The Bailey Lab shows off their work!



Recently, various members of the Bailey lab showed off their hard work. At the University's Karle Symposium, Colleen picked up an award for her oral presentation, "Microfluidic Platform for Rapidly Incorporating Total Membrane Protein Content from Whole Cell Lysate into Nanodisc Libraries Enables Activity-Based Profiling." Cole also talked his poster about "Cytokine Profiling for the Diagnosis of Chorioamnionitis Using Silicon Photonic Microring Resonator Arrays" up for an award. 

Heather also just got back from the AACC Annual Scientific Meeting & Clinical Lab Expo where she presented her work, "A Machine Learning Approach to Inflammatory Cytokine Profiling Reveals Diagnostic Signatures for Latent Tuberculosis Infection and Reactivation Risk Stratification," and earned the AACC Student Poster Contest, Second Place and Personalized Medicine Division Outstanding Abstract Award! 

Way to go, everyone!

New publication from Vishal and Steve!


Droplet Microfluidics in Thermoplastics: Device Fabrication, Droplet Generation, and Content Manipulation using Integrated Electric and Magnetic Fields


We have developed droplet microfluidic devices in thermoplastics and demonstrated the integration of key functional components that not only facilitate droplet generation, but also include electric field-assisted reagent injection, droplet splitting, and magnetic field-assisted bead extraction. We manufactured devices in poly(methyl methacrylate) and cyclic olefin polymer using a hot-embossing procedure employing silicon masters fabricated via photolithography and deep reactive ion etching techniques. Device characterization showed robust fabrication with uniform feature transfer and good embossing yield. Channel modification with heptadecafluoro-1,1,2,2-tetrahydrodecyltrichlorosilane increased device hydrophobicity, allowing stable generation of 330-pL aqueous droplets using T-junction configuration. Picoinjector and K-channel motifs were also both successfully integrated into the thermoplastic devices, allowing for robust control over electric field-assisted reagent injection, as well as droplet splitting with the K-channel. A magnetic field was also introduced to the K-channel geometry to allow for selective concentration of magnetic beads while decanting waste volume through droplet splitting. To show the ability to link multiple, modular features in a single thermoplastic device, we integrated droplet generation, reagent injection, and magnetic field-assisted droplet splitting on a single device, realizing a magnetic bead washing scheme to selectively exchange the fluid composition around the magnetic particles, analogous to the washing steps in many common biochemical assays. Finally, integrated devices were used to perform a proof-of-concept in-droplet β-galactosidase enzymatic assay combining enzyme-magnetic bead containing droplet generation, resorufin-β-D-galactopyranoside substrate injection, enzyme-substrate reaction, and enzyme-magnetic bead washing. By integrating multiple droplet operations and actuation forces we have demonstrated the potential of thermoplastic droplet microfluidic devices for complex (bio)chemical analysis, and we envision a path toward mass fabrication of droplet microfluidic devices for a range of (bio)chemical applications.

Click here to read the manuscript!

Congrats to Yi on the new publication in Lab on a Chip!


A droplet microfluidic platform for efficient enzymatic chromatin digestion enables robust determination of nucleosome positioning


The first step in chromatin-based epigenetic assays involves the fragmentation of chromatin to facilitate precise genomic localization of the associated DNA. Here, we report the development of a droplet microfluidic device that can rapidly and efficiently digest chromatin into single nucleosomes starting from whole-cell input material offering simplified and automated processing compared to conventional manual preparation. We demonstrate the digestion of chromatin from 2500–125 000 Jurkat cells using micrococcal nuclease for enzymatic processing. We show that the yield of mononucleosomal DNA can be optimized by controlling enzyme concentration and incubation time, with resulting mononucleosome yields exceeding 80%. Bioinformatic analysis of sequenced mononucleosomal DNA (MNase-seq) indicated a high degree of reproducibility and concordance (97–99%) compared with conventionally processed preparations. Our results demonstrate the feasibility of robust and automated nucleosome preparation using a droplet microfluidic platform for nucleosome positioning and downstream epigenomic assays. 

Click here to read the manuscript!

New publication from Richard and Mari in Analytical Methods!


