Hello Qiaobing Xu! Good questions!
The antibody coating allows us to specifically direct where these particles accumulate inside the body. Normally if these particles were injected into the body, they would circulate and be filtered out, thus provide no localized benefit.
In this specific case, we are using an antibody coating that adheres to vascular cells. The concept is we can easily substitute the coating to any other antibody to target a multitude of other cell types (Inflamed cells, cancer cells, etc)
A few examples of this concept are listed here:
http://jpet.aspetjournals.org/content/317/3/116...
http://www.sciencedirect.com/science/article/pi...

As for your other question, we have previously published on the uncoated nanoparticle form of PTx. We saw no toxicity up to 2 mg/mL in our endothelial cell line, whereas we DID see toxicity with trolox (the monomer, it is a modification of vitamin E) at lower concentrations. Most interestingly we saw that PTx can inhibit protein carbonyl formation, a symptom of oxidative stress injury, that we did not observe in the native form of trolox.
The full article is listed here: http://onlinelibrary.wiley.com/doi/10.1002/jbm....

Hi Aparna. This is correct, we have a culture of vascular cells, and show that our anti-PECAM-1 antibody coated particles adhere specifically, as opposed to a non specific IgG.
There are a couple of ways to demonstrate in vivo. We can look either directly at particle adhesion, or downstream effects.
To look directly, we can utilize a radioactive label on the antibody (Similar to our cell culture studies), and analyze tissue samples directly to correlate distribution. We have done this in the past in animal studies to determine distribution in the blood brain barrier. To be even more precise, there are methods to separate blood vessels from tissue.
For an indirect method, we can excise tissue of interest, and observe the antioxidant effects through the use of assays such as DCF (A dye that becomes fluorescent in the presence of free radicals, treated areas would be less fluorescent due to the scavenging ability of the PTx), or a Trolox Equivalent Antioxidant Capacity (TEAC), which can measure the antioxidant potential of the tissue.
This has been done in a previous publication using targeting in vivo listed here:
http://www.sciencedirect.com/science/article/pi...

As for an in vitro experiment demonstrating targeting, we have actually inadvertently done this. In utilizing these nanoparticles for a different cell culture application, we discovered we were using a blood brain barrier epithelial cell culture line that did not express PECAM-1. As a result, these nanoparticles (specific to PECAM-1), did not adhere at all to those BBB epithelial cells. In contrast, vascular endothelial cells express PECAM-1, thus can adhere with this targeting method.

Hi Natalia. We have found that the antibody coating increases the nanoparticle size by 15-20 nm. This has been consistent with a few of the nanoparticle systems we have used.
As for your second question, that is an interesting idea and this does tie into the first question.
The physical size of the antibody (by X-ray crystallography) is roughly 5nm x 5nm x 10nm. Because of this size, I don’t think this would be possible to coat ultra-small particles of 5 nm.
In fact a quick calculation I did based on the surface area of a particle that size and the smallest surface area (5nm x 5nm) of an antibody would yield 0.62 antibodies per particle.
However! With that being said, I think it would be possible using just the Fab fragments of an antibody (The portion of the antibody responsible for binding), rather than the entire molecule.
Additionally, it would also be possible to develop an peptide aptamer-type system, which consists of very small sequence of peptides to target the sites we want, which also would be significantly smaller than a whole antibody molecule.

See this link for an easier visual representation of the Fab fragments if you are unfamiliar:
http://en.wikipedia.org/wiki/Fragment_antigen-b...

Hello Hyunjoon! Great question!
While we haven’t looked at stability of THIS specific antioxidant nanoparticle system, we do have a publication coming out on another particle system using the same exact antibody coating.
In this other publication, I had incubated the nanoparticles in whole blood taken from our sample animals. I found that the particles remained stable, along with the antibody coating, for up to 6 hours in blood. After 24 hours, the particles were still stable in terms of suspension, but approximately 50% of the coating had dissociated (And probably replaced with serum proteins).

Hi Qi-Huo, good question! This is an interesting concept in that the term “biocompatibility” always changing.
Currently, iron oxide nanoparticles are already deemed as biodegradable in literature. Indeed this is the case for particles circulating in the blood stream. Through IV injections (Such as in MRI applications), the particles will eventually be deposited in the liver, where Kupffer cells can metabolize the nanoparticles. The digested iron is then kept in the body’s iron stores. There have been reports in literature that even these cells experience a severe rise in oxidative stress, resulting in toxicity.
The issue, however, comes to light when these particles are injected or accumulate in a specific area of tissue (Such as in cancer applications). Endothelial cells have been shown to be able to metabolize iron oxide, but at a slower rate than Kupffer cells, with a significant generation of oxidative stress.
A well written review about iron oxide metabolism is listed here: http://www.sciencedirect.com/science/article/pi...

In order to “increase” biodegradability, many groups have utilized biocompatible coatings or polymer encapsulating delivery systems. While this does not directly influence how fast the iron can be metabolized, some of the coatings do influence how quickly the particles can be cleared, thus potentially lowering long term toxicity.

Thank you Dr. Shin!

Hi Dr. Kwiatkowski! I think you should be able to view the poster.
At the top of the video, there should be two tabs, one that says view video, the other view poster. If you click view poster, it should come up for viewing.