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2022
Kanik, Fulya Ekiz; Celebi, Iris; Sevenler, Derin; Tanriverdi, Kahraman; Ünlü, Nese Lortlar; Freedman, Jane E.; Ünlü, M. Selim
Attomolar sensitivity microRNA detection using real‑time digital microarrays Journal Article
In: Scientific Reports, vol. 2022, no. 12, pp. 16220, 2022.
Abstract | Links | BibTeX | Tags: Oligo
@article{nokey,
title = {Attomolar sensitivity microRNA detection using real‑time digital microarrays},
author = {Fulya Ekiz Kanik and Iris Celebi and Derin Sevenler and Kahraman Tanriverdi and Nese Lortlar Ünlü and Jane E. Freedman and M. Selim Ünlü},
url = {https://axivend.com/wp-content/uploads/2022/09/Attomolar-sensitivity-microRNA-detection.pdf},
doi = {10.1038/s41598-022-19912-z},
year = {2022},
date = {2022-09-27},
urldate = {2022-09-27},
journal = {Scientific Reports},
volume = {2022},
number = {12},
pages = {16220},
abstract = {MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA‑451a (miR‑451) and miRNA‑223‑3p (miR‑223). We demonstrated improvements in sensitivity in comparison to traditional end‑point assays that employ capture on solid phase support, by implementing real‑time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low‑abundance biomarkers in the presence
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.},
keywords = {Oligo},
pubstate = {published},
tppubtype = {article}
}
MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA‑451a (miR‑451) and miRNA‑223‑3p (miR‑223). We demonstrated improvements in sensitivity in comparison to traditional end‑point assays that employ capture on solid phase support, by implementing real‑time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low‑abundance biomarkers in the presence
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.
Kanik, Fulya Ekiz; Celebi, Iris; Sevenler, Derin; Tanriverdi, Kahraman; Ünlü, Nese Lortlar; Freedman, Jane E.; Ünlü, M. Selim
Attomolar sensitivity microRNA detection using real‑time digital microarrays Journal Article
In: Scientific Reports, vol. 2022, iss. 2, no. 12, pp. 16220, 2022.
Abstract | Links | BibTeX | Tags: Oligo
@article{nokey,
title = {Attomolar sensitivity microRNA detection using real‑time digital microarrays},
author = {Fulya Ekiz Kanik and Iris Celebi and Derin Sevenler and Kahraman Tanriverdi and Nese Lortlar Ünlü and Jane E. Freedman and M. Selim Ünlü},
url = {https://mhg912.p3cdn1.secureserver.net/wp-content/uploads/2022/09/Attomolar-sensitivity-microRNA-detection.pdf},
doi = {10.1038/s41598-022-19912-z},
year = {2022},
date = {2022-09-27},
urldate = {2022-09-27},
booktitle = {Imaging Systems and Applications},
journal = {Scientific Reports},
volume = {2022},
number = {12},
issue = {2},
pages = {16220},
publisher = {Optica Publishing Group},
series = {Biophysics At the Nanoscale},
abstract = {MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA‑451a (miR‑451) and miRNA‑223‑3p (miR‑223). We demonstrated improvements in sensitivity in comparison to traditional end‑point assays that employ capture on solid phase support, by implementing real‑time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low‑abundance biomarkers in the presence
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.},
howpublished = {https://chemrxiv.org/engage/chemrxiv/article-details/6565fc1829a13c4d4730f055},
keywords = {Oligo},
pubstate = {published},
tppubtype = {article}
}
MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA‑451a (miR‑451) and miRNA‑223‑3p (miR‑223). We demonstrated improvements in sensitivity in comparison to traditional end‑point assays that employ capture on solid phase support, by implementing real‑time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low‑abundance biomarkers in the presence
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.
Chiodi, E; Marn, AM; Bakhshpour, M; ans MS Ünlü, N Lortlar Ünlü
The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors Journal Article
In: Polymers, vol. 14, iss. 2, pp. 241, 2022.
Abstract | Links | BibTeX | Tags: Oligo, Protein
@article{nokey,
title = {The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors},
author = {E Chiodi and AM Marn and M Bakhshpour and N Lortlar Ünlü ans MS Ünlü},
editor = {Ick-Soo Kim},
url = {https://www.mdpi.com/2073-4360/14/2/241},
doi = {https://doi.org/10.3390/polym14020241},
year = {2022},
date = {2022-01-07},
urldate = {2022-01-07},
journal = {Polymers},
volume = {14},
issue = {2},
pages = {241},
abstract = {The field of biosensing is in constant evolution, propelled by the need for sensitive, reliable platforms that provide consistent results, especially in the drug development industry, where small molecule characterization is of uttermost relevance. Kinetic characterization of small biochemicals is particularly challenging, and has required sensor developers to find solutions to compensate for the lack of sensitivity of their instruments. In this regard, surface chemistry plays a crucial role. The ligands need to be efficiently immobilized on the sensor surface, and probe distribution, maintenance of their native structure and efficient diffusion of the analyte to the surface need to be optimized. In order to enhance the signal generated by low molecular weight targets, surface plasmon resonance sensors utilize a high density of probes on the surface by employing a thick dextran matrix, resulting in a three-dimensional, multilayer distribution of molecules. Despite increasing the binding signal, this method can generate artifacts, due to the diffusion dependence of surface binding, affecting the accuracy of measured affinity constants. On the other hand, when working with planar surface chemistries, an incredibly high sensitivity is required for low molecular weight analytes, and furthermore the standard method for immobilizing single layers of molecules based on self-assembled monolayers (SAM) of epoxysilane has been demonstrated to promote protein denaturation, thus being far from ideal. Here, we will give a concise overview of the impact of tridimensional immobilization of ligands on label-free biosensors, mostly focusing on the effect of diffusion on binding affinity constants measurements. We will comment on how multilayering of probes is certainly useful in terms of increasing the sensitivity of the sensor, but can cause steric hindrance, mass transport and other diffusion effects. On the other hand, probe monolayers on epoxysilane chemistries do not undergo diffusion effect but rather other artifacts can occur due to probe distortion. Finally, a combination of tridimensional polymeric chemistry and probe monolayer is presented and reviewed, showing advantages and disadvantages over the other two approaches},
keywords = {Oligo, Protein},
pubstate = {published},
tppubtype = {article}
}
