Effects of Microchannel Shape and Ultrasonic Mixing on Microfluidic Padlock Probe Rolling Circle Amplification (RCA) Reactions
Abstract
:1. Introduction
2. Materials and Methods
2.1. Device Fabrication
2.2. Device Cell Culture
2.3. Padlock/RCA Reactions
2.4. Ultrasonic Mixing
2.5. Image Analysis
3. Results and Discussion
3.1. Effect of Microchannel Shape
3.2. Effects of Adsorption
3.3. Effects of Ultrasonic Mixing
4. Conclusions
Author Contributions
Acknowledgments
Conflicts of Interest
References
- Bauman, J.G.; Wiegant, J.; Borst, P.; van Duijn, P. A new method for fluorescence microscopical localization of specific DNA sequences by in situ hybridization of fluorochromelabelled RNA. Exp. Cell Res. 1980, 128, 485–490. [Google Scholar] [CrossRef]
- Wang, F.; Flanagan, J.; Su, N.; Wang, L.C.; Bui, S.; Nielson, A.; Wu, X.; Vo, H.T.; Ma, X.J.; Luo, Y. RNAscope: A novel in situ RNA analysis platform for formalin-fixed, paraffin-embedded tissues. J. Mol. Diagn. 2012, 14, 22–29. [Google Scholar] [CrossRef] [PubMed]
- Wang, Z.; Portier, B.P.; Gruver, A.M.; Bui, S.; Wang, H.; Su, N.; Vo, H.T.; Ma, X.J.; Luo, Y.; Budd, G.T.; et al. Automated quantitative RNA in situ hybridization for resolution of equivocal and heterogeneous ERBB2 (HER2) status in invasive breast carcinoma. J. Mol. Diagn. 2013, 15, 210–219. [Google Scholar] [CrossRef] [PubMed]
- Trcek, T.; Chao, J.A.; Larson, D.R.; Park, H.Y.; Zenklusen, D.; Shenoy, S.M.; Singer, R.H. Single-mRNA counting using fluorescent in situ hybridization in budding yeast. Nat. Protoc. 2012, 7, 408–419. [Google Scholar] [CrossRef] [PubMed]
- Kwon, S. Single-molecule fluorescence in situ hybridization: Quantitative imaging of single RNA molecules. BMB Rep. 2013, 46, 65–72. [Google Scholar] [CrossRef] [PubMed]
- Krzywkowski, T.; Hauling, T.; Nilsson, M. In Situ Single-Molecule RNA Genotyping Using Padlock Probes and Rolling Circle Amplification. Methods Mol. Biol. 2017, 1492, 59–76. [Google Scholar] [CrossRef] [PubMed]
- Krzywkowski, T.; Nilsson, M. Padlock Probes to Detect Single Nucleotide Polymorphisms. Methods Mol. Biol. 2018, 1649, 209–229. [Google Scholar] [CrossRef] [PubMed]
- Larsson, C.; Koch, J.; Nygren, A.; Janssen, G.; Raap, A.K.; Landegren, U.; Nilsson, M. In situ genotyping individual DNA molecules by target-primed rolling-circle amplification of padlock probes. Nat. Methods 2004, 1, 227–232. [Google Scholar] [CrossRef] [PubMed]
- Larsson, C.; Grundberg, I.; Soderberg, O.; Nilsson, M. In situ detection and genotyping of individual mRNA molecules. Nat. Methods 2010, 7, 395–397. [Google Scholar] [CrossRef] [PubMed]
- Grundberg, I.; Kiflemariam, S.; Mignardi, M.; Imgenberg-Kreuz, J.; Edlund, K.; Micke, P.; Sundstrom, M.; Sjoblom, T.; Botling, J.; Nilsson, M. In situ mutation detection and visualization of intratumor heterogeneity for cancer research and diagnostics. Oncotarget 2013, 4, 2407–2418. [Google Scholar] [CrossRef] [PubMed]
- Ge, J.; Zhang, L.L.; Liu, S.J.; Yu, R.Q.; Chu, X. A highly sensitive target-primed rolling circle amplification (TPRCA) method for fluorescent in situ hybridization detection of microRNA in tumor cells. Anal. Chem. 2014, 86, 1808–1815. [Google Scholar] [CrossRef] [PubMed]
