnosis and Therapy. Int J Mol Sci 23, 7167 (2022).
55. Soleymani, F., Paquet, E., Viktor, H., Michalowski, W. & Spinello, D. Protein–protein interaction prediction with deep learning: A comprehensive review. Comput Struct Biotechnol J 20, 5316–5341 (2022).
56. Fionda, V. Networks in Biology. in Encyclopedia of Bioinformatics and Computational Biology 915–921 (Elsevier, 2019). doi:10.1016/B978-0-12-809633-8.20420-2.
57. Lu, H. et al. Recent advances in the development of protein–protein interactions modulators: mechanisms and clinical trials. Signal Transduct Target Ther 5, 213 (2020).
58. Armingol, E., Officer, A., Harismendy, O. & Lewis, N. E. Deciphering cell–cell interactions and communication from gene expression. Nat Rev Genet 22, 71–88 (2021).
59. Roy, S., Manners, H. N., Elmsallati, A. & Kalita, J. K. Alignment of Protein-Protein Interaction Networks. in Encyclopedia of Bioinformatics and Computational Biology 997–1015 (Elsevier, 2019). doi:10.1016/B978-0-12-809633-8.20429-9.
60. Jurkovic, C.-M. & Boisvert, F.-M. Evolution of techniques and tools for replication fork proteome and protein interaction studies. Biochemistry and Cell Biology 102, 135–144 (2024).
61. Wang, S. et al. Protein‐protein interaction networks as miners of biological discovery. Proteomics 22, (2022).
62. Chan, A. M., Goodis, C. C., Pommier, E. G. & Fletcher, S. Recent applications of covalent chemistries in protein–protein interaction inhibitors. RSC Med Chem 13, 921–928 (2022).
63. Meftahi, G. H., Bahari, Z., Zarei Mahmoudabadi, A., Iman, M. & Jangravi, Z. Applications of western blot technique: From bench to bedside. Biochemistry and Molecular Biology Education 49, 509–517 (2021).
64. Begum, H., Murugesan, P. & Tangutur, A. D. Western Blotting: A Powerful Staple In Scientific and Biomedical Research. Biotechniques 73, 58–69 (2022).
65. Towbin, H., Staehelin, T. & Gordon, J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A 76, 4350–4 (1979).
66. Sule, R., Rivera, G. & Gomes, A. V. Western Blotting (immunoblotting): History, Theory, Uses, Protocol and Problems. Biotechniques 75, 99–114 (2023).
67. Pillai-Kastoori, L., Schutz-Geschwender, A. R. & Harford, J. A. A systematic approach to quantitative Western blot analysis. Anal Biochem 593, 113608 (2020).
68. Tsurusawa, N. et al. Modified ELISA for Ultrasensitive Diagnosis. J Clin Med 10, 5197 (2021).
69. Alhajj, M., Zubair, M. & Farhana, A. Enzyme Linked Immunosorbent Assay. (2024).
70. Portilho, A. I., Gimenes Lima, G. & De Gaspari, E. Enzyme-Linked Immunosorbent Assay: An Adaptable Methodology to Study SARS-CoV-2 Humoral and Cellular Immune Responses. J Clin Med 11, 1503 (2022).
71. Sakamoto, S. et al. Enzyme-linked immunosorbent assay for the quantitative/qualitative analysis of plant secondary metabolites. J Nat Med 72, 32–42 (2018).
72. Alhajj, M., Zubair, M. & Farhana, A. Enzyme Linked Immunosorbent Assay. (2024).
73. Hayrapetyan, H., Tran, T., Tellez-Corrales, E. & Madiraju, C. Enzyme-Linked Immunosorbent Assay: Types and Applications. Methods Mol Biol 2612, 1–17 (2023).
74. Peng, P. et al. Emerging ELISA derived technologies for in vitro diagnostics. TrAC Trends in Analytical Chemistry 152, 116605 (2022).
75. Barsanti, L., Birindelli, L., Sbrana, F., Lombardi, G. & Gualtieri, P. Advanced Microscopy Techniques for Molecular Biophysics. Int J Mol Sci 24, 9973 (2023).
76. Jacquemet, G., Carisey, A. F., Hamidi, H., Henriques, R. & Leterrier, C. The cell biologist’s guide to super-resolution microscopy. J Cell Sci 133, (2020).
77. Siegerist, F., Drenic, V., Koppe, sssssssssssssssssssssT.-M., Telli, N. & Endlich, N. Super-Resolution Microscopy: A Technique to Revolutionize Research and Diagnosis of Glomerulopathies. Glomerular Dis 3, 19–28 (2022).