The field of biosensing is in constant evolution, propelled by the need for sensitive, reliable platforms that provide consistent results, especially in the drug development industry, where small molecule characterization is of uttermost relevance. Kinetic characterization of small biochemicals is particularly challenging, and has required sensor developers to find solutions to compensate for the lack of sensitivity of their instruments. In this regard, surface chemistry plays a crucial role. The ligands need to be efficiently immobilized on the sensor surface, and probe distribution, maintenance of their native structure and efficient diffusion of the analyte to the surface need to be optimized. In order to enhance the signal generated by low molecular weight targets, surface plasmon resonance sensors utilize a high density of probes on the surface by employing a thick dextran matrix, resulting in a three-dimensional, multilayer distribution of molecules. Despite increasing the binding signal, this method can generate artifacts, due to the diffusion dependence of surface binding, affecting the accuracy of measured affinity constants. On the other hand, when working with planar surface chemistries, an incredibly high sensitivity is required for low molecular weight analytes, and furthermore the standard method for immobilizing single layers of molecules based on self-assembled monolayers (SAM) of epoxysilane has been demonstrated to promote protein denaturation, thus being far from ideal. Here, we will give a concise overview of the impact of tridimensional immobilization of ligands on label-free biosensors, mostly focusing on the effect of diffusion on binding affinity constants measurements. We will comment on how multilayering of probes is certainly useful in terms of increasing the sensitivity of the sensor, but can cause steric hindrance, mass transport and other diffusion effects. On the other hand, probe monolayers on epoxysilane chemistries do not undergo diffusion effect but rather other artifacts can occur due to probe distortion. Finally, a combination of tridimensional polymeric chemistry and probe monolayer is presented and reviewed, showing advantages and disadvantages over the other two approaches
Chiodi, E; Marn, AM; Bakhshpour, M; Ünlü, N Lortlar Ünlü MS
The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors Journal Article
In: Polymers, vol. 14, iss. 2, pp. 241, 2022.
Abstract | Links | BibTeX | Tags: Oligo, Protein
@article{nokeyd,
title = {The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors},
author = {E Chiodi and AM Marn and M Bakhshpour and N Lortlar Ünlü MS Ünlü},
editor = {Ick-Soo Kim},
url = {https://www.mdpi.com/2073-4360/14/2/241},
doi = {https://doi.org/10.3390/polym14020241},
year = {2022},
date = {2022-01-07},
urldate = {2022-01-07},
journal = {Polymers},
volume = {14},
issue = {2},
pages = {241},
abstract = {The field of biosensing is in constant evolution, propelled by the need for sensitive, reliable platforms that provide consistent results, especially in the drug development industry, where small molecule characterization is of uttermost relevance. Kinetic characterization of small biochemicals is particularly challenging, and has required sensor developers to find solutions to compensate for the lack of sensitivity of their instruments. In this regard, surface chemistry plays a crucial role. The ligands need to be efficiently immobilized on the sensor surface, and probe distribution, maintenance of their native structure and efficient diffusion of the analyte to the surface need to be optimized. In order to enhance the signal generated by low molecular weight targets, surface plasmon resonance sensors utilize a high density of probes on the surface by employing a thick dextran matrix, resulting in a three-dimensional, multilayer distribution of molecules. Despite increasing the binding signal, this method can generate artifacts, due to the diffusion dependence of surface binding, affecting the accuracy of measured affinity constants. On the other hand, when working with planar surface chemistries, an incredibly high sensitivity is required for low molecular weight analytes, and furthermore the standard method for immobilizing single layers of molecules based on self-assembled monolayers (SAM) of epoxysilane has been demonstrated to promote protein denaturation, thus being far from ideal. Here, we will give a concise overview of the impact of tridimensional immobilization of ligands on label-free biosensors, mostly focusing on the effect of diffusion on binding affinity constants measurements. We will comment on how multilayering of probes is certainly useful in terms of increasing the sensitivity of the sensor, but can cause steric hindrance, mass transport and other diffusion effects. On the other hand, probe monolayers on epoxysilane chemistries do not undergo diffusion effect but rather other artifacts can occur due to probe distortion. Finally, a combination of tridimensional polymeric chemistry and probe monolayer is presented and reviewed, showing advantages and disadvantages over the other two approaches},
keywords = {Oligo, Protein},
pubstate = {published},
tppubtype = {article}
}
The field of biosensing is in constant evolution, propelled by the need for sensitive, reliable platforms that provide consistent results, especially in the drug development industry, where small molecule characterization is of uttermost relevance. Kinetic characterization of small biochemicals is particularly challenging, and has required sensor developers to find solutions to compensate for the lack of sensitivity of their instruments. In this regard, surface chemistry plays a crucial role. The ligands need to be efficiently immobilized on the sensor surface, and probe distribution, maintenance of their native structure and efficient diffusion of the analyte to the surface need to be optimized. In order to enhance the signal generated by low molecular weight targets, surface plasmon resonance sensors utilize a high density of probes on the surface by employing a thick dextran matrix, resulting in a three-dimensional, multilayer distribution of molecules. Despite increasing the binding signal, this method can generate artifacts, due to the diffusion dependence of surface binding, affecting the accuracy of measured affinity constants. On the other hand, when working with planar surface chemistries, an incredibly high sensitivity is required for low molecular weight analytes, and furthermore the standard method for immobilizing single layers of molecules based on self-assembled monolayers (SAM) of epoxysilane has been demonstrated to promote protein denaturation, thus being far from ideal. Here, we will give a concise overview of the impact of tridimensional immobilization of ligands on label-free biosensors, mostly focusing on the effect of diffusion on binding affinity constants measurements. We will comment on how multilayering of probes is certainly useful in terms of increasing the sensitivity of the sensor, but can cause steric hindrance, mass transport and other diffusion effects. On the other hand, probe monolayers on epoxysilane chemistries do not undergo diffusion effect but rather other artifacts can occur due to probe distortion. Finally, a combination of tridimensional polymeric chemistry and probe monolayer is presented and reviewed, showing advantages and disadvantages over the other two approaches
2021
Chiodi, Elisa; Marn, Allison M; Geib, Matthew T; Ünlü, M Selim
The Role of Surface Chemistry in the Efficacy of Protein and DNA Microarrays for Label-Free Detection: An Overview Journal Article
In: Polymers, vol. 13, iss. 7, pp. 1026, 2021.