- Wamsley, H.L.; Barbet, A.F. In situ detection of Anaplasma spp. by DNA target-primed rolling-circle amplification of a padlock probe and intracellular colocalization with immunofluorescently labeled host cell von Willebrand factor. J. Clin. Microbiol. 2008, 46, 2314–2319. [Google Scholar] [CrossRef] [PubMed]
- Sato, K. Microdevice in Cellular Pathology: Microfluidic Platforms for Fluorescence in situ Hybridization and Analysis of Circulating Tumor Cells. Anal. Sci. 2015, 31, 867–873. [Google Scholar] [CrossRef] [PubMed]
- Ghodbane, M.; Stucky, E.C.; Maguire, T.J.; Schloss, R.S.; Shreiber, D.I.; Zahn, J.D.; Yarmush, M.L. Development and validation of a microfluidic immunoassay capable of multiplexing parallel samples in microliter volumes. Lab Chip 2015, 15, 3211–3221. [Google Scholar] [CrossRef] [PubMed]
- Hasegawa, K.; Negishi, R.; Matsumoto, M.; Yohda, M.; Hosokawa, K.; Maeda, M. Specificity of MicroRNA Detection on a Power-free Microfluidic Chip with Laminar Flow-assisted Dendritic Amplification. Anal. Sci. 2017, 33, 171–177. [Google Scholar] [CrossRef] [PubMed]
- Shirai, A.; Nakashima, K.; Sueyoshi, K.; Endo, T.; Hisamoto, H. Fast and Single-step Fluorescence-based Competitive Bioassay Microdevice Combined PDMS Microchannel Arrays Separately Immobilizing Graphene Oxide-Analyte Conjugates and Fluorescently-labelled Receptor Proteins. Anal. Sci. 2017, 33, 969–972. [Google Scholar] [CrossRef] [PubMed]
- Furutani, S.; Nishio, K.; Naruishi, N.; Akazawa-Ogawa, Y.; Hagihara, Y.; Yoshida, Y.; Nagai, H. Rapid Enzyme-linked Immunosorbent Assays for Diagnosis of Diabetes in a Compact Disc-shaped Microfluidic Device. Anal. Sci. 2018, 34, 379–382. [Google Scholar] [CrossRef] [PubMed]
- Neoh, K.H.; Hassan, A.A.; Chen, A.; Sun, Y.; Liu, P.; Xu, K.F.; Wong, A.S.T.; Han, R.P.S. Rethinking liquid biopsy: Microfluidic assays for mobile tumor cells in human body fluids. Biomaterials 2018, 150, 112–124. [Google Scholar] [CrossRef] [PubMed]
- Kuroda, A.; Ishigaki, Y.; Nilsson, M.; Sato, K.; Sato, K. Microfluidics-based in situ padlock/rolling circle amplification system for counting single DNA molecules in a cell. Anal. Sci. 2014, 30, 1107–1112. [Google Scholar] [CrossRef] [PubMed]
- Na, W.; Nam, D.; Lee, H.; Shin, S. Rapid molecular diagnosis of infectious viruses in microfluidics using DNA hydrogel formation. Biosens. Bioelectron. 2018, 108, 9–13. [Google Scholar] [CrossRef] [PubMed]
- Sarkar, S.; Sabhachandani, P.; Konry, T. Isothermal Amplification Strategies for Detection in Microfluidic Devices. Trends Biotechnol. 2017, 35, 186–189. [Google Scholar] [CrossRef] [PubMed]
- Heo, H.Y.; Chung, S.; Kim, Y.T.; Kim, D.H.; Seo, T.S. A valveless rotary microfluidic device for multiplex point mutation identification based on ligation-rolling circle amplification. Biosens. Bioelectron. 2016, 78, 140–146. [Google Scholar] [CrossRef] [PubMed]
- Lee, C.Y.; Chang, C.L.; Wang, Y.N.; Fu, L.M. Microfluidic mixing: A review. Int. J. Mol. Sci. 2011, 12, 3263–3287. [Google Scholar] [CrossRef] [PubMed]
