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Impedimetric CRISPR-dCas9 Based Biosensor System for Sickle Cell Anemia Mutation

Year 2022, Volume: 9 Issue: 3, 631 - 638, 31.08.2022
https://doi.org/10.18596/jotcsa.1033237

Abstract

Sickle cell anemia is one of the single point mutation diseases with symptoms such as stroke, lethargy, chronic anemia, and increased mortality, and it causes red blood cells to become sickle-shaped. In the study, a biosensor system was developed to detect this mutation quickly and cost-effectively. This biosensor system was prepared by forming a SAM layer with 4-Aminothiophenol (4-ATP) on the gold electrode, and coating it with amino graphene. It was then modified with SG-RNA with the sequence of the target mutation after CRISPR-dCas9 immobilization. The nanomaterial used in the preparation of the biosensor increased the sensitivity of the method by increasing the surface area. The biosensor prepared in this way was optimized and made to perform DNA analysis. As a measurement method, electrochemical impedance spectroscopy (EIS) was used. Electrochemical measurements were carried out in 50 mM pH 7.0 phosphate buffer solution, which includes 5 mM Fe(CN)64- /3- and 10 mM KCl, as redox probe solution by CV and EIS in this redox probe solution. EIS parameters were 10,000–0.05 Hz frequency, 10 mV AC and 180 mV DC potentials, and CV parameters were between - 0.2 to 0.5 V potential, 100 mV/s scan rate for 5 cycles. The DNA measurement time of the biosensor system was determined by the chronoimpedance measurements taken by applying a frequency of 500 Hz under 200 mV DC current. Measurement time of the biosensor was found to be 100 seconds. With the CRISPR-Cas9 based electrochemical biosensor system, which gives faster results compared to the measurement methods in the literature, a linear measurement between 40 pM and 1000 pM with a length of 400 base pairs was taken.

Thanks

The author want to thank Center for Fabrication and Applications of Electronic Materials at Dokuz Eylul University where characterization measurements were performed.

References

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  • 2. Kasiri M, Rahaie M. A visible and colorimetric nanobiosensor based on DNA-CuO nanoparticle for detection of single nucleotide polymorphism involved in sickle cell anemia disease. Materials Today Communications. 2021 Jun;27:102423.
  • 3. Sahu M, Biswas AK, Uma K. Detection of Sickle Cell Anemia in Red Blood Cell. A International Journal of Engineering and Applied Sciences (IJEAS). 2015;2(3).
  • 4. Brazaca LC, Bramorski CB, Cancino-Bernardi J, Janegitz BC, Zucolotto V. A Genosensor for Sickle Cell Anemia Trait Determination. Electroanalysis. 2017 Mar;29(3):773–7.
  • 5. Barrett AN, McDonnell TCR, Chan KCA, Chitty LS. Digital PCR Analysis of Maternal Plasma for Noninvasive Detection of Sickle Cell Anemia. Clinical Chemistry. 2012 Jun 1;58(6):1026–32.
  • 6. Galvão AF, Petta T, Flamand N, Bollela VR, Silva CL, Jarduli LR, et al. Plasma eicosanoid profiles determined by high-performance liquid chromatography coupled with tandem mass spectrometry in stimulated peripheral blood from healthy individuals and sickle cell anemia patients in treatment. Analytical and Bioanalytical Chemistry. 2016 May;408(13):3613–23.
  • 7. Ali TH, Alamiry AAN, Majeed MN. Detection of hemoglobinopathies in hypochromic, microcytic and sickeled cell blood films by hemoglobin electrophoresis. University of Thi-Qar Journal Of Medicine. 2011;5(1):139–48.
  • 8. Tothill IE. Biosensors for cancer markers diagnosis. Seminars in Cell & Developmental Biology. 2009 Feb;20(1):55–62.
  • 9. Uygun ZO, Yeniay L, Girgin Sağın F. CRISPR-dCas9 powered impedimetric biosensor for label-free detection of circulating tumor DNAs. Analytica Chimica Acta. 2020 Jul;1121:35–41.
  • 10. Huang M, Zhou X, Wang H, Xing D. Clustered Regularly Interspaced Short Palindromic Repeats/Cas9 Triggered Isothermal Amplification for Site-Specific Nucleic Acid Detection. Analytical Chemistry 2018 Feb 6;90(3):2193–200. 11. Mehrotra P. Biosensors and their applications – A review. Journal of Oral Biology and Craniofacial Research. 2016 May;6(2):153–9.
  • 12. Uygun ZO, Sezgintürk MK. A novel, ultra sensible biosensor built by layer-by-layer covalent attachment of a receptor for diagnosis of tumor growth. Analytica Chimica Acta. 2011 Nov 14;706(2):343–8.
  • 13. Uygun ZO, Ertuğrul Uygun HD. A short footnote: Circuit design for faradaic impedimetric sensors and biosensors. Sensors and Actuators B: Chemical. 2014 Oct;202:448–53.
  • 14. Chaki NK, Vijayamohanan K. Self-assembled monolayers as a tunable platform for biosensor applications. Biosensors and Bioelectronics. 2002 Jan;17(1–2):1–12.
  • 15. Uygun ZO, Şahin Ç, Yılmaz M, Akçay Y, Akdemir A, Sağın F. Fullerene-PAMAM(G5) composite modified impedimetric biosensor to detect Fetuin-A in real blood samples. Analytical Biochemistry. 2018 Feb;542:11–5.
  • 16. Ertuğrul Uygun HD, Uygun ZO, Canbay E, Gi̇rgi̇n Sağın F, Sezer E. Non-invasive cortisol detection in saliva by using molecularly cortisol imprinted fullerene-acrylamide modified screen printed electrodes. Talanta. 2020 Jan;206:120225.
  • 17. Uygun ZO, Atay S. Label-free highly sensitive detection of DNA approximate length and concentration by impedimetric CRISPR-dCas9 based biosensor technology. Bioelectrochemistry. 2021 Aug;140:107812.
  • 18. Arishi WA, Alhadrami HA, Zourob M. Techniques for the Detection of Sickle Cell Disease: A Review. Micromachines. 2021 May 5;12(5):519.
Year 2022, Volume: 9 Issue: 3, 631 - 638, 31.08.2022
https://doi.org/10.18596/jotcsa.1033237