Abstract | Links | BibTeX | Tags: Oligo, Protein
@article{nokey,
title = {The Role of Surface Chemistry in the Efficacy of Protein and DNA Microarrays for Label-Free Detection: An Overview},
author = {Elisa Chiodi and Allison M Marn and Matthew T Geib and M Selim Ünlü},
url = {https://www.mdpi.com/2073-4360/13/7/1026/pdf},
doi = {https://doi.org/10.3390/polym13071026},
year = {2021},
date = {2021-01-03},
urldate = {2021-01-03},
journal = {Polymers},
volume = {13},
issue = {7},
pages = {1026},
abstract = {The importance of microarrays in diagnostics and medicine has drastically increased in the last few years. Nevertheless, the efficiency of a microarray-based assay intrinsically depends on the density and functionality of the biorecognition elements immobilized onto each sensor spot. Recently, researchers have put effort into developing new functionalization strategies and technologies which provide efficient immobilization and stability of any sort of molecule. Here, we present an overview of the most widely used methods of surface functionalization of microarray substrates, as well as the most recent advances in the field, and compare their performance in terms of optimal immobilization of the bioreceptor molecules. We focus on label-free microarrays and, in particular, we aim to describe the impact of surface chemistry on two types of microarray-based sensors: microarrays for single particle imaging and for label-free measurements of binding kinetics. Both protein and DNA microarrays are taken into consideration, and the effect of different polymeric coatings on the molecules’ functionalities is critically analyzed.},
keywords = {Oligo, Protein},
pubstate = {published},
tppubtype = {article}
}
The importance of microarrays in diagnostics and medicine has drastically increased in the last few years. Nevertheless, the efficiency of a microarray-based assay intrinsically depends on the density and functionality of the biorecognition elements immobilized onto each sensor spot. Recently, researchers have put effort into developing new functionalization strategies and technologies which provide efficient immobilization and stability of any sort of molecule. Here, we present an overview of the most widely used methods of surface functionalization of microarray substrates, as well as the most recent advances in the field, and compare their performance in terms of optimal immobilization of the bioreceptor molecules. We focus on label-free microarrays and, in particular, we aim to describe the impact of surface chemistry on two types of microarray-based sensors: microarrays for single particle imaging and for label-free measurements of binding kinetics. Both protein and DNA microarrays are taken into consideration, and the effect of different polymeric coatings on the molecules’ functionalities is critically analyzed.
Francesco Damin Elisa Chiodi, Laura Sola
A reliable, label free quality control method for the production of DNA microarrays with clinical applications Journal Article
In: Polymers, vol. 13, iss. 36, pp. 340, 2021.
Abstract | Links | BibTeX | Tags: Oligo
@article{nokey,
title = {A reliable, label free quality control method for the production of DNA microarrays with clinical applications},
author = {Elisa Chiodi, Francesco Damin, Laura Sola, Lucia Ferraro, Dario Brambilla, M Selim Ünlü, Marcella Chiari},
url = {https://www.mdpi.com/2073-4360/13/3/340/pdf},
doi = {https://doi.org/10.3390/polym13030340},
year = {2021},
date = {2021-01-03},
urldate = {2021-01-03},
journal = {Polymers},
volume = {13},
issue = {36},
pages = {340},
abstract = {The manufacture of a very high-quality microarray support is essential for the adoption of this assay format in clinical routine. In fact, poorly surface-bound probes can affect the diagnostic sensitivity or, in worst cases, lead to false negative results. Here we report on a reliable and easy quality control method for the evaluation of spotted probe properties in a microarray test, based on the Interferometric Reflectance Imaging Sensor (IRIS) system, a high-resolution label free technique able to evaluate the variation of the mass bound to a surface. In particular, we demonstrated that the IRIS analysis of microarray chips immediately after probe immobilization can detect the absence of probes, which recognizably causes a lack of signal when performing a test, with clinical relevance, using fluorescence detection. Moreover, the use of the IRIS technique allowed also to determine the optimal concentration of the probe, that has to be immobilized on the surface, to maximize the target recognition, thus the signal, but to avoid crowding effects. Finally, through this preliminary quality inspection it is possible to highlight differences in the immobilization chemistries. In particular, we have compared NHS ester versus click chemistry reactions using two different surface coatings, demonstrating that, in the diagnostic case used as an example (colorectal cancer) a higher probe density does not reflect a higher binding signal, probably because of a crowding effect.},
keywords = {Oligo},
pubstate = {published},
tppubtype = {article}
}
The manufacture of a very high-quality microarray support is essential for the adoption of this assay format in clinical routine. In fact, poorly surface-bound probes can affect the diagnostic sensitivity or, in worst cases, lead to false negative results. Here we report on a reliable and easy quality control method for the evaluation of spotted probe properties in a microarray test, based on the Interferometric Reflectance Imaging Sensor (IRIS) system, a high-resolution label free technique able to evaluate the variation of the mass bound to a surface. In particular, we demonstrated that the IRIS analysis of microarray chips immediately after probe immobilization can detect the absence of probes, which recognizably causes a lack of signal when performing a test, with clinical relevance, using fluorescence detection. Moreover, the use of the IRIS technique allowed also to determine the optimal concentration of the probe, that has to be immobilized on the surface, to maximize the target recognition, thus the signal, but to avoid crowding effects. Finally, through this preliminary quality inspection it is possible to highlight differences in the immobilization chemistries. In particular, we have compared NHS ester versus click chemistry reactions using two different surface coatings, demonstrating that, in the diagnostic case used as an example (colorectal cancer) a higher probe density does not reflect a higher binding signal, probably because of a crowding effect.
