skip to main content

Attention:

The NSF Public Access Repository (NSF-PAR) system and access will be unavailable from 11:00 PM ET on Thursday, May 23 until 2:00 AM ET on Friday, May 24 due to maintenance. We apologize for the inconvenience.


Title: Toward Automated Additive Manufacturing of Living Bio-Tubes Using Ring-Shaped Building Units
Tissue engineering has been largely confined to academic research institutions with limited success in commercial settings. To help address this issue, more work is needed to develop new automated manufacturing processes for tissue-related technologies. In this article, we describe the automation of the funnel-guide, an additive manufacturing method that uses living tissue rings as building units to form bio-tubes. We developed a method based on 96-well plates and a modified off-the-shelf liquid-handling robot to retrieve, perform real-time quality control, and transfer tissue rings to the funnel-guide. Cells seeded into 96-well plates containing specially designed agarose micromolds self-assembled and formed ring-shaped microtissues that could be retrieved using a liquid-handling robot. We characterized the effects of time, cell type, and mold geometry on the morphology of the ring-shaped microtissues to inform optimal use of the building parts. We programmed and modified an off-the-shelf liquid-handling robot to retrieve ring-shaped microtissues from the 96-well plates, and we fabricated a custom illuminated pipette to visualize each ring-shaped microtissue prior to deposit in the funnel guide. Imaging at the liquid-air interface presented challenges that were overcome by controlling lighting conditions and liquid curvature. Based on these images, we incorporated into our workflow a real-time quality control step based on visual inspection and morphological criteria to assess each ring prior to use. We used this system to fabricate bio-tubes of endothelial cells with luminal alignment.  more » « less
Award ID(s):
1827422
NSF-PAR ID:
10157744
Author(s) / Creator(s):
; ; ; ;
Date Published:
Journal Name:
SLAS TECHNOLOGY: Translating Life Sciences Innovation
ISSN:
2472-6303
Page Range / eLocation ID:
247263032092089
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. A critical role of vascular endothelium is as a semi-permeable barrier, dynamically regulating the flux of solutes between blood and the surrounding tissue. Existing platforms that quantify endothelial function in vitro are either significantly throughput limited or overlook physiologically relevant extracellular matrix (ECM) interactions and thus do not recapitulate in vivo function. Leveraging droplet microfluidics, we developed a scalable platform to measure endothelial function in nanoliter-volume, ECM-based microtissues. In this study, we describe our high-throughput method for fabricating endothelial-coated collagen microtissues that incorporate physiologically relevant cell–ECM interactions. We showed that the endothelial cells had characteristic morphology, expressed tight junction proteins, and remodeled the ECM via compaction and deposition of basement membrane. We also measured macromolecular permeability using two optical modalities, and found the cell layers: (1) had permeability values comparable to in vivo measurements and (2) were responsive to physiologically-relevant modulators of endothelial permeability (TNF-α and TGF-β). This is the first demonstration, to the authors’ knowledge, of high-throughput assessment ( n > 150) of endothelial permeability on natural ECM. Additionally, this technology is compatible with standard cell culture equipment ( e.g. multi-well plates) and could be scaled up further to be integrated with automated liquid handling systems and automated imaging platforms. Overall, this platform recapitulates the functions of traditional transwell inserts, but extends application to high-throughput studies and introduces new possibilities for interrogating cell–cell and cell–matrix interactions. 
    more » « less
  2. Abstract

    Strained rings are increasingly important for the design of pharmaceutical candidates, but cross‐coupling of strained rings remains challenging. An attractive, but underdeveloped, approach to diverse functionalized carbocyclic and heterocyclic frameworks containing all‐carbon quaternary centers is the coupling of abundant strained‐ring carboxylic acids with abundant aryl halides. Herein we disclose the development of a nickel‐catalyzed cross‐electrophile approach that couples a variety of strained ringN‐hydroxyphthalimide (NHP) esters, derived from the carboxylic acid in one step, with various aryl and heteroaryl halides under reductive conditions. The chemistry is enabled by the discovery of methods to control NHP ester reactivity, by tuning the solvent or using modified NHP esters, and the discovery thatt‐BuBpyCamCN, an L2X ligand, avoids problematic side reactions. This method can be run in flow and in 96‐well plates.

     
    more » « less
  3. Abstract

    Strained rings are increasingly important for the design of pharmaceutical candidates, but cross‐coupling of strained rings remains challenging. An attractive, but underdeveloped, approach to diverse functionalized carbocyclic and heterocyclic frameworks containing all‐carbon quaternary centers is the coupling of abundant strained‐ring carboxylic acids with abundant aryl halides. Herein we disclose the development of a nickel‐catalyzed cross‐electrophile approach that couples a variety of strained ringN‐hydroxyphthalimide (NHP) esters, derived from the carboxylic acid in one step, with various aryl and heteroaryl halides under reductive conditions. The chemistry is enabled by the discovery of methods to control NHP ester reactivity, by tuning the solvent or using modified NHP esters, and the discovery thatt‐BuBpyCamCN, an L2X ligand, avoids problematic side reactions. This method can be run in flow and in 96‐well plates.

     
    more » « less
  4. Abstract

    High-throughput experimentation (HTE) is an increasingly important tool in reaction discovery. While the hardware for running HTE in the chemical laboratory has evolved significantly in recent years, there remains a need for software solutions to navigate data-rich experiments. Here we have developed phactor™, a software that facilitates the performance and analysis of HTE in a chemical laboratory. phactor™ allows experimentalists to rapidly design arrays of chemical reactions or direct-to-biology experiments in 24, 96, 384, or 1,536 wellplates. Users can access online reagent data, such as a chemical inventory, to virtually populate wells with experiments and produce instructions to perform the reaction array manually, or with the assistance of a liquid handling robot. After completion of the reaction array, analytical results can be uploaded for facile evaluation, and to guide the next series of experiments. All chemical data, metadata, and results are stored in machine-readable formats that are readily translatable to various software. We also demonstrate the use of phactor™ in the discovery of several chemistries, including the identification of a low micromolar inhibitor of the SARS-CoV-2 main protease. Furthermore, phactor™ has been made available for free academic use in 24- and 96-well formats via an online interface.

     
    more » « less
  5. We report simulation studies on the self-assembly of hard-lobed particles (patchy particles where patches appear as lobes around a seed) of different shapes and show that various types of self-assembled morphologies can be achieved by tuning inter-lobe interactions. On self-assembly, the linear building blocks having two lobes around the seed formed rings, the trigonal planar building blocks formed cylindrical hollow tubes and two-dimensional sheets, and the square planar building blocks formed spherical clathrates. The tetrahedral, trigonal bipyramidal, and the octahedral-shaped particles formed compact porous crystalline structures which are constituted by either hexagonal close packed or face centered cubic lattices. The pore size distributions revealed that linear, trigonal planar, and square planar building blocks create highly porous self-assembled structures. Our results suggest that these self-assembled morphologies will potentially find applications in tissue engineering, host-guest chemistry, adsorption, and catalysis. 
    more » « less