Biology subjects

Haack, A. J.

Publications and source records attributed to Haack, A. J..

11 recordsLinked to original sources

bioRxiv2026

homeRNA Just Dropped! Streamlining Remote Blood Collection and RNA Stabilization for Smaller Sample Volumes with homeRNAdrop

At-home sampling can address obstacles to participation in transcriptomics research. We previously developed the homeRNA and homeRNAmax platforms; these custom-designed tubes containing RNA stabilizer interface with a blood tube compatible with upper-arm blood collection devices, allowing for blood collection and RNA stabilization to occur from a participant's home. In this work we introduce homeRNAdrop, an insert for a commercially available blood tube (BD Microtainer) that divides the tube into two compartments: the top compartment holds 200 L blood and the bottom compartment holds ~590 L liquid stabilizer (RNAlater). By combining blood collection and RNA stabilization into the same tube, homeRNAdrop streamlines sample preparation for participants and allows for more compact sample storage for the lab. In a remote pilot study, n=25 participants from across the United States tested the homeRNAdrop kit. The n=23 samples successfully received by the lab show that samples collected using homeRNAdrop are of sufficient RNA quality (all RINs > 6, a common cutoff for analysis) and yield (mean yield=1.44 g) for downstream gene expression analysis. Almost all participants who successfully collected blood using the Tasso Mini device found the homeRNAdrop device easy to use (n=23/24). Overall, this work adds a simplified, user-friendly platform to the growing suite of tools for remote blood RNA stabilization.

bioengineering↗

bioRxiv2026

Curved by Design: Applying Microfluidic Principles for Nonplanar and Planar Suspended Tissue Patterning to the Development of Bladder Models with Tunable Mechanics

Cells in vivo exist in a complex environment where they receive chemical and physical cues from neighboring cells and the extracellular matrix. Suspended, three dimensional (3D) cell culture enables the study of mechanical signals in a controlled in vitro setting where cells can exert forces on the extracellular matrix, and mechanical stimulation can be externally applied. In addition to mechanical cues, tissues in vivo also exhibit spatial heterogeneity and nonplanar topography. To facilitate the development of suspended 3D cell culture models with both spatial and geometric complexity, we previously introduced Suspended Tissue Engineering with Assemblable Microfluidics (STEAM). STEAM is an accessible, modular platform that utilizes fluidic patterning to create multiregional planar and nonplanar suspended cell-embedded 3D tissues. Herein, we further characterize the STEAM dome platform by developing a theoretical model that explains some experimental considerations necessary for successful two-region patterning in a nonplanar construct. We highlight a brief biological application of the planar and nonplanar STEAM platforms by creating simple but physiologically relevant model systems for the bladder, a sphere-like organ with concentric tissue layers and a central lumen that dynamically expands and contracts during filling and voiding. We demonstrate that the suspended configuration of the planar bladder smooth muscle tissue patch induces inherent tension, which can be increased by further straining the tissues; both result in muscle cell alignment along the axis of stretch as shown by a clear peak at 90 degrees in a radial sum analysis of the two dimensional Fast Fourier Transform of images with fluorescent signal from myosin heavy chain 11 immunostaining. Further, we utilize the nonplanar STEAM platform with a human urothelial cell line (HBLAK) and primary bladder smooth muscle cells (HBdSMC) to create a bladder wall model, resulting in a domed, bilayered tissue. STEAM integrates patterning precision, mechanical functionality, and customizability to actualize an accessible and low cost alternative to generate spatially and geometrically complex suspended tissues.

bioengineering↗

bioRxiv2025

To the homeRNAmax: Developing an Improved Blood Self-Collection and Stabilization Platform for Remote Transcriptomic Studies

