Evaluation and optimization of PolyJet 3D-printed materials for cell culture studies.

Use of 3D printing for microfluidics is a rapidly growing area, with applications involving cell culture in these devices also becoming of interest. 3D printing can be used to create custom-designed devices that have complex features and integrate different material types in one device; however, there are fewer studies studying the ability to culture cells on the various substrates that are available. This work describes the effect of PolyJet 3D-printing technology on cell culture of two cell lines, bovine pulmonary artery endothelial cells (BPAECs) and Madin-Darby Canine Kidney (MDCK) cells, on two different types of printed materials (VeroClear or MED610). It was found that untreated devices, when used for studies of 1 day or more, led to unsuccessful culture. A variety of device treatment methodologies were investigated, with the most success coming from the use of sodium hydroxide/sodium metasilicate solution. Devices treated with this cleaning step resulted in culture of BPAECs and MDCK cells that were more similar to what is obtained in traditional culture flasks (in terms of cell morphology, viability, and cell density). LC-MS/MS analysis (via Orbitrap MS) was used to determine potential leachates from untreated devices. Finally, the use of a fiber scaffold in the devices was utilized to further evaluate the treatment methodology and to also demonstrate the ability to perform 3D culture in such devices. This study will be of use for researchers wanting to utilize these or other cell types in PolyJet-based 3D-printed devices.

Direct embedding and versatile placement of electrodes in 3D printed microfluidic-devices.

In this paper, we describe how PolyJet 3D printing technology can be used to fully integrate electrode materials into microfluidic devices during the print process. This approach uses stacked printing (separate printing steps and stage drops) with liquid support to result in devices where electrodes and a capillary fluidic connection are directly integrated and ready to use when printing is complete. A key feature of this approach is the ability to directly incorporate electrode materials into the print process so that the electrode(s) can be placed anywhere in the channel (at any height). We show that this can be done with a single electrode or an electrode array (which led to increases in signal). In both cases, we found that a middle electrode configuration leads to a significant increase in the sensitivity, as opposed to more traditional bottom channel placement. Since the electrode is embedded in the device, in situ platinum black deposition was performed to aid in the detection of nitric oxide. Finally, a generator-collector configuration with an opposed counter electrode was made by placing two working electrodes ∼750 µm apart (in the middle of the channel) and a platinum counter electrode at the bottom of the channel. The utility of this configuration was demonstrated by dual electrode detection of catechol. This 3D printing approach affords robust electrochemical detection schemes with new electrode configurations being possible in a manner that also increases the ease of use and transferability of the 3D printed devices with integrated electrode materials.

Identifying cysteine residues susceptible to oxidation by photoactivatable atomic oxygen precursors using a proteome-wide analysis.

The reactivity profile of atomic oxygen [O(3P)] in the condensed phase has shown a preference for the thiol group of cysteines. In this work, water-soluble O(3P)-precursors were synthesized by adding aromatic burdens and water-soluble sulphonic acid groups to the core structure of dibenzothiophene-S-oxide (DBTO) to study O(3P) reactivity in cell lysates and live cells. The photodeoxygenation of these compounds was investigated using common intermediates, which revealed that an increase in aromatic burdens to the DBTO core structure decreases the total oxidation yield due to competitive photodeoxygenation mechanisms. These derivatives were then tested in cell lysates and live cells to profile changes in cysteine reactivity using the isoTOP-ABPP chemoproteomics platform. The results from this analysis indicated that O(3P) significantly affects cysteine reactivity in the cell. Additionally, O(3P) was found to oxidize cysteines within peptide sequences with leucine and serine conserved at the sites surrounding the oxidized cysteine. O(3P) was also found to least likely oxidize cysteines among membrane proteins.