Chiodi, Elisa; Marn, Allison M; Geib, Matthew T; Ünlü, M Selim
The Role of Surface Chemistry in the Efficacy of Protein and DNA Microarrays for Label-Free Detection: An Overview Journal Article
In: Polymers, vol. 13, iss. 7, pp. 1026, 2021.
Abstract | Links | BibTeX | Tags: Oligo, Protein
@article{nokeyo,
title = {The Role of Surface Chemistry in the Efficacy of Protein and DNA Microarrays for Label-Free Detection: An Overview},
author = {Elisa Chiodi and Allison M Marn and Matthew T Geib and M Selim Ünlü},
url = {https://www.mdpi.com/2073-4360/13/7/1026/pdf},
doi = {https://doi.org/10.3390/polym13071026},
year = {2021},
date = {2021-01-03},
urldate = {2021-01-03},
journal = {Polymers},
volume = {13},
issue = {7},
pages = {1026},
abstract = {The importance of microarrays in diagnostics and medicine has drastically increased in the last few years. Nevertheless, the efficiency of a microarray-based assay intrinsically depends on the density and functionality of the biorecognition elements immobilized onto each sensor spot. Recently, researchers have put effort into developing new functionalization strategies and technologies which provide efficient immobilization and stability of any sort of molecule. Here, we present an overview of the most widely used methods of surface functionalization of microarray substrates, as well as the most recent advances in the field, and compare their performance in terms of optimal immobilization of the bioreceptor molecules. We focus on label-free microarrays and, in particular, we aim to describe the impact of surface chemistry on two types of microarray-based sensors: microarrays for single particle imaging and for label-free measurements of binding kinetics. Both protein and DNA microarrays are taken into consideration, and the effect of different polymeric coatings on the molecules’ functionalities is critically analyzed.},
keywords = {Oligo, Protein},
pubstate = {published},
tppubtype = {article}
}
The importance of microarrays in diagnostics and medicine has drastically increased in the last few years. Nevertheless, the efficiency of a microarray-based assay intrinsically depends on the density and functionality of the biorecognition elements immobilized onto each sensor spot. Recently, researchers have put effort into developing new functionalization strategies and technologies which provide efficient immobilization and stability of any sort of molecule. Here, we present an overview of the most widely used methods of surface functionalization of microarray substrates, as well as the most recent advances in the field, and compare their performance in terms of optimal immobilization of the bioreceptor molecules. We focus on label-free microarrays and, in particular, we aim to describe the impact of surface chemistry on two types of microarray-based sensors: microarrays for single particle imaging and for label-free measurements of binding kinetics. Both protein and DNA microarrays are taken into consideration, and the effect of different polymeric coatings on the molecules’ functionalities is critically analyzed.
Chiodi, Laura Sola Francesco Damin Elisa
A reliable, label free quality control method for the production of DNA microarrays with clinical applications Journal Article
In: Polymers, vol. 13, iss. 36, pp. 340, 2021.
Abstract | Links | BibTeX | Tags: Oligo
@article{nokeyn,
title = {A reliable, label free quality control method for the production of DNA microarrays with clinical applications},
author = {Laura Sola Francesco Damin Elisa Chiodi},
url = {https://www.mdpi.com/2073-4360/13/3/340/pdf},
doi = {https://doi.org/10.3390/polym13030340},
year = {2021},
date = {2021-01-03},
urldate = {2021-01-03},
journal = {Polymers},
volume = {13},
issue = {36},
pages = {340},
abstract = {The manufacture of a very high-quality microarray support is essential for the adoption of this assay format in clinical routine. In fact, poorly surface-bound probes can affect the diagnostic sensitivity or, in worst cases, lead to false negative results. Here we report on a reliable and easy quality control method for the evaluation of spotted probe properties in a microarray test, based on the Interferometric Reflectance Imaging Sensor (IRIS) system, a high-resolution label free technique able to evaluate the variation of the mass bound to a surface. In particular, we demonstrated that the IRIS analysis of microarray chips immediately after probe immobilization can detect the absence of probes, which recognizably causes a lack of signal when performing a test, with clinical relevance, using fluorescence detection. Moreover, the use of the IRIS technique allowed also to determine the optimal concentration of the probe, that has to be immobilized on the surface, to maximize the target recognition, thus the signal, but to avoid crowding effects. Finally, through this preliminary quality inspection it is possible to highlight differences in the immobilization chemistries. In particular, we have compared NHS ester versus click chemistry reactions using two different surface coatings, demonstrating that, in the diagnostic case used as an example (colorectal cancer) a higher probe density does not reflect a higher binding signal, probably because of a crowding effect.},
keywords = {Oligo},
pubstate = {published},
tppubtype = {article}
}
The manufacture of a very high-quality microarray support is essential for the adoption of this assay format in clinical routine. In fact, poorly surface-bound probes can affect the diagnostic sensitivity or, in worst cases, lead to false negative results. Here we report on a reliable and easy quality control method for the evaluation of spotted probe properties in a microarray test, based on the Interferometric Reflectance Imaging Sensor (IRIS) system, a high-resolution label free technique able to evaluate the variation of the mass bound to a surface. In particular, we demonstrated that the IRIS analysis of microarray chips immediately after probe immobilization can detect the absence of probes, which recognizably causes a lack of signal when performing a test, with clinical relevance, using fluorescence detection. Moreover, the use of the IRIS technique allowed also to determine the optimal concentration of the probe, that has to be immobilized on the surface, to maximize the target recognition, thus the signal, but to avoid crowding effects. Finally, through this preliminary quality inspection it is possible to highlight differences in the immobilization chemistries. In particular, we have compared NHS ester versus click chemistry reactions using two different surface coatings, demonstrating that, in the diagnostic case used as an example (colorectal cancer) a higher probe density does not reflect a higher binding signal, probably because of a crowding effect.
1.