Shifting human subjects research from research sites to participants homes removes barriers to participation, including transportation and scheduling difficulties. Previously, we developed homeRNA, a kit for immediate stabilization of RNA in self-collected blood using a custom-engineered tube containing RNA stabilizer fluid. The stabilized RNA is extracted and used for downstream gene expression analysis. Here, we introduce homeRNAmax, which improves our original design by interfacing with a commercially available blood collection tube (BD Microtainer), allowing homeRNAmax to be used with any blood collection method that uses this tube and doubling the possible sample volume that can be collected and stabilized compared to the original homeRNA. Through a pilot study (n=19 participants), we show that homeRNAmax (with the Tasso+ blood collection device) produces RNA samples of sufficient quality (mean RIN=7.8) and yield (mean yield=1.93 g) for downstream analysis and can reach participants across the United States, who generally (n=17/19) found the homeRNAmax kit easy to use. A key aspect of the homeRNA and homeRNAmax platforms is a fluidic feature that prevents the RNA stabilizer from spilling, however our previous work had not yet fully characterized the mechanism of this feature. Here, we developed a theoretical model of the spill-resistant feature. In brief, fluid in the tube is suspended due to a balance of pressures; an increase in air volume within the tube reduces the air pressure above the fluid, creating a small vacuum, and preventing fluid leakage. Overall, we show that homeRNAmax is a user-friendly, effective tool for remote blood RNA stabilization.

bioengineering↗

bioRxiv2025

Suspended Tissue Engineering with Assemblable Microfluidics (STEAM)

Suspended tissue culture systems enable cellular responses to mechanical forces critical for tissue development and function. Tissues develop in complex environments containing both mechanical and chemical cues that vary spatiotemporally; thus modeling both of these physiochemistries in vitro through integration of spatial patterning with mechanical manipulation simultaneously is an important aspect in microphysiological tissue modeling which has yet to be achieved. Here, we introduce Suspended Tissue Engineering with Assemblable Microfluidics (STEAM), a modular tissue fabrication platform that allows for spatially heterogeneous suspended tissue architectures. With STEAM, we achieve tissue constructs with multiple regions through the addition of capillary pinning features to control hydrogel precursor flow. STEAM tissues can easily be moved from patterning setup to well-plate to microscope slide, which also enables stacking of separately generated layers. Mechanical manipulation post-fabrication is also possible via static stretching, where cell-embedded 3D tissues can be stretched farther apart to induce strain along an axis. To demonstrate the utility of post fabrication strain ability, we showed that myotube alignment increases when strain is applied to STEAM generated engineered muscle tissue containing mouse myoblasts. Finally, by modifying the channel geometry of the fluidic-based patterning rails, we generate complex nonplanar suspended tissues. STEAM leverages microfluidic principles to generate suspended tissues that integrate patterning precision, mechanical functionality, and experimental versatility, providing a suite of construct combinations for modeling tissue behaviors from the interplay of spatial organization and mechanical forces.

bioengineering↗

bioRxiv2025

From Home to Transcriptome: Comparing the transcriptomic profile of induced immune response via lipopolysaccharide stimulation in homeRNA and venous blood

Remote blood sampling offers multiple advantages over traditional clinic-based blood sampling studies, including greater patient inclusion, more frequent sampling, and broader geographical reach. Combining remote blood sampling with transcriptomic analysis opens potential in translational applications for capturing acute and dynamic immune responses to various exposures. In this study, we establish the feasibility of homeRNA, a capillary blood collection and RNAlater-based stabilization kit, for use in downstream bulk RNA-sequencing applications via capturing a lipopolysaccharide (LPS)-induced inflammatory response. We also compared the baseline gene expression profiles and induced inflammatory response following LPS stimulation between homeRNA-stabilized samples and venous blood stabilized with RNAlater or PAXgene. We found that homeRNA was successfully able to capture an inflammatory response to LPS, specifically targeting various cytokines (e.g., IL6, IL12B, IL1B), chemokines (e.g., CCL3, CXCL10, CCL4), and other transcriptional factors in the toll-like receptor pathway, the primary pathway activated during LPS stimulation. Importantly, we also found that homeRNA captured a LPS-induced inflammatory response comparable to that of venous blood samples stabilized with either RNAlater or PAXgene. Overall, this work demonstrates that the homeRNA platform is compatible with downstream bulk RNA-sequencing analysis and can capture transcriptomic immune responses to a known stimulus which are analogous to results in traditional stabilized venous blood samples.

bioengineering↗

bioRxiv2024

Suspended Tissue Open Microfluidic Patterning (STOMP)

