The purpose of this experiment was to quantify the effectiveness of the kit membrane at separating free antibody and qdots from qdot-bound antibodies. The manual advertises an approximate 80% yield for this step and no purity reading. It was originally planned to directly measure the purity based on the amount of free antibody in the…

Alright, so this is kind of a big update and I will probably split it into a couple of parts. Basically, in a search for a way of confirming that the particles we are seeing on the membranes, I have tried several techniques with varying degrees of success. The initial experiment involved me conjugating an…

Since I am posting data that is acquired using Auger electron spectroscopy, I thought that it might be a good idea to write a post about the theory of the instrument, to better understand why this data is useful and relevant. The Auger effect is a phenomenon in which an atom that is excited by…

Summary Statement: We have doubled our MgF2 yields (compared to nanoporous MgF2) by moving to thicker microporous membranes. While the increase in strength is more modest than initially expected, we have plenty of substrates for use in cell culture. Statistics: Window [mm] Average Std Dev Count 0.1 21.02 8.10 14 0.3 8.18 5.62 17 0.5 4.33…

In attempting to generate control systems for our exosome work as well as to potentially develop a system that would help some collaborators at the University of Illinois, I have been working with the ultracentrifuge again to hopefully isolate a pure fraction of exosomes, devoid of any protein components. This would allow us to start…

Used Thermofisher SiteClick qdot antibody labeling kit 625.  Began with 4.0mg/mL IgG and conjugated it with an unknown amount of qDots, methods provided by the SiteClick kit. The final stage in the manual [1] is an optional cleaning of the qDots to remove any unconjugated and free-floating qDots from the mixture using the nanosep omega…

I am trying to understand the development of endothelial barrier properties as the cells remodel under the influence of physiological shear stresses. I culture HUVECs inside my device on 100 micron thin type I collagen gel. I coat the gel with fibronectin on top to provide adhesion for the cells. It helps them to stay…

First off, this is the protocol for the staining and analyzing the data is shown below and was used for all of the YAP experiments: For the substrates, we wanted to use 0.5μm HP, 0.5μm LP, 3μm HP, TCP, and nonporous SiO2.  Cyto-Vu devices were made using the aforementioned substrates and 300 ADSCs were seeded per membrane.  The…

Goal: study HUVEC migration on different porous substrates. HP = high porosity 0.5 μm SiO2 membrane LP = low porosity 0.5 μm SiO2 membrane NP = non-porous SiO2 membrane TCP = tissue culture polystyrene (~3 GPa [ref 1, 2]) P184 = sylgard 184 (a commonly used PDMS; Young’s modulus = 1.74-1.84 MPa [ref 3]) P527 = sylgard 527…