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Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements

Protein microarrays have gained popularity as an attractive tool for various fields, including drug and biomarker development, and diagnostics. Thus, multiplexed binding affinity measurements in microarray format has become crucial. The preparation of microarray-based protein assays relies on precis...

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Autores principales: Celebi, Iris, Geib, Matthew T., Chiodi, Elisa, Lortlar Ünlü, Nese, Ekiz Kanik, Fulya, Ünlü, Selim
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7692379/
https://www.ncbi.nlm.nih.gov/pubmed/33138051
http://dx.doi.org/10.3390/bios10110158
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author Celebi, Iris
Geib, Matthew T.
Chiodi, Elisa
Lortlar Ünlü, Nese
Ekiz Kanik, Fulya
Ünlü, Selim
author_facet Celebi, Iris
Geib, Matthew T.
Chiodi, Elisa
Lortlar Ünlü, Nese
Ekiz Kanik, Fulya
Ünlü, Selim
author_sort Celebi, Iris
collection PubMed
description Protein microarrays have gained popularity as an attractive tool for various fields, including drug and biomarker development, and diagnostics. Thus, multiplexed binding affinity measurements in microarray format has become crucial. The preparation of microarray-based protein assays relies on precise dispensing of probe solutions to achieve efficient immobilization onto an active surface. The prohibitively high cost of equipment and the need for trained personnel to operate high complexity robotic spotters for microarray fabrication are significant detriments for researchers, especially for small laboratories with limited resources. Here, we present a low-cost, instrument-free dispensing technique by which users who are familiar with micropipetting can manually create multiplexed protein assays that show improved capture efficiency and noise level in comparison to that of the robotically spotted assays. In this study, we compare the efficiency of manually and robotically dispensed α-lactalbumin probe spots by analyzing the binding kinetics obtained from the interaction with anti-α-lactalbumin antibodies, using the interferometric reflectance imaging sensor platform. We show that the protein arrays prepared by micropipette manual spotting meet and exceed the performance of those prepared by state-of-the-art robotic spotters. These instrument-free protein assays have a higher binding signal (~4-fold improvement) and a ~3-fold better signal-to-noise ratio (SNR) in binding curves, when compared to the data acquired by averaging 75 robotic spots corresponding to the same effective sensor surface area. We demonstrate the potential of determining antigen-antibody binding coefficients in a 24-multiplexed chip format with less than 5% measurement error.
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spelling pubmed-76923792020-11-28 Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements Celebi, Iris Geib, Matthew T. Chiodi, Elisa Lortlar Ünlü, Nese Ekiz Kanik, Fulya Ünlü, Selim Biosensors (Basel) Article Protein microarrays have gained popularity as an attractive tool for various fields, including drug and biomarker development, and diagnostics. Thus, multiplexed binding affinity measurements in microarray format has become crucial. The preparation of microarray-based protein assays relies on precise dispensing of probe solutions to achieve efficient immobilization onto an active surface. The prohibitively high cost of equipment and the need for trained personnel to operate high complexity robotic spotters for microarray fabrication are significant detriments for researchers, especially for small laboratories with limited resources. Here, we present a low-cost, instrument-free dispensing technique by which users who are familiar with micropipetting can manually create multiplexed protein assays that show improved capture efficiency and noise level in comparison to that of the robotically spotted assays. In this study, we compare the efficiency of manually and robotically dispensed α-lactalbumin probe spots by analyzing the binding kinetics obtained from the interaction with anti-α-lactalbumin antibodies, using the interferometric reflectance imaging sensor platform. We show that the protein arrays prepared by micropipette manual spotting meet and exceed the performance of those prepared by state-of-the-art robotic spotters. These instrument-free protein assays have a higher binding signal (~4-fold improvement) and a ~3-fold better signal-to-noise ratio (SNR) in binding curves, when compared to the data acquired by averaging 75 robotic spots corresponding to the same effective sensor surface area. We demonstrate the potential of determining antigen-antibody binding coefficients in a 24-multiplexed chip format with less than 5% measurement error. MDPI 2020-10-29 /pmc/articles/PMC7692379/ /pubmed/33138051 http://dx.doi.org/10.3390/bios10110158 Text en © 2020 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/).
spellingShingle Article
Celebi, Iris
Geib, Matthew T.
Chiodi, Elisa
Lortlar Ünlü, Nese
Ekiz Kanik, Fulya
Ünlü, Selim
Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements
title Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements
title_full Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements
title_fullStr Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements
title_full_unstemmed Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements
title_short Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements
title_sort instrument-free protein microarray fabrication for accurate affinity measurements
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7692379/
https://www.ncbi.nlm.nih.gov/pubmed/33138051
http://dx.doi.org/10.3390/bios10110158
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