Free-standing tissue structures tethered between pillars are powerful mechanobiology tools for studying cell contraction. To model interfaces ubiquitous in natural tissues and upgrade existing single-region suspended constructs, we developed Suspended Tissue Open Microfluidic Patterning (STOMP), a method to create multi-regional suspended tissues. STOMP uses open microfluidics and capillary pinning to pattern subregions within free-standing tissues, facilitating the study of complex tissue interfaces, such as diseased-healthy boundaries (e.g., fibrotic-healthy) and tissue-type interfaces (e.g., bone-ligament). We observed altered contractile dynamics in fibrotic-healthy engineered heart tissues compared to single-region tissues and differing contractility in bone-ligament enthesis constructs compared to single-tissue periodontal ligament models. STOMP is a versatile platform - surface tension-driven patterning removes material requirements common with other patterning methods (e.g., shear-thinning, photopolymerizable) allowing tissue generation in multiple geometries with native extracellular matrices and advanced 4D materials. STOMP combines the contractile functionality of suspended tissues with precise patterning, enabling dynamic and spatially controlled studies.

bioengineering↗

bioRxiv2024

Your Blood is Out for Delivery: Considerations of Shipping Time and Temperature on Degradation of RNA from Stabilized Whole Blood

Remote research studies are an invaluable tool for reaching populations in geographical regions with limited access to large medical centers or universities. To expand the remote study toolkit, we have previously developed homeRNA, which allows for at-home self-collection and stabilization of blood and demonstrated the feasibility of using homeRNA in high temperature climates. Here, we expand upon this work through a systematic study exploring the effects of high temperature on RNA integrity through in-lab and field experiments. Compared to the frozen controls (overall mean RIN of 8.2, n = 8), samples kept at 37{degrees}C for 2, 4, and 8 days had mean RINs of 7.6, 5.9, and 5.2 (n = 3), respectively, indicating that typical shipping conditions ([~]2 days) yield samples suitable for downstream RNA sequencing. Shorter time intervals (6 hours) resulted in minimal RNA degradation (median RIN of 6.4, n = 3) even at higher temperatures (50{degrees}C) compared to the frozen control (mean RIN of 7.8, n = 3). Additionally, we shipped homeRNA-stabilized blood from a single donor to 14 different states and back during the summer with continuous temperature probes (7.1 median RIN, n = 42). Samples from all locations were analyzed with 3 mRNA-seq to assess differences in gene counts, with the transcriptomic data suggesting that there was no preferential degradation of transcripts as a result of different shipping times, temperatures, and regions. Overall, our data support that homeRNA can be used in elevated temperature conditions, enabling decentralized sample collection for telemedicine, global health, and clinical research.

bioengineering↗

bioRxiv2022

At-home blood collection and stabilization in high temperature climates using homeRNA

Expanding whole blood sample collection for transcriptome analysis beyond traditional phlebotomy clinics will open new frontiers for remote immune research and telemedicine. Determining the stability of RNA in blood samples exposed to high ambient temperatures (>30{degrees}C) is necessary for deploying home-sampling in settings with elevated temperatures (e.g., studying physiological response to natural disasters that occur in warm locations or in the summer). Recently, we have developed homeRNA, a technology that allows for self-blood sampling and RNA stabilization remotely. homeRNA consists of a lancet-based blood collection device, the Tasso-SST which collects up to 0.5 mL of blood from the upper arm, and a custom-built stabilization transfer tube containing RNAlater. In this study, we investigated the robustness of our homeRNA kit in high temperature settings via two small pilot studies in Doha, Qatar (no. participants = 8), and the Western and South Central USA during the summer of 2021, which included a heatwave of unusually high temperatures in some locations (no. participants = 11). Samples collected from participants in Doha were subjected to rapid external temperature fluctuations from being moved to and from air-conditioned areas and extreme heat environments (up to 41{degrees}C external temperature during brief temperature spikes). In the USA pilot study, regions varied in outdoor temperature highs (between 25{degrees}C and 43.4{degrees}C). All samples that returned a RNA integrity number (RIN) value from the Doha, Qatar group had a RIN [≥]7.0, a typical integrity threshold for downstream transcriptomics analysis. RIN values for the Western and South Central USA samples (n=12 samples) ranged from 6.9-8.7 with 9 out of 12 samples reporting RINs [≥]7.0. Overall, our pilot data suggest that homeRNA can be used in some regions that experience elevated temperatures, opening up new geographical frontiers in disseminated transcriptome analysis for applications critical to telemedicine, global health, and expanded clinical research. Further studies, including our ongoing work in Qatar, USA, and Thailand, will continue to test the robustness of homeRNA.