Kanik, Fulya Ekiz; Celebi, Iris; Sevenler, Derin; Tanriverdi, Kahraman; Ünlü, Nese Lortlar; Freedman, Jane E.; Ünlü, M. Selim
Attomolar sensitivity microRNA detection using real‑time digital microarrays Journal Article
In: Scientific Reports, vol. 2022, no. 12, pp. 16220, 2022.
Abstract | Links | BibTeX | Tags: Oligo
@article{nokey,
title = {Attomolar sensitivity microRNA detection using real‑time digital microarrays},
author = {Fulya Ekiz Kanik and Iris Celebi and Derin Sevenler and Kahraman Tanriverdi and Nese Lortlar Ünlü and Jane E. Freedman and M. Selim Ünlü},
url = {https://axivend.com/wp-content/uploads/2022/09/Attomolar-sensitivity-microRNA-detection.pdf},
doi = {10.1038/s41598-022-19912-z},
year = {2022},
date = {2022-09-27},
urldate = {2022-09-27},
journal = {Scientific Reports},
volume = {2022},
number = {12},
pages = {16220},
abstract = {MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA‑451a (miR‑451) and miRNA‑223‑3p (miR‑223). We demonstrated improvements in sensitivity in comparison to traditional end‑point assays that employ capture on solid phase support, by implementing real‑time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low‑abundance biomarkers in the presence
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.},
keywords = {Oligo},
pubstate = {published},
tppubtype = {article}
}
MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA‑451a (miR‑451) and miRNA‑223‑3p (miR‑223). We demonstrated improvements in sensitivity in comparison to traditional end‑point assays that employ capture on solid phase support, by implementing real‑time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low‑abundance biomarkers in the presence
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.
2.
Kanik, Fulya Ekiz; Celebi, Iris; Sevenler, Derin; Tanriverdi, Kahraman; Ünlü, Nese Lortlar; Freedman, Jane E.; Ünlü, M. Selim
Attomolar sensitivity microRNA detection using real‑time digital microarrays Journal Article
In: Scientific Reports, vol. 2022, iss. 2, no. 12, pp. 16220, 2022.
Abstract | Links | BibTeX | Tags: Oligo
@article{nokey,
title = {Attomolar sensitivity microRNA detection using real‑time digital microarrays},
author = {Fulya Ekiz Kanik and Iris Celebi and Derin Sevenler and Kahraman Tanriverdi and Nese Lortlar Ünlü and Jane E. Freedman and M. Selim Ünlü},
url = {https://mhg912.p3cdn1.secureserver.net/wp-content/uploads/2022/09/Attomolar-sensitivity-microRNA-detection.pdf},
doi = {10.1038/s41598-022-19912-z},
year = {2022},
date = {2022-09-27},
urldate = {2022-09-27},
booktitle = {Imaging Systems and Applications},
journal = {Scientific Reports},
volume = {2022},
number = {12},
issue = {2},
pages = {16220},
publisher = {Optica Publishing Group},
series = {Biophysics At the Nanoscale},
abstract = {MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA‑451a (miR‑451) and miRNA‑223‑3p (miR‑223). We demonstrated improvements in sensitivity in comparison to traditional end‑point assays that employ capture on solid phase support, by implementing real‑time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low‑abundance biomarkers in the presence
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.},
howpublished = {https://chemrxiv.org/engage/chemrxiv/article-details/6565fc1829a13c4d4730f055},
keywords = {Oligo},
pubstate = {published},
tppubtype = {article}
}
MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA‑451a (miR‑451) and miRNA‑223‑3p (miR‑223). We demonstrated improvements in sensitivity in comparison to traditional end‑point assays that employ capture on solid phase support, by implementing real‑time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low‑abundance biomarkers in the presence
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.
of low‑affinity but high‑abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to ∼ 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.
3.
Chiodi, E; Marn, AM; Bakhshpour, M; ans MS Ünlü, N Lortlar Ünlü
The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors Journal Article
In: Polymers, vol. 14, iss. 2, pp. 241, 2022.
Abstract | Links | BibTeX | Tags: Oligo, Protein
@article{nokey,
title = {The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors},
author = {E Chiodi and AM Marn and M Bakhshpour and N Lortlar Ünlü ans MS Ünlü},
editor = {Ick-Soo Kim},
url = {https://www.mdpi.com/2073-4360/14/2/241},
doi = {https://doi.org/10.3390/polym14020241},
year = {2022},
date = {2022-01-07},
urldate = {2022-01-07},
journal = {Polymers},
volume = {14},
issue = {2},
pages = {241},
abstract = {The field of biosensing is in constant evolution, propelled by the need for sensitive, reliable platforms that provide consistent results, especially in the drug development industry, where small molecule characterization is of uttermost relevance. Kinetic characterization of small biochemicals is particularly challenging, and has required sensor developers to find solutions to compensate for the lack of sensitivity of their instruments. In this regard, surface chemistry plays a crucial role. The ligands need to be efficiently immobilized on the sensor surface, and probe distribution, maintenance of their native structure and efficient diffusion of the analyte to the surface need to be optimized. In order to enhance the signal generated by low molecular weight targets, surface plasmon resonance sensors utilize a high density of probes on the surface by employing a thick dextran matrix, resulting in a three-dimensional, multilayer distribution of molecules. Despite increasing the binding signal, this method can generate artifacts, due to the diffusion dependence of surface binding, affecting the accuracy of measured affinity constants. On the other hand, when working with planar surface chemistries, an incredibly high sensitivity is required for low molecular weight analytes, and furthermore the standard method for immobilizing single layers of molecules based on self-assembled monolayers (SAM) of epoxysilane has been demonstrated to promote protein denaturation, thus being far from ideal. Here, we will give a concise overview of the impact of tridimensional immobilization of ligands on label-free biosensors, mostly focusing on the effect of diffusion on binding affinity constants measurements. We will comment on how multilayering of probes is certainly useful in terms of increasing the sensitivity of the sensor, but can cause steric hindrance, mass transport and other diffusion effects. On the other hand, probe monolayers on epoxysilane chemistries do not undergo diffusion effect but rather other artifacts can occur due to probe distortion. Finally, a combination of tridimensional polymeric chemistry and probe monolayer is presented and reviewed, showing advantages and disadvantages over the other two approaches},
keywords = {Oligo, Protein},
pubstate = {published},
tppubtype = {article}
}