Multiplexed microRNA expression profiling by combined asymmetric PCR and label-free detection using silicon photonic sensor arrays


Analysis methods based upon the quantitative, real-time polymerase chain reaction are extremely powerful; however, they face intrinsic limitations in terms of target multiplexing. In contrast, silicon photonic microring resonators represent a modularly multiplexable sensor array technology that is well-suited to the analysis of targeted biomarker panels. In this manuscript we employ an asymmetric polymerase chain reaction approach to selectively amplify copies of cDNAs generated from targeted miRNAs before multiplexed, label-free quantitation through hybridization to microring resonator arrays pre-functionalized with capture sequences. This method, which shows applicability to low input amounts and a large dynamic range, was demonstrated for the simultaneous detection of eight microRNA targets from twenty primary brain tumor samples with expression profiles in good agreement with literature precedent. Click here to read the manuscript.

Congrats to Mari on the recent publication!


Combining asymmetric PCR-based enzymatic amplification with silicon photonic microring resonators for the detection of lncRNAs from low input human RNA samples


A method for quantifying biologically relevant long-non-coding RNAs by combining nucleic acid amplification via asymmetric polymerase chain reaction (PCR) with label-free PCR product detection using silicon photonic microring resonator arrays is described. This approach eliminates the need for fluorophores, which presents a limit for spectral multiplexing in conventional qPCR methods, and rather offers potential for much higher levels of plexity by spatially arraying capture probes. Here, we demonstrate the potential of this technique to detect two differentially expressed lncRNA transcripts and an internal control mRNA transcript in different commercial human tissue specimens, as well as in a glioblastoma cell line using only nanogram input amounts of total RNA. The obtained results were validated using single-plex RT-qPCR and found to be in good agreement, demonstrating the potential of this technique for lncRNA quantification applications. Click here to read the manuscript.

Congrats to Heather on her recent publication!


A Guide to Quantitative Biomarker Assay
Development using Whispering Gallery
Mode Biosensors


Whispering gallery mode (WGM) sensors are a class of powerful analytical techniques defined by the measurement of changes in the local refractive index at or near the sensor surface. When functionalized with target-specific capture agents, analyte binding can be measured with very low limits of detection. There are many geometric manifestations of WGM sensors, with chip-integrated silicon photonic devices first commercialized because of the robust, wafer-scale device fabrication, facile optical interrogation, and amenability to the creation of multiplexed sensor arrays. Using these arrays, a number of biomolecular targets have been detected in both label-free and label-enhanced assay formats. For example, sub-picomolar detection limits for multiple cytokines were achieved using an enzymatically enhanced sandwich immunoassay that showed high analyte specificity suitable for detection in complex, clinical matrices. This protocol describes a generalizable approach for the development of quantitative, multiplexed immunoassays using silicon photonic microrings as an example WGM platform.

Click here to read the manuscript.

Congrats to Ellen and Sara on the recent publication!

Multiplexed silicon photonic sensor arrays enable facile characterization of coagulation protein binding to Nanodiscs with variable lipid content


Interactions of soluble proteins with the cell membrane are critical within the blood coagulation cascade. Of particular interest are the interactions of γ-carboxyglutamic acid-rich (GLA) domain-containing clotting proteins with lipids. Variability among conventional analytical methods presents challenges for comparing clotting protein-lipid interactions. Most previous studies have investigated only a single clotting protein and lipid composition and have yielded widely different binding constants. Herein, we demonstrate that a combination of lipid bilayer Nanodiscs and a multiplexed silicon photonic analysis technology enables high-throughput probing of many protein-lipid interactions among blood-clotting proteins. This approach allowed direct comparison of the binding constants of prothrombin, factor X, activated factor VII, and activated protein C to seven different binary lipid compositions. In a single experiment, the binding constants of one protein interacting with all lipid compositions were simultaneously determined. A simple surface regeneration then facilitated similar binding measurements for three other coagulation proteins. As expected, our results indicated that all proteins exhibit tighter binding (lower Kd) as the proportion of anionic lipid increases. Interestingly, at high proportions of phosphatidylserine, the Kd values of all four proteins began to converge. We also found that although koff values for all four proteins followed trends similar to those observed for the Kd values, the variation among the proteins was much lower, indicating that much of the variation came from the kinetic binding (kon) of the proteins. These findings indicate that the combination of silicon photonic microring resonator arrays and Nanodiscs enables rapid interrogation of biomolecular binding interactions at model cell membrane interfaces.

Click here to read the manuscript.