bioengineering↗

bioRxiv2021

homeRNA: A self-sampling kit for the collection of peripheral blood and stabilization of RNA

Gene expression analysis (e.g., targeted gene panels, transcriptomics) from whole blood can elucidate mechanisms of immune function and aid in the discovery of biomarkers. Conventional venipuncture offers only a small snapshot of our broad immune landscape as immune responses may occur outside of the time and location parameters available for conventional venipuncture. A self-operated method that enables flexible sampling of liquid whole blood coupled with immediate stabilization of cellular RNA is instrumental in facilitating capture and preservation of acute or transient immune fluxes. To this end, we developed homeRNA: a kit for self-collection of peripheral blood ([~]0.5 mL) and immediate stabilization of cellular RNA, using the Tasso- SST blood collection device with a specially designed stabilizer tube containing RNAlater. To assess the feasibility of homeRNA for self-collection and stabilization of whole blood RNA, we conducted a pilot study (n = 47 participants) where we sent homeRNA to participants aged 21-69, located across 10 US states (94% successful blood collections, n = 61/65). Among participants who successfully collected blood, 93% reported no or minimal pain/discomfort using the kit (n = 39/42), and 79% reported very easy/somewhat easy stabilization protocol (n = 33/42). Total RNA yield from the stabilized samples ranged between 0.20 {micro}g and 5.99 {micro}g (mean = 1.51 {micro}g), and all but one RNA Integrity Number (RIN) values were above 7.0 (mean = 8.1), indicating limited RNA degradation. Results from this study demonstrate the self-collection and RNA stabilization of whole blood with homeRNA by participants themselves, in their own home.

bioengineering↗

bioRxiv2019

Layer-by-Layer Fabrication of 3D Hydrogel Structures Using Open Microfluidics

Patterning and 3D fabrication techniques have enabled the use of hydrogels for a number of applications including microfluidics, sensors, separations, and tissue engineering in which form fits function. Devices such as reconfigurable microvalves or implantable tissues have been created using lithography or casting techniques. Here, we present a novel open microfluidic patterning method that utilizes surface tension forces to pattern hydrogel layers on top of each other, producing 3D hydrogel structures. We use a patterning device to form a temporary open microfluidic channel on an existing gel layer, allowing the controlled flow of unpolymerized gel in regions defined by the device. Once the gel is polymerized, the patterning device can then be removed, and subsequent layers added to create a multi-layered 3D structure. The use of open-microfluidic and surface tension-based methods to define the shape of each layer enables patterning to be performed with a simple pipette, minimizing dead-volume and shear stress applied on the fluid. Our method is compatible with unmodified (native) biological hydrogels, or other non-biological materials with fluid properties compatible with capillary flow. With our open-microfluidic layer-by-layer fabrication method, we demonstrate the capability to build agarose and type I collagen structures featuring asymmetric designs, multiple components, overhanging features, and cell laden regions.

bioengineering↗

bioRxiv2018

Fast fluorescence lifetime imaging reveals the maturation process of α-synuclein aggregates in ageing Caenorhabditis elegans

The nematode worm Caenorhabditis elegans has emerged as an important model organism to study the molecular mechanisms of protein misfolding diseases associated with amyloid formation because of its small size, ease of genetic manipulation and optical transparency. Obtaining a reliable and quantitative read-out of protein aggregation in this system, however, remains a challenge. To address this problem, we here present a fast time-gated fluorescence lifetime imaging (TG-FLIM) method and show that it provides functional insights into the process of protein aggregation in living animals by enabling the rapid characterisation of different types of aggregates. More specifically, in longitudinal studies of C. elegans models of Parkinsons and Huntingtons diseases, we observed marked differences in the aggregation kinetics and the nature of the protein inclusions formed by -synuclein and polyglutamine. In particular, we found that -synuclein inclusions do not display amyloid-like features until late in the life of the worms, whereas polyglutamine forms amyloid characteristics rapidly in early adulthood. Furthermore, we show that the TG-FLIM method is capable of imaging live and non-anaesthetised worms moving in specially designed agarose micro-chambers. Taken together, our results show that the TG-FLIM method enables high-throughput functional imaging of living C. elegans that can be used to study in vivo mechanisms of aggregation and that has the potential to aid the search for therapeutic modifiers of protein aggregation and toxicity.

biophysics↗