Abstract

References

  • 1. Breveglieri G, D’Aversa E, Cosenza LC, Boutou E, Balassopoulou A, Voskaridou E, et al. Detection of the sickle hemoglobin allele using a surface plasmon resonance based biosensor. Sensors and Actuators B: Chemical. 2019 Oct;296:126604.
  • 2. Kasiri M, Rahaie M. A visible and colorimetric nanobiosensor based on DNA-CuO nanoparticle for detection of single nucleotide polymorphism involved in sickle cell anemia disease. Materials Today Communications. 2021 Jun;27:102423.
  • 3. Sahu M, Biswas AK, Uma K. Detection of Sickle Cell Anemia in Red Blood Cell. A International Journal of Engineering and Applied Sciences (IJEAS). 2015;2(3).
  • 4. Brazaca LC, Bramorski CB, Cancino-Bernardi J, Janegitz BC, Zucolotto V. A Genosensor for Sickle Cell Anemia Trait Determination. Electroanalysis. 2017 Mar;29(3):773–7.
  • 5. Barrett AN, McDonnell TCR, Chan KCA, Chitty LS. Digital PCR Analysis of Maternal Plasma for Noninvasive Detection of Sickle Cell Anemia. Clinical Chemistry. 2012 Jun 1;58(6):1026–32.
  • 6. Galvão AF, Petta T, Flamand N, Bollela VR, Silva CL, Jarduli LR, et al. Plasma eicosanoid profiles determined by high-performance liquid chromatography coupled with tandem mass spectrometry in stimulated peripheral blood from healthy individuals and sickle cell anemia patients in treatment. Analytical and Bioanalytical Chemistry. 2016 May;408(13):3613–23.
  • 7. Ali TH, Alamiry AAN, Majeed MN. Detection of hemoglobinopathies in hypochromic, microcytic and sickeled cell blood films by hemoglobin electrophoresis. University of Thi-Qar Journal Of Medicine. 2011;5(1):139–48.
  • 8. Tothill IE. Biosensors for cancer markers diagnosis. Seminars in Cell & Developmental Biology. 2009 Feb;20(1):55–62.
  • 9. Uygun ZO, Yeniay L, Girgin Sağın F. CRISPR-dCas9 powered impedimetric biosensor for label-free detection of circulating tumor DNAs. Analytica Chimica Acta. 2020 Jul;1121:35–41.
  • 10. Huang M, Zhou X, Wang H, Xing D. Clustered Regularly Interspaced Short Palindromic Repeats/Cas9 Triggered Isothermal Amplification for Site-Specific Nucleic Acid Detection. Analytical Chemistry 2018 Feb 6;90(3):2193–200. 11. Mehrotra P. Biosensors and their applications – A review. Journal of Oral Biology and Craniofacial Research. 2016 May;6(2):153–9.
  • 12. Uygun ZO, Sezgintürk MK. A novel, ultra sensible biosensor built by layer-by-layer covalent attachment of a receptor for diagnosis of tumor growth. Analytica Chimica Acta. 2011 Nov 14;706(2):343–8.
  • 13. Uygun ZO, Ertuğrul Uygun HD. A short footnote: Circuit design for faradaic impedimetric sensors and biosensors. Sensors and Actuators B: Chemical. 2014 Oct;202:448–53.
  • 14. Chaki NK, Vijayamohanan K. Self-assembled monolayers as a tunable platform for biosensor applications. Biosensors and Bioelectronics. 2002 Jan;17(1–2):1–12.
  • 15. Uygun ZO, Şahin Ç, Yılmaz M, Akçay Y, Akdemir A, Sağın F. Fullerene-PAMAM(G5) composite modified impedimetric biosensor to detect Fetuin-A in real blood samples. Analytical Biochemistry. 2018 Feb;542:11–5.
  • 16. Ertuğrul Uygun HD, Uygun ZO, Canbay E, Gi̇rgi̇n Sağın F, Sezer E. Non-invasive cortisol detection in saliva by using molecularly cortisol imprinted fullerene-acrylamide modified screen printed electrodes. Talanta. 2020 Jan;206:120225.
  • 17. Uygun ZO, Atay S. Label-free highly sensitive detection of DNA approximate length and concentration by impedimetric CRISPR-dCas9 based biosensor technology. Bioelectrochemistry. 2021 Aug;140:107812.
  • 18. Arishi WA, Alhadrami HA, Zourob M. Techniques for the Detection of Sickle Cell Disease: A Review. Micromachines. 2021 May 5;12(5):519.
There are 17 citations in total.

Details

Primary Language English
Subjects Chemical Engineering
Journal Section Articles
Authors

Hilmiye Deniz Ertuğrul Uygun 0000-0003-1631-527X

Publication Date August 31, 2022
Submission Date December 6, 2021
Acceptance Date March 31, 2022
Published in Issue Year 2022 Volume: 9 Issue: 3

Cite

Vancouver Ertuğrul Uygun HD. Impedimetric CRISPR-dCas9 Based Biosensor System for Sickle Cell Anemia Mutation. JOTCSA. 2022;9(3):631-8.