The field of biosensing is in constant evolution, propelled by the need for sensitive, reliable platforms that provide consistent results, especially in the drug development industry, where small molecule characterization is of uttermost relevance. Kinetic characterization of small biochemicals is particularly challenging, and has required sensor developers to find solutions to compensate for the lack of sensitivity of their instruments. In this regard, surface chemistry plays a crucial role. The ligands need to be efficiently immobilized on the sensor surface, and probe distribution, maintenance of their native structure and efficient diffusion of the analyte to the surface need to be optimized. In order to enhance the signal generated by low molecular weight targets, surface plasmon resonance sensors utilize a high density of probes on the surface by employing a thick dextran matrix, resulting in a three-dimensional, multilayer distribution of molecules. Despite increasing the binding signal, this method can generate artifacts, due to the diffusion dependence of surface binding, affecting the accuracy of measured affinity constants. On the other hand, when working with planar surface chemistries, an incredibly high sensitivity is required for low molecular weight analytes, and furthermore the standard method for immobilizing single layers of molecules based on self-assembled monolayers (SAM) of epoxysilane has been demonstrated to promote protein denaturation, thus being far from ideal. Here, we will give a concise overview of the impact of tridimensional immobilization of ligands on label-free biosensors, mostly focusing on the effect of diffusion on binding affinity constants measurements. We will comment on how multilayering of probes is certainly useful in terms of increasing the sensitivity of the sensor, but can cause steric hindrance, mass transport and other diffusion effects. On the other hand, probe monolayers on epoxysilane chemistries do not undergo diffusion effect but rather other artifacts can occur due to probe distortion. Finally, a combination of tridimensional polymeric chemistry and probe monolayer is presented and reviewed, showing advantages and disadvantages over the other two approaches
4.
Chiodi, E; Marn, AM; Bakhshpour, M; Ünlü, N Lortlar Ünlü MS
The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors Journal Article
In: Polymers, vol. 14, iss. 2, pp. 241, 2022.
Abstract | Links | BibTeX | Tags: Oligo, Protein
@article{nokeyd,
title = {The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors},
author = {E Chiodi and AM Marn and M Bakhshpour and N Lortlar Ünlü MS Ünlü},
editor = {Ick-Soo Kim},
url = {https://www.mdpi.com/2073-4360/14/2/241},
doi = {https://doi.org/10.3390/polym14020241},
year = {2022},
date = {2022-01-07},
urldate = {2022-01-07},
journal = {Polymers},
volume = {14},
issue = {2},
pages = {241},
abstract = {The field of biosensing is in constant evolution, propelled by the need for sensitive, reliable platforms that provide consistent results, especially in the drug development industry, where small molecule characterization is of uttermost relevance. Kinetic characterization of small biochemicals is particularly challenging, and has required sensor developers to find solutions to compensate for the lack of sensitivity of their instruments. In this regard, surface chemistry plays a crucial role. The ligands need to be efficiently immobilized on the sensor surface, and probe distribution, maintenance of their native structure and efficient diffusion of the analyte to the surface need to be optimized. In order to enhance the signal generated by low molecular weight targets, surface plasmon resonance sensors utilize a high density of probes on the surface by employing a thick dextran matrix, resulting in a three-dimensional, multilayer distribution of molecules. Despite increasing the binding signal, this method can generate artifacts, due to the diffusion dependence of surface binding, affecting the accuracy of measured affinity constants. On the other hand, when working with planar surface chemistries, an incredibly high sensitivity is required for low molecular weight analytes, and furthermore the standard method for immobilizing single layers of molecules based on self-assembled monolayers (SAM) of epoxysilane has been demonstrated to promote protein denaturation, thus being far from ideal. Here, we will give a concise overview of the impact of tridimensional immobilization of ligands on label-free biosensors, mostly focusing on the effect of diffusion on binding affinity constants measurements. We will comment on how multilayering of probes is certainly useful in terms of increasing the sensitivity of the sensor, but can cause steric hindrance, mass transport and other diffusion effects. On the other hand, probe monolayers on epoxysilane chemistries do not undergo diffusion effect but rather other artifacts can occur due to probe distortion. Finally, a combination of tridimensional polymeric chemistry and probe monolayer is presented and reviewed, showing advantages and disadvantages over the other two approaches},
keywords = {Oligo, Protein},
pubstate = {published},
tppubtype = {article}
}
The field of biosensing is in constant evolution, propelled by the need for sensitive, reliable platforms that provide consistent results, especially in the drug development industry, where small molecule characterization is of uttermost relevance. Kinetic characterization of small biochemicals is particularly challenging, and has required sensor developers to find solutions to compensate for the lack of sensitivity of their instruments. In this regard, surface chemistry plays a crucial role. The ligands need to be efficiently immobilized on the sensor surface, and probe distribution, maintenance of their native structure and efficient diffusion of the analyte to the surface need to be optimized. In order to enhance the signal generated by low molecular weight targets, surface plasmon resonance sensors utilize a high density of probes on the surface by employing a thick dextran matrix, resulting in a three-dimensional, multilayer distribution of molecules. Despite increasing the binding signal, this method can generate artifacts, due to the diffusion dependence of surface binding, affecting the accuracy of measured affinity constants. On the other hand, when working with planar surface chemistries, an incredibly high sensitivity is required for low molecular weight analytes, and furthermore the standard method for immobilizing single layers of molecules based on self-assembled monolayers (SAM) of epoxysilane has been demonstrated to promote protein denaturation, thus being far from ideal. Here, we will give a concise overview of the impact of tridimensional immobilization of ligands on label-free biosensors, mostly focusing on the effect of diffusion on binding affinity constants measurements. We will comment on how multilayering of probes is certainly useful in terms of increasing the sensitivity of the sensor, but can cause steric hindrance, mass transport and other diffusion effects. On the other hand, probe monolayers on epoxysilane chemistries do not undergo diffusion effect but rather other artifacts can occur due to probe distortion. Finally, a combination of tridimensional polymeric chemistry and probe monolayer is presented and reviewed, showing advantages and disadvantages over the other two approaches
5.