78. Badawi, Y. & Nishimune, H. Super-resolution microscopy for analyzing neuromuscular junctions and synapses. Neurosci Lett 715, 134644 (2020).
79. Elliott, A. D. Confocal Microscopy: Principles and Modern Practices. Curr Protoc Cytom 92, (2020).
80. Wu, Y. et al. Multiview confocal super-resolution microscopy. Nature 600, 279–284 (2021).
81. Mendes, A., Heil, H. S., Coelho, S., Leterrier, C. & Henriques, R. Mapping molecular complexes with super-resolution microscopy and single-particle analysis. Open Biol 12, (2022).
82. Cardellini, J., Balestri, A., Montis, C. & Berti, D. Advanced Static and Dynamic Fluorescence Microscopy Techniques to Investigate Drug Delivery Systems. Pharmaceutics 13, 861 (2021).
83. Kaur, A., Kaur, P. & Ahuja, S. Förster resonance energy transfer (FRET) and applications thereof. Analytical Methods 12, 5532–5550 (2020).
84. Menaesse, A. et al. Simplified Instrument Calibration for Wide‐Field Fluorescence Resonance Energy Transfer (FRET) Measured by the Sensitized Emission Method. Cytometry Part A 99, 407–416 (2021).
85. Wallace, B. & Atzberger, P. J. Förster resonance energy transfer: Role of diffusion of fluorophore orientation and separation in observed shifts of FRET efficiency. PLoS One 12, e0177122 (2017).
86. Shrestha, D., Jenei, A., Nagy, P., Vereb, G. & Szöllősi, J. Understanding FRET as a Research Tool for Cellular Studies. Int J Mol Sci 16, 6718–6756 (2015).
87. Förster, Th. Zwischenmolekulare Energiewanderung und Fluoreszenz. Ann Phys 437, 55–75 (1948).
88. Agam, G. et al. Reliability and accuracy of single-molecule FRET studies for characterization of structural dynamics and distances in proteins. Nat Methods 20, 523–535 (2023).
89. Hirata, E. & Kiyokawa, E. Future Perspective of Single-Molecule FRET Biosensors and Intravital FRET Microscopy. Biophys J 111, 1103–1111 (2016).
90. Varghese, S. S., Zhu, Y., Davis, T. J. & Trowell, S. C. FRET for lab-on-a-chip devices — current trends and future prospects. Lab Chip 10, 1355 (2010).
91. Lim, J., Petersen, M., Bunz, M., Simon, C. & Schindler, M. Flow cytometry based-FRET: basics, novel developments and future perspectives. Cellular and Molecular Life Sciences 79, 217 (2022).
92. Shen, Y. et al. FRET-based innovative assays for precise detection of the residual heavy metals in food and agriculture-related matrices. Coord Chem Rev 469, 214676 (2022).
93. Sorzano, C. O. S. & Carazo, J. M. Cryo-Electron Microscopy: The field of 1,000+ methods. J Struct Biol 214, 107861 (2022).
94. Leung, M. R. & Zeev‐Ben‐Mordehai, T. Cryo‐electron microscopy of cholinesterases, present and future. J Neurochem 158, 1236–1243 (2021).
95. Benjin, X. & Ling, L. Developments, applications, and prospects of cryo‐electron microscopy. Protein Science 29, 872–882 (2020).
96. Renaud, J.-P. et al. Cryo-EM in drug discovery: achievements, limitations and prospects. Nat Rev Drug Discov 17, 471–492 (2018).
97. Danev, R., Yanagisawa, H. & Kikkawa, M. Cryo-Electron Microscopy Methodology: Current Aspects and Future Directions. Trends Biochem Sci 44, 837–848 (2019).
98. Ştefan, A. E. et al. Tissue microarrays – brief history, techniques and clinical future. Romanian Journal of Morphology and Embryology 61, 1077–1083 (2021).
99. Kaur, R., A, N., Bansal, R., Saxena, S. & Rai, B. Tissue microarray- A review. International Journal of Oral Health Dentistry 4, 152–155 (2020).
100. Zhang, W., Qiu, F., Jiang, Q., Liu, S. & Xiong, Z. Simple method for constructing and repairing tissue microarrays using simple equipment. Journal of International Medical Research 49, 030006052110001 (2021).
101. Oyejide, L., Mendes, O. R. & Mikaelian, I. Molecular Pathology. in A Comprehensive Guide to Toxicology in Nonclinical Drug Development 407–445 (Elsevier, 2017). doi:10.1016/B978-0-12-803620-4.00016-5.