- Chang, B.S.; Mahoney, R.R. Enzyme thermostabilization by bovine serum albumin and other proteins: Evidence for hydrophobic interactions. Biotechnol. Appl. Biochem. 1995, 22, 203–214. [Google Scholar] [PubMed]
- Taylor, T.B.; Winn-Deen, E.S.; Picozza, E.; Woudenberg, T.M.; Albin, M. Optimization of the performance of the polymerase chain reaction in silicon-based microstructures. Nucleic Acids Res. 1997, 25, 3164–3168. [Google Scholar] [CrossRef] [PubMed]
- Bergethon, P.R. The Physical Basis of Biochemistry: The Foundations of Molecular Biophysics, 2nd ed.; Springer Science & Business Media: Berlin, Germany, 2013. [Google Scholar]
- Gaigalas, A.K.; Hubbard, J.B.; McCurley, M.; Woo, S. Diffusion of bovine serum albumin in aqueous solutions. J. Phys. Chem. 1992, 96, 2355–2359. [Google Scholar] [CrossRef]
Name | Oligonucleotide Sequence (5′→3′) | Kind of DNA | Modification 5′ | References |
---|---|---|---|---|
ppMscs | TAAGAAGAGGAATTGCCTTTCCTTTCCTACGACCTCAATGAACATGTTTGGCTCCTCTTCCCATGGGTATGTTGT | Padlock probe | Phosphate | [8] |
Lin33Alexa555 | CCTCAATGCACATGTTTGGCTCC | Detection probe | Alexa Fluor 555 | [8] |
P-KRAS | CC(L)TC(L)TA(L)TT(L)GT(L)TG(L)GA(L)TCATATTCGTC | cDNA primer | - | [9] |
PLP-KRASwtGGT | GGCGTAGGCAAGAGTTCCTGTAGTAAAGTAGCCGTGACTATCGACTGAATCTAAGGTAGTTGGAGCTGGT | Padlock probe | Phosphate | [9] |
DP-3 | AGTAGCCGTGACTATCGACT | Detection probe | Cyanine3 | [9] |
P-ACTB | CG(L)GG(L)CG(L)GC(L)GG(L)ATCGGCAAAG | cDNA primer | - | [10] |
PLP-b-actin_hum | GCCGGCTTCGCGGGCGACGATTCCTCTATGATTACTGACCTATGCGTCTATTTAGTGGAGCCTCTTCTTTACGGCGCCGGCATGTGCAAG | Padlock probe | Phosphate | [10] |
DP-4 | TGCGTCTATTTAGTGGAGCC | Detection probe | Cyanine3 | [10] |
(L) = Locked Nucleic Acid (LNA)-modified base |
Device | Size | Injection Volume (µL) | Chamber Volume (µL) | Bottom Area (cm2) | Volume Per Unit Area (µL/cm2) |
---|---|---|---|---|---|
Well device | 5 mm diameter | 20.0 | 20.0 | 20 | 1.0 |
I-shaped microchannel | width × depth × length, 1 mm × 200 μm × 10 mm | 3.5 | 2.0 | 10 | 0.2 |
Circular-shaped microchamber | 4 mm in diameter, 200 μm in depth | 3.5 | 2.5 | 13 | 0.2 |
© 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
Share and Cite
Ishigaki, Y.; Sato, K. Effects of Microchannel Shape and Ultrasonic Mixing on Microfluidic Padlock Probe Rolling Circle Amplification (RCA) Reactions. Micromachines 2018, 9, 272. https://doi.org/10.3390/mi9060272
Ishigaki Y, Sato K. Effects of Microchannel Shape and Ultrasonic Mixing on Microfluidic Padlock Probe Rolling Circle Amplification (RCA) Reactions. Micromachines. 2018; 9(6):272. https://doi.org/10.3390/mi9060272
Chicago/Turabian StyleIshigaki, Yuri, and Kae Sato. 2018. "Effects of Microchannel Shape and Ultrasonic Mixing on Microfluidic Padlock Probe Rolling Circle Amplification (RCA) Reactions" Micromachines 9, no. 6: 272. https://doi.org/10.3390/mi9060272
APA StyleIshigaki, Y., & Sato, K. (2018). Effects of Microchannel Shape and Ultrasonic Mixing on Microfluidic Padlock Probe Rolling Circle Amplification (RCA) Reactions. Micromachines, 9(6), 272. https://doi.org/10.3390/mi9060272