Since we last discovered our polyurethane acrylate (PUA) has a shelf life (described here) and received a new solution of PUA, we have been trying to replicate the results that we had before. Unfortunately, we have realized that this is much easier said than done. What we have found is the specific dilution of PUA…
Introduction While we have optimized coculture models for commercial brain endothelial cells and pericytes, we needed to do the same for iPSC-derived cocultures. In this post, I show the optimization process using IMR90-4-derived EECM-BMEC-like cells [1] and brain pericyte-like cells (BPLC) [2]. Methods The top well chip surface was coated with collagen iv (Sigma C5533), bovine fibronectin…
Introduction The µSiM BBB model is being developed at UR with the goal to understand the impact of sepsis on the brain. Here, we use an EECM-BMEC-like cell (iSPC-derived brain endothelial cells) [1] monoculture model and an EECM-BMEC-like cell and brain pericyte-like cell (BPLC; iSPC-derived brain pericytes) [2] coculture model to optimize the concentration of…
Introduction High throughput imaging techniques in conjunction with in vitro micro-physiological systems (MPS) allow for comprehensive exploration of physiologically relevant phenomena in a living 3D mimetic of a tissue microenvironment. Prior work in this lab has readily demonstrated that vascular tissue mimetics (the µSiM) facilitate real time phase-microscopy visualization of immune cell trafficking in response…
Introduction The blood-brain barrier (BBB) plays an important role in maintaining brain chemistry by restricting the permeability of small molecules and cells. Inflammatory diseases may disrupt the BBB and promote the development of neurological disorders [1]. One potential mechanism of BBB disruption is the degradation of the extracellular matrix proteins that make up the endothelial…
The fabrication of nano pocket membrane to capture and release the EV faced challenges during the etching step. There was a suggestion after the last NRG post to use RIE-2000 located in the UR. This post shows the update on the methodology of creating the nano pocket membranes and the preliminary results after using the…
Summary We look at nanoparticle counting and colocalization after an ALine device membrane capture and compare IMARIS and ImageJ methods with an example. Procedures for particle counting and colocalization for IMARIS are discussed and a walkthrough is provided An example of the observed difference between ImageJ and IMARIS on the observed counts showing a near…
The battle for soft membrane coatings wages on as we continue to optimize polyurethane acrylate (PUA) coatings for our osteomyelitis project, whose background is described here and here and initial progress is described here. Since our last update, we have honed in on our most promising recipe and tried to fine tune its settings in…
The first DNA extraction was done over 150 years ago in 18691. Since then, the process has been refined and expanded to include multiple physical and or chemical methods. For the waste water epidemiology project (described here), we will be focusing on DNA extraction via silica adsorption without the use of chaotropic agents. The mechanism…
Introduction In the Glading Lab we study Cerebral Cavernous Malformations (CCMs). In CCMs, brain capillaries develop enlarged lumens and a disrupted blood-brain barrier. The presence of a mutation in KRIT1 causes loss of KRIT1 protein function in the endothelial cells. KRIT 1 plays a role in the integrity of endothelial junctions, therefore the lack of…