Chiodi, Elisa; Marn, Allison M; Geib, Matthew T; Ünlü, M Selim
The Role of Surface Chemistry in the Efficacy of Protein and DNA Microarrays for Label-Free Detection: An Overview Journal Article
In: Polymers, vol. 13, iss. 7, pp. 1026, 2021.
Abstract | Links | BibTeX | Tags: Oligo, Protein
@article{nokey,
title = {The Role of Surface Chemistry in the Efficacy of Protein and DNA Microarrays for Label-Free Detection: An Overview},
author = {Elisa Chiodi and Allison M Marn and Matthew T Geib and M Selim Ünlü},
url = {https://www.mdpi.com/2073-4360/13/7/1026/pdf},
doi = {https://doi.org/10.3390/polym13071026},
year = {2021},
date = {2021-01-03},
urldate = {2021-01-03},
journal = {Polymers},
volume = {13},
issue = {7},
pages = {1026},
abstract = {The importance of microarrays in diagnostics and medicine has drastically increased in the last few years. Nevertheless, the efficiency of a microarray-based assay intrinsically depends on the density and functionality of the biorecognition elements immobilized onto each sensor spot. Recently, researchers have put effort into developing new functionalization strategies and technologies which provide efficient immobilization and stability of any sort of molecule. Here, we present an overview of the most widely used methods of surface functionalization of microarray substrates, as well as the most recent advances in the field, and compare their performance in terms of optimal immobilization of the bioreceptor molecules. We focus on label-free microarrays and, in particular, we aim to describe the impact of surface chemistry on two types of microarray-based sensors: microarrays for single particle imaging and for label-free measurements of binding kinetics. Both protein and DNA microarrays are taken into consideration, and the effect of different polymeric coatings on the molecules’ functionalities is critically analyzed.},
keywords = {Oligo, Protein},
pubstate = {published},
tppubtype = {article}
}
The importance of microarrays in diagnostics and medicine has drastically increased in the last few years. Nevertheless, the efficiency of a microarray-based assay intrinsically depends on the density and functionality of the biorecognition elements immobilized onto each sensor spot. Recently, researchers have put effort into developing new functionalization strategies and technologies which provide efficient immobilization and stability of any sort of molecule. Here, we present an overview of the most widely used methods of surface functionalization of microarray substrates, as well as the most recent advances in the field, and compare their performance in terms of optimal immobilization of the bioreceptor molecules. We focus on label-free microarrays and, in particular, we aim to describe the impact of surface chemistry on two types of microarray-based sensors: microarrays for single particle imaging and for label-free measurements of binding kinetics. Both protein and DNA microarrays are taken into consideration, and the effect of different polymeric coatings on the molecules’ functionalities is critically analyzed.
6.
Francesco Damin Elisa Chiodi, Laura Sola
A reliable, label free quality control method for the production of DNA microarrays with clinical applications Journal Article
In: Polymers, vol. 13, iss. 36, pp. 340, 2021.
Abstract | Links | BibTeX | Tags: Oligo
@article{nokey,
title = {A reliable, label free quality control method for the production of DNA microarrays with clinical applications},
author = {Elisa Chiodi, Francesco Damin, Laura Sola, Lucia Ferraro, Dario Brambilla, M Selim Ünlü, Marcella Chiari},
url = {https://www.mdpi.com/2073-4360/13/3/340/pdf},
doi = {https://doi.org/10.3390/polym13030340},
year = {2021},
date = {2021-01-03},
urldate = {2021-01-03},
journal = {Polymers},
volume = {13},
issue = {36},
pages = {340},
abstract = {The manufacture of a very high-quality microarray support is essential for the adoption of this assay format in clinical routine. In fact, poorly surface-bound probes can affect the diagnostic sensitivity or, in worst cases, lead to false negative results. Here we report on a reliable and easy quality control method for the evaluation of spotted probe properties in a microarray test, based on the Interferometric Reflectance Imaging Sensor (IRIS) system, a high-resolution label free technique able to evaluate the variation of the mass bound to a surface. In particular, we demonstrated that the IRIS analysis of microarray chips immediately after probe immobilization can detect the absence of probes, which recognizably causes a lack of signal when performing a test, with clinical relevance, using fluorescence detection. Moreover, the use of the IRIS technique allowed also to determine the optimal concentration of the probe, that has to be immobilized on the surface, to maximize the target recognition, thus the signal, but to avoid crowding effects. Finally, through this preliminary quality inspection it is possible to highlight differences in the immobilization chemistries. In particular, we have compared NHS ester versus click chemistry reactions using two different surface coatings, demonstrating that, in the diagnostic case used as an example (colorectal cancer) a higher probe density does not reflect a higher binding signal, probably because of a crowding effect.},
keywords = {Oligo},
pubstate = {published},
tppubtype = {article}
}
The manufacture of a very high-quality microarray support is essential for the adoption of this assay format in clinical routine. In fact, poorly surface-bound probes can affect the diagnostic sensitivity or, in worst cases, lead to false negative results. Here we report on a reliable and easy quality control method for the evaluation of spotted probe properties in a microarray test, based on the Interferometric Reflectance Imaging Sensor (IRIS) system, a high-resolution label free technique able to evaluate the variation of the mass bound to a surface. In particular, we demonstrated that the IRIS analysis of microarray chips immediately after probe immobilization can detect the absence of probes, which recognizably causes a lack of signal when performing a test, with clinical relevance, using fluorescence detection. Moreover, the use of the IRIS technique allowed also to determine the optimal concentration of the probe, that has to be immobilized on the surface, to maximize the target recognition, thus the signal, but to avoid crowding effects. Finally, through this preliminary quality inspection it is possible to highlight differences in the immobilization chemistries. In particular, we have compared NHS ester versus click chemistry reactions using two different surface coatings, demonstrating that, in the diagnostic case used as an example (colorectal cancer) a higher probe density does not reflect a higher binding signal, probably because of a crowding effect.