102. Cui, M., Cheng, C. & Zhang, L. High-throughput proteomics: a methodological mini-review. Laboratory Investigation 102, 1170–1181 (2022).
103. Robinson, J. P., Ostafe, R., Iyengar, S. N., Rajwa, B. & Fischer, R. Flow Cytometry: The Next Revolution. Cells 12, 1875 (2023).
104. D’Amato Figueiredo, M. V., Alexiou, G. A., Vartholomatos, G. & Rehder, R. Advances in Intraoperative Flow Cytometry. Int J Mol Sci 23, 13430 (2022).
105. Robinson, J. P. Flow Cytometry: Past and Future. Biotechniques 72, 159–169 (2022).
106. Welsh, J. A. et al. A compendium of single extracellular vesicle flow cytometry. J Extracell Vesicles 12, (2023).
107. Weiskirchen, S., Schröder, S. K., Buhl, E. M. & Weiskirchen, R. A Beginner’s Guide to Cell Culture: Practical Advice for Preventing Needless Problems. Cells 12, 682 (2023).
108. Kapałczyńska, M. et al. 2D and 3D cell cultures – a comparison of different types of cancer cell cultures. Archives of Medical Science (2016) doi:10.5114/aoms.2016.63743.
109. Zuščíková, L., Greifová, H., Bažány, D., Lukáč, N. & Jambor, T. Current approaches and techniques of 3D cell culture systems: a review. Archives of Ecotoxicology 6, 22–27 (2024).
110. Ylostalo, J. H. 3D Stem Cell Culture. Cells 9, 2178 (2020).
111. Langhans, S. A. Three-Dimensional in Vitro Cell Culture Models in Drug Discovery and Drug Repositioning. Front Pharmacol 9, (2018).
112. Ryu, N.-E., Lee, S.-H. & Park, H. Spheroid Culture System Methods and Applications for Mesenchymal Stem Cells. Cells 8, 1620 (2019).
113. Tatullo, M. et al. Organoids in Translational Oncology. J Clin Med 9, 2774 (2020).
114. Yan, J., Li, Z., Guo, J., Liu, S. & Guo, J. Organ-on-a-chip: A new tool for in vitro research. Biosens Bioelectron 216, 114626 (2022).
115. Leung, C. M. et al. A guide to the organ-on-a-chip. Nature Reviews Methods Primers 2, 33 (2022).
116. Sonagra, A. D. & Dholariya, S. J. Electrophoresis. (2024).
117. Maity, A., Kesh, S. S., Palai, S. & Egbuna, C. Electrophoretic techniques. in Analytical Techniques in Biosciences 59–72 (Elsevier, 2022). doi:10.1016/B978-0-12-822654-4.00007-5.
118. Gummadi, S. & Kandula, V. N. A review on electrophoresis, capillary electrophoresis and hyphenations. Int J Pharm Sci Res 11, 6038–6056 (2020).
119. Lee, P. Y., Saraygord-Afshari, N. & Low, T. Y. The evolution of two-dimensional gel electrophoresis - from proteomics to emerging alternative applications. J Chromatogr A 1615, 460763 (2020).
120. Li, Y., Miao, S., Tan, J., Zhang, Q. & Chen, D. D. Y. Capillary Electrophoresis: A Three-Year Literature Review. Anal Chem 96, 7799–7816 (2024).
121. Hassan, S. Microchip Electrophoresis. Encyclopedia 1, 30–41 (2020).
122. Amatori, S. & Fanelli, M. The Current State of Chromatin Immunoprecipitation (ChIP) from FFPE Tissues. Int J Mol Sci 23, 1103 (2022).
123. Huang, X., Pan, Q., Lin, Y., Gu, T. & Li, Y. A native chromatin immunoprecipitation (ChIP) protocol for studying histone modifications in strawberry fruits. Plant Methods 16, 10 (2020).
124. Saini, A., Rawat, Y., Jain, K. & Mani, I. State-of-the-art techniques to study epigenetics. in 23–50 (2023). doi:10.1016/bs.pmbts.2023.02.004.
125. Yadav, M. P. & Kundra, V. Chromatin Immunoprecipitation and Library Preparation: a Powerful Tool to Unravel the Epigenome. Biotechniques 75, 7–10 (2023).
126. Nakato, R. & Sakata, T. Methods for ChIP-seq analysis: A practical workflow and advanced applications. Methods 187, 44–53 (2021).
127. Zou, Z., Iwata, M., Yamanishi, Y. & Oki, S. Epigenetic landscape of drug responses revealed through large-scale ChIP-seq data analyses. BMC Bioinformatics 23, 51 (2022).
