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The concentration of nitrate (NO3−) in Narragansett Bay has been shown to undergo considerable temporal and spatial variation. However, the dynamics of this flux has never been monitored on a fine-scale (<100 m, < 1 d) or in real-time. Whole-cell bio-reporters are promising candidates for low cost environmental sensing of bioavailable nutrients. Yet difficulties remain in creating sensors for long term deployment in the marine environment. This paper describes the creation and validation of a low-cost sensor using a self-bioluminescent strain of the cyanobacteria Synechococcus elongatus pcc 7942 for the direct measurement of bioavailable nitrate. Nitrate bioavailability was measured by monitoring light emission from a luxAB based promotor fusion to glnA using a light to frequency sensor and single board microcontroller. Sensor designs are presented in this manuscript with specific focus on storage, cell viability, and compatibility with the marine environment. Sensors were able to consistently assess nitrate standards as low as 1 ppm (16.3 μM). Using a wavelet denoising approach to reduce white noise and hardware noise, nitrate detection of standards as low as 0.037 ppm (0.65 μM) was achieved. Good sensitivity and low cost make these sensors ideal candidates for continuous monitoring of biological nitrates in estuarine systems. 

Extravillous trophoblasts (EVTs) have the potential to provide the entire fetal genome for prenatal testing. Previous studies have demonstrated the presence of EVTs in the cervical canal and the ability to retrieve a small quantity of these cells by cervical sampling. However, these small quantities of trophoblasts are far outnumbered by the population of cervical cells in the sample, making isolation of the trophoblasts challenging. We have developed a method to enrich trophoblast cells from a cervical sample using differential settling of the cells in polystyrene wells. We tested the addition of small quantities of JEG-3 trophoblast cell line cells into clinical samples from standard Pap tests taken at 5 to 20 weeks of gestation to determine the optimal work flow. We observed that a 4 min incubation in the capture wells led to a maximum in JEG-3 cell settling on the surface (71 ± 10% of the initial amount added) with the removal of 91 ± 3% of the cervical cell population, leading to a 700% enrichment in JEG-3 cells. We hypothesized that settling of mucus in the cervical sample affects the separation. Finally, we performed a proof-of-concept study using our work flow and CyteFinder cell picking to verify enrichment and pick individual JEG-3 and trophoblast cells free of cervical cells. Ultimately, this work provides a rapid, facile, and cost-effective method for enriching native trophoblasts from cervical samples for use in subsequent non-invasive prenatal testing using methods including single cell picking.

 

Engineered tissues usually fall short of physiological cell densities and sizes, resulting in limited functional performance. Viability of large tissues is constrained by inadequate diffusion‐driven nutrient exchange. Methods to form large viable tissues are lacking and are constrained by diffusion‐driven nutrient exchange. Here, the use of the Bio‐Pick, Place, and Perfuse (Bio‐P3) is reported, an integrated biofabrication‐bioreactor platform that semiautomatically and rapidly assembles physiologically cell‐dense macrotissues with 100 million cells while being actively perfused. The Bio‐P3 grips, aligns, and stacks prefabricated, scaffold‐free microtissue parts with integrated lumens on a perfusable build‐platform. Parts spontaneously fuse into one continuous macrotissue with perfusable channels. Customizable microtissues are rapidly prepared up to centimeter‐scale with sustained functional performance. Computational models are developed and experimentally validated to elucidate the effects of perfusion rate and tissue geometry on convective nutrient transport in built macrotissues. It is shown that macrotissues constructed from human hepatocellular microtissues maintain geometry and function (albumin and urea secretion) over 5 days. The Bio‐P3 technology fabricates massive solid tissues with high cell numbers and densities to mimic human physiology for preclinical and clinical applications.