7.
Chiodi, Elisa; Marn, Allison M; Geib, Matthew T; Ünlü, M Selim
The Role of Surface Chemistry in the Efficacy of Protein and DNA Microarrays for Label-Free Detection: An Overview Journal Article
In: Polymers, vol. 13, iss. 7, pp. 1026, 2021.
Abstract | Links | BibTeX | Tags: Oligo, Protein
@article{nokeyo,
title = {The Role of Surface Chemistry in the Efficacy of Protein and DNA Microarrays for Label-Free Detection: An Overview},
author = {Elisa Chiodi and Allison M Marn and Matthew T Geib and M Selim Ünlü},
url = {https://www.mdpi.com/2073-4360/13/7/1026/pdf},
doi = {https://doi.org/10.3390/polym13071026},
year = {2021},
date = {2021-01-03},
urldate = {2021-01-03},
journal = {Polymers},
volume = {13},
issue = {7},
pages = {1026},
abstract = {The importance of microarrays in diagnostics and medicine has drastically increased in the last few years. Nevertheless, the efficiency of a microarray-based assay intrinsically depends on the density and functionality of the biorecognition elements immobilized onto each sensor spot. Recently, researchers have put effort into developing new functionalization strategies and technologies which provide efficient immobilization and stability of any sort of molecule. Here, we present an overview of the most widely used methods of surface functionalization of microarray substrates, as well as the most recent advances in the field, and compare their performance in terms of optimal immobilization of the bioreceptor molecules. We focus on label-free microarrays and, in particular, we aim to describe the impact of surface chemistry on two types of microarray-based sensors: microarrays for single particle imaging and for label-free measurements of binding kinetics. Both protein and DNA microarrays are taken into consideration, and the effect of different polymeric coatings on the molecules’ functionalities is critically analyzed.},
keywords = {Oligo, Protein},
pubstate = {published},
tppubtype = {article}
}
The importance of microarrays in diagnostics and medicine has drastically increased in the last few years. Nevertheless, the efficiency of a microarray-based assay intrinsically depends on the density and functionality of the biorecognition elements immobilized onto each sensor spot. Recently, researchers have put effort into developing new functionalization strategies and technologies which provide efficient immobilization and stability of any sort of molecule. Here, we present an overview of the most widely used methods of surface functionalization of microarray substrates, as well as the most recent advances in the field, and compare their performance in terms of optimal immobilization of the bioreceptor molecules. We focus on label-free microarrays and, in particular, we aim to describe the impact of surface chemistry on two types of microarray-based sensors: microarrays for single particle imaging and for label-free measurements of binding kinetics. Both protein and DNA microarrays are taken into consideration, and the effect of different polymeric coatings on the molecules’ functionalities is critically analyzed.
8.
Chiodi, Laura Sola Francesco Damin Elisa
A reliable, label free quality control method for the production of DNA microarrays with clinical applications Journal Article
In: Polymers, vol. 13, iss. 36, pp. 340, 2021.
Abstract | Links | BibTeX | Tags: Oligo
@article{nokeyn,
title = {A reliable, label free quality control method for the production of DNA microarrays with clinical applications},
author = {Laura Sola Francesco Damin Elisa Chiodi},
url = {https://www.mdpi.com/2073-4360/13/3/340/pdf},
doi = {https://doi.org/10.3390/polym13030340},
year = {2021},
date = {2021-01-03},
urldate = {2021-01-03},
journal = {Polymers},
volume = {13},
issue = {36},
pages = {340},
abstract = {The manufacture of a very high-quality microarray support is essential for the adoption of this assay format in clinical routine. In fact, poorly surface-bound probes can affect the diagnostic sensitivity or, in worst cases, lead to false negative results. Here we report on a reliable and easy quality control method for the evaluation of spotted probe properties in a microarray test, based on the Interferometric Reflectance Imaging Sensor (IRIS) system, a high-resolution label free technique able to evaluate the variation of the mass bound to a surface. In particular, we demonstrated that the IRIS analysis of microarray chips immediately after probe immobilization can detect the absence of probes, which recognizably causes a lack of signal when performing a test, with clinical relevance, using fluorescence detection. Moreover, the use of the IRIS technique allowed also to determine the optimal concentration of the probe, that has to be immobilized on the surface, to maximize the target recognition, thus the signal, but to avoid crowding effects. Finally, through this preliminary quality inspection it is possible to highlight differences in the immobilization chemistries. In particular, we have compared NHS ester versus click chemistry reactions using two different surface coatings, demonstrating that, in the diagnostic case used as an example (colorectal cancer) a higher probe density does not reflect a higher binding signal, probably because of a crowding effect.},
keywords = {Oligo},
pubstate = {published},
tppubtype = {article}
}
The manufacture of a very high-quality microarray support is essential for the adoption of this assay format in clinical routine. In fact, poorly surface-bound probes can affect the diagnostic sensitivity or, in worst cases, lead to false negative results. Here we report on a reliable and easy quality control method for the evaluation of spotted probe properties in a microarray test, based on the Interferometric Reflectance Imaging Sensor (IRIS) system, a high-resolution label free technique able to evaluate the variation of the mass bound to a surface. In particular, we demonstrated that the IRIS analysis of microarray chips immediately after probe immobilization can detect the absence of probes, which recognizably causes a lack of signal when performing a test, with clinical relevance, using fluorescence detection. Moreover, the use of the IRIS technique allowed also to determine the optimal concentration of the probe, that has to be immobilized on the surface, to maximize the target recognition, thus the signal, but to avoid crowding effects. Finally, through this preliminary quality inspection it is possible to highlight differences in the immobilization chemistries. In particular, we have compared NHS ester versus click chemistry reactions using two different surface coatings, demonstrating that, in the diagnostic case used as an example (colorectal cancer) a higher probe density does not reflect a higher binding signal, probably because of a crowding effect.