128. Anwar, F., Asrafuzzaman, Amin, K. F. & Hoque, M. E. Tools and techniques for characterizing sustainable hydrogels. in Sustainable Hydrogels 47–77 (Elsevier, 2023). doi:10.1016/B978-0-323-91753-7.00014-4.
129. Kaliva, M. & Vamvakaki, M. Nanomaterials characterization. in Polymer Science and Nanotechnology 401–433 (Elsevier, 2020). doi:10.1016/B978-0-12-816806-6.00017-0.
130. Phạm, T. T. T. & Rainey, J. K. On-cell nuclear magnetic resonance spectroscopy to probe cell surface interactions. Biochemistry and Cell Biology 99, 683–692 (2021).
131. Wishart, D. S. et al. NMR and Metabolomics—A Roadmap for the Future. Metabolites 12, 678 (2022).
132. Joss, D. & Häussinger, D. Design and applications of lanthanide chelating tags for pseudocontact shift NMR spectroscopy with biomacromolecules. Prog Nucl Magn Reson Spectrosc 114–115, 284–312 (2019).
133. Shukla, V. K., Heller, G. T. & Hansen, D. F. Biomolecular NMR spectroscopy in the era of artificial intelligence. Structure 31, 1360–1374 (2023).
134. Pintér, G. et al. Real-time nuclear magnetic resonance spectroscopy in the study of biomolecular kinetics and dynamics. Magnetic Resonance 2, 291–320 (2021).
135. Ude, A., Afi-Leslie, K., Okeke, K. & Ogbodo, E. Trypan Blue Exclusion Assay, Neutral Red, Acridine Orange and Propidium Iodide. in Cytotoxicity - Understanding Cellular Damage and Response (IntechOpen, 2023). doi:10.5772/intechopen.105699.
136. Aslantürk, Ö. S. In Vitro Cytotoxicity and Cell Viability Assays: Principles, Advantages, and Disadvantages. in Genotoxicity - A Predictable Risk to Our Actual World (InTech, 2018). doi:10.5772/intechopen.71923.
137. Kamiloglu, S., Sari, G., Ozdal, T. & Capanoglu, E. Guidelines for cell viability assays. Food Front 1, 332–349 (2020).
138. Larsson, P. et al. Optimization of cell viability assays to improve replicability and reproducibility of cancer drug sensitivity screens. Sci Rep 10, 5798 (2020).
139. World Health Organization & Food and Agriculture Organization of the United Nations. Principles and Methods for the Risk Assessment of Chemicals in Food. (2009).
140. Beal, M. A. et al. Quantitative in vitro to in vivo extrapolation of genotoxicity data provides protective estimates of in vivo dose. Environ Mol Mutagen 64, 105–122 (2023).
141. Dusinska, M., Rundén-Pran, E., Schnekenburger, J. & Kanno, J. Toxicity Tests: In Vitro and In Vivo. in Adverse Effects of Engineered Nanomaterials 51–82 (Elsevier, 2017). doi:10.1016/B978-0-12-809199-9.00003-3.
142. Marchetti, F. et al. Error-corrected next generation sequencing – Promises and challenges for genotoxicity and cancer risk assessment. Mutation Research/Reviews in Mutation Research 792, 108466 (2023).
143. Menz, J. et al. Genotoxicity assessment: opportunities, challenges and perspectives for quantitative evaluations of dose–response data. Arch Toxicol 97, 2303–2328 (2023).
144. Bhinder, B., Gilvary, C., Madhukar, N. S. & Elemento, O. Artificial Intelligence in Cancer Research and Precision Medicine. Cancer Discov 11, 900–915 (2021).
145. Schwalbe, N. & Wahl, B. Artificial intelligence and the future of global health. The Lancet 395, 1579–1586 (2020).
146. Yan, X., Liu, X., Zhao, C. & Chen, G.-Q. Applications of synthetic biology in medical and pharmaceutical fields. Signal Transduct Target Ther 8, 199 (2023).
147. Liu, A. P. et al. The living interface between synthetic biology and biomaterial design. Nat Mater 21, 390–397 (2022).
148. Wu, Q. et al. Organ-on-a-chip: recent breakthroughs and future prospects. Biomed Eng Online 19, 9 (2020).
149. Picollet-D’hahan, N., Zuchowska, A., Lemeunier, I. & Le Gac, S. Multiorgan-on-a-Chip: A Systemic Approach To Model and Decipher Inter-Organ Communication. Trends Biotechnol 39, 788–810 (2021).