Evaluation of Transfection Agent Complexation with Ferucarbotran to Enhance T Cell Labeling

Leyda Marrero-Morales

Authors:  Leyda S. Marrero-Morales, Angelie Rivera-Rodriguez, Nicole Sarna, Duane Mitchell, Carlos Rinaldi

Faculty Mentor:  Carlos Rinaldi

College:  Herbert Wertheim College of Engineering

Abstract

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.

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Andres Osuna
Andres Osuna (@guest_494)
1 year ago

Very interesting imaging modality! I had never heard of MPI! Great job.

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_4124)
Reply to  Andres Osuna
1 year ago

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!)

Angelie
Angelie (@guest_666)
1 year ago

Great work Leyda!

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_1608)
Reply to  Angelie
1 year ago

Thanks!

Nicole Sarna
Nicole Sarna (@guest_2388)
1 year ago

Great poster and video explanation!

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_6576)
Reply to  Nicole Sarna
1 year ago

Thank you Nicole!

Isabella Campbell
Isabella Campbell (@guest_2506)
1 year ago

Amazing work Leyda! Could you explain why is necessary to track T-cells?

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_4754)
Reply to  Isabella Campbell
1 year ago

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.

Whitney Stoppel
Whitney Stoppel (@guest_3224)
1 year ago

Excellent explanation, Leyda!

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_4262)
Reply to  Whitney Stoppel
1 year ago

Thank you Dr. Stoppel 🙂

Anays Hernandez
Anays Hernandez (@guest_3312)
1 year ago

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.

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_4220)
Reply to  Anays Hernandez
1 year ago

Thanks. We’re very excited about understanding how T cells migrate in biological systems!

Kamila Hernandez
Kamila Hernandez (@guest_3586)
1 year ago

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?

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_6204)
Reply to  Kamila Hernandez
1 year ago

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.

Duncan Ross
Duncan Ross (@guest_3714)
1 year ago

Great work, Leyda! How do you plan on removing the free particles from the supernatant?

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_6558)
Reply to  Duncan Ross
1 year ago

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.

Miguel Perez Blanco
Miguel Perez Blanco (@guest_4490)
1 year ago

Well done, Leyda! Keep up the good work.

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_6642)
Reply to  Miguel Perez Blanco
1 year ago

Thanks!

Lisa Duran
Lisa Duran (@guest_4714)
1 year ago

This is a very interesting topic and great explanation as well. Awesome work Leyda!

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_6678)
Reply to  Lisa Duran
1 year ago

Glad you think so too, magnetic imaging is pretty cool! Thanks.

Ashley Foster
Ashley Foster (@guest_4734)
1 year ago

Your poster and presentation were thorough and phenomenal. Thank you and great job!

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_6604)
Reply to  Ashley Foster
1 year ago

Glad you enjoyed!

Anonymous
Anonymous (@guest_5986)
1 year ago

Hi Leyda!

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?

Great job!

Leyda Marrero-Morales
Leyda Marrero-Morales (@guest_7124)
Reply to  Anonymous
1 year ago

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!

Andres Pulido
Andres Pulido (@guest_6168)
1 year ago

Thorough explanation. Congratulations