Evaluation of Transfection Agent Complexation with Ferucarbotran to Enhance T Cell Labeling
Authors: Leyda S. Marrero-Morales, Angelie Rivera-Rodriguez, Nicole Sarna, Duane Mitchell, Carlos Rinaldi
Faculty Mentor: Carlos Rinaldi
College: Herbert Wertheim College of Engineering
Biomedical imaging can be used to track T cells to develop cell-based immunotherapies. Magnetic Particle Imaging (MPI) is a novel technique that allows non-invasive visualization of superparamagnetic iron-oxide (SPIO) tracers. T cell uptake of SPIO is generally poor, so there is a need to optimize labeling. We investigated the effects of ferucarbotran complexation with two transfection agents (TA), poly-L-lysine and protamine sulfate. These cationic molecules interact electrostatically with the SPIO surface, changing the particle charge and potentially increasing SPIO uptake by T cells. Here we characterize SPIO-TA complexes by evaluating magnetic relaxation, hydrodynamic diameter, zeta potential, and MPI performance. We found that SPIO-TA complexes caused particle aggregation in cell media and water. We also show that increasing concentration of TA in water resulted in a more positive zeta potential, while the opposite trend occurred in media for poly-L-lysine. MPI measurements show that increasing TA concentration did not affect MPI signal. T cell viability studies show that protamine sulfate decreases cell viability more than poly-L-lysine. MPI measurements of SPIO-labeled T cells show a signal increase for both SPIO-TA complexes. In the future, we will investigate the effects of the ferucarbotran and poly-L-lysine complex on T cell labeling at different conditions.
Click the video below to view the student's poster pitch.
Very interesting imaging modality! I had never heard of MPI! Great job.
Hi Andres. MPI has a variety of biomedical applications! Some reasons for this:
-its a non-invasive imaging modality
-the signal intensity is proportional to the nanoparticle tracer mass
-tissue depth does not decrease the signal
(These are just some of the reasons!)
Great work Leyda!
Great poster and video explanation!
Thank you Nicole!
Amazing work Leyda! Could you explain why is necessary to track T-cells?
Hi Isabella. We want to understand how T cells migrate in the body as an immune response against malignant cancer cells. We can then help advance the field of cancer immunotherapy by providing a non-invasive and quantitative way of visualizing the immune system’s (T cells) mobilization.
Excellent explanation, Leyda!
Thank you Dr. Stoppel 🙂
Hi Leyda! What an interesting use of transfection agents to affect the charge of the tracer!
Great work, I’m looking forward to see what advancements in this or in other research you make in the future.
Thanks. We’re very excited about understanding how T cells migrate in biological systems!
Hi Leyda, I loved learning about Ferucarbotran and its effects on T cell labeling. Did you encounter any failures throughout the process of your research? If so, how did you get through them?
Glad you enjoyed it Kamila! The biggest impact to this study was the severe aggregation in our samples. This did not help SPIO labeling efficiency because–even though the mechanism for nanoparticle uptake in T cells is not yet well understood–we can rationalize that a large clump of particles (~3000nm for our largest measurement) is not going to be readily internalized in such small cells.
We’ll be moving forward with our Poly-l-Lysine samples that showed less aggregation (Fig 1) to try and disrupt aggregation for better data acquisition.
Great work, Leyda! How do you plan on removing the free particles from the supernatant?
Hi Duncan. Great question! I have plans to vary the ratio of concentrations of transfection agent to ferucarbotran, as well as perform some aggregate disruption techniques. This will let me remove free particles from the supernatant–that is, particles that are not bound to or interacting with the T cells when I go to fix the pelleted samples for MPI measurements.
Well done, Leyda! Keep up the good work.
This is a very interesting topic and great explanation as well. Awesome work Leyda!
Glad you think so too, magnetic imaging is pretty cool! Thanks.
Your poster and presentation were thorough and phenomenal. Thank you and great job!
Glad you enjoyed!
Super interesting topic! Its incredible to see the endless applications of magnetic imaging. I see that your next steps include furthering your research by varying concentrations and mixing conditions. Did you vary the mixing conditions this time around? If so, how did it affect your results?
Hello. The mixing conditions on my poster were the same throughout the results, where we first mixed ferucarbotran (SPIO) and the transfection agents (TA), and then the solvent (water or media) was added. The reasoning behind this was to allow for adequate complexation between the SPIO and the TA, since proteins in the cell media could’ve interacted with the TAs and interrupted SPIO-TA complexation.
We still believe that a protein corona formed around our complexes, contributing to the aggregation of our samples, mainly shown in the DLS measurements in Fig 1.
Thanks for your question!
Thorough explanation. Congratulations