Subscriber access provided by Stockholm University Library
Emerging Trends
Maskless Photochemical Printing of Multiplexed Glycan Microarrays for High-Throughput Binding Studies Daniel Valles, Yasir Naeem, Carlos Carbonell, Alexa Wong, David R. Mootoo, and Adam B Braunschweig ACS Biomater. Sci. Eng., Just Accepted Manuscript • DOI: 10.1021/acsbiomaterials.9b00033 • Publication Date (Web): 01 May 2019 Downloaded from http://pubs.acs.org on May 2, 2019
Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.
is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.
Page 1 of 27 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
ACS Biomaterials Science & Engineering
Maskless Photochemical Printing of Multiplexed Microarrays for High-Throughput Binding Studies
Glycan
Daniel J. Valles,1,2,3 Yasir Naeem2,3 Carlos Carbonell,2,3 Alexa M. Wong,2,3 David R. Mootoo*1,3 and Adam B. Braunschweig*1,2,3,4 1
The PhD program in Chemistry, Graduate Center of the City University of New York, 365 5th
Ave, New York, NY 10016, United States 2
The Advanced Science Research Center at the Graduate Center of the City University of New
York, 85 St. Nicholas Terrace, New York, NY 10031, United States 3Department
4The
of Chemistry, Hunter College, 695 Park Ave, New York, NY 10065, United States
PhD program in Biochemistry, Graduate Center of the City University of New York, 365 5th
Ave, New York, NY 10016, United States Email:
[email protected] 1 ACS Paragon Plus Environment
ACS Biomaterials Science & Engineering 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Page 2 of 27
ABSTRACT Spatially-encoded glycan microarrays promise to rapidly accelerate our understanding of glycan binding in myriad biological processes, which could lead to new therapeutics and previously unknown drug targets. Here, we bring together a digital micromirror device, microfluidic introduction of inks, and advanced surface photochemistry to produce multiplexed glycan microarrays with reduced feature diameters, an increased number of features per array, and with precise control of glycan density at each feature. The versatility of this platform was validated by printing two distinct glycan microarrays where, in the first, different glycans were immobilized to create a multiplexed array, and another where the density of a single glycan was varied systematically to explore the effect of surface presentation on lectin-glycan binding. For lectin binding studies on these miniaturized microarrays, a microfluidic incubation chip was developed that channels multiple different protein solutions over the array. Using the multiplexed array, binding between eight lectin solutions and five different glycosides were determined, such that a single array can interrogate the binding between 40 lectin-glycan combinations. The incubation chip was then used on the array with varied glycan density to study the effects of glycan density on lectin binding. These results show that this novel printer could rapidly advance our understanding of critical unresolved questions in glycobiology, while simultaneously increasing the throughput and reducing the cost of these experiments.
Keywords:
digital
micromirror
device,
glycan
microarrays,
microfluidics,
thiol-ene,
photochemistry, surface science
2 ACS Paragon Plus Environment
Page 3 of 27 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
ACS Biomaterials Science & Engineering
INTRODUCTION Glycan microarrays have accelerated substantially our knowledge of important biological processes, including the immune response1-3, pathogen-cell interactions4-6, and tumor metastasis78.
Spatially encoded glycan microarrays consist of glycans patterned onto a surface with
micrometer-scale feature diameters, and they have received considerable recent attention9-12 because they can test binding interactions in parallel and under identical experimental conditions, while minimizing the quantity of expensive glycan needed to carry out binding experiments. Moreover, studying glycan binding on surfaces is particularly desirable because of the outsized role of cooperative and multivalent interactions on glycan recognition in the dense glycocalyx.13 Therefore, to accurately represent how binding occurs in a biologically relevant environment, glycan-functionalized surfaces that can replicate the density and orientation found in natural biointerfaces, must be prepared. Despite this motivation for studying glycan binding in arrays, the development and adoption of glycan microarray technologies has lagged behind DNA14-16, peptide17-18, protein19-20, and antibody21-22 microarrays because of substantial challenges unique to printing glycans and interrogating their binding. Specifically, glycan samples are difficult to synthesize or isolate from natural sources, so there is a pressing need to reduce feature dimensions and thereby minimize the quantity of glycans needed to prepare the arrays. Glycan arrays are generally prepared by inkjet23 or pin-printing24, leaving features with diameters that are generally >100 µm, which requires considerable volume of glycans or lectins needed for binding studies and restricts the number of spots in an array.25 Additionally, the sensitive functional group tolerance of glycans has tempered the ability to either synthesize them combinatorially on surfaces or control their density and orientation with the surface immobilization chemistries that are commonly used in microarray technologies. Because of the sensitivity of glycan-lectin binding towards these 3 ACS Paragon Plus Environment
ACS Biomaterials Science & Engineering 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Page 4 of 27
different printing parameters, arrays prepared by different approaches can often provide inconsistent results.26 Given these challenges, there is a pressing need for preparing glycan microarrays that, first, can reduce substantially feature area, which minimizes the amount of expensive glycan sample needed for each print and increases the number of features on a substrate, and, second, can vary the glycan density within each feature to investigate how multivalency affects binding. Such a platform will increase the number of experiments that can be carried out with limited glycan samples, considerably reduce costs, and improve experimental throughput, thereby opening opportunities to ask new scientific questions that cannot be addressed with existing methods, and transforming glycoscience research. Efforts to control feature presentation and reduce feature diameter in glycan microarrays have attracted considerable recent attention from both the instrumentation development and the surface chemistry communities. Cutting-edge patterning methods, including microcontact printing27-28 and scanning probe lithography29-32, have been explored as alternatives to inkjet and pin-printing because they can produce feature diameters of
