Exo-Fect is a novel nucleic acid transfer agent that enables the transfection of nucleic acids directly into isolated exosomes. The transfected siRNA, microRNA, mRNA or even plasmid DNA can then be shuttled into target cells via the transfected exosome vesicles. Simply combine isolated exosomes with Exo-Fect and the nucleic acid of your choice to generate exosome delivery vehicles. The protocol takes less than an hour and is highly efficient at placing nucleic acids into exosomes for transport. All Exo-Fect kits come with a Texas-Red labeled positive control, non-targeting siRNA to confirm exosome transfections. The term “FedExosomes” was coined by Dr. Michelle E. Marcus and Dr. Joshua N. Leonard in their article, “FedExosomes: Engineering Therapeutic Biological Nanoparticles that Truly Deliver”published in Pharmaceuticals (Basel), May 2013; 6(5): 659-680.
SIMPLE, FAST PROTOCOL
A Texas-Red end-labeled siRNA (20 pmol) was transfected into exosomes using Exo-Fect. The far left panels show the transfection of the fluorescently labeled siRNA into exosomes that were then immunocaptured on CD63 magnetic beads. The right panels show the transfection of unlabeled siRNA into exosomes that were then immunocaptured on CD63 magnetic beads. The magnetic beads were imaged after overnight capture of transfected HEK293 exosomes on CD63 immunobeads (catalog# EXOFLOW32A-CD63).
The transfected, CD63-captured exosomes were eluted from the beads and then added to HEK293 cells grown in exosome-depleted media (catalog# EXOFBS-50A-1). The transfected exosomes were allowed to dock and interact with the cells for 1-2 days and then the cells were imaged to monitor siRNA cargo delivery.
ExoFect kits work with longer RNAs, like mRNA. We transfected 1 ug of an mRNA encoding RFP into exosomes. These exosomes were then added to HEK293 cells and imaged for RFP protein production after 24 hours. Exo-Fect also enables the transfection of plasmid DNA into exosomes. We used 5 ug of plasmid DNA encoding a GFP gene, transfected exosomes and then added to HEK293 cells. The cells were imaged for GFP protein presence after 48 hours. The image data are shown below.
Small moleculae drugs can also be Exo-Fected into exosomes for delivery to target cells. SBI’s cumate system features a cumate switch promoter that is highly suppressed by its cognate CymR repressor protein abd can switch on with the addition of the small moleucle inducer, cumate. We tested the loading of cumate into exosomes using the Exo-Fect system and then added those exosomes to cells containing a cumate switch GFP reporter construct. The control exosomes were untransfected exosomes
and the cumate-transfected exosomes were treated with Exo-Fect and 450 ug cumate solution. The experimental approach and reporter HEK293 cell data are shown below. The Exo-Fect technology is able to successfully load small molecule drugs into exosomes that are then able to deliver the drug to target cells.
|Exo-Fect Exosome Transfection Kit||10 reactions||EXFT10A-1|
|Exo-Fect Exosome Transfection Kit||20 reactions||EXFT20A-1|
|ExoQuick serum exosome precipitation solution (5 ml)||75 reactions||EXOQ5A-1|
|ExoQuick Plasma prep and Exosome precipitation kit (5 ml ExoQuick plus 500 ul Thrombin at 500U/mL), replaces EXOQ5TD-1 product||75 reactions||EXOQ5TM-1|
|Thrombin Plasma prep for Exosome precipitation (500 ul at 500U/mL), replaces TDEXO-1 product||100 reactions||TMEXO-1|
|ExoQuick serum exosome precipitation solution (20 ml)||300 reactions||EXOQ20A-1|
|ExoQuick-TC for Tissue Culture Media and Urine (10ml)||10 reactions||EXOTC10A-1|
|ExoQuick-TC for Tissue Culture Media and Urine (50ml)||50 reactions||EXOTC50A-1|
|Exosome-depleted FBS Media Supplement – USA Certified||50 ml||EXO-FBS-50A-1|
|Exosome-depleted FBS Media Supplement – USA Certified||250 ml||EXO-FBS-250A-1|
|ExoQuick-LP Lipoprotein Pre-Clear & Exosome Precipitation Kit||5 reactions||EXOLP5A-1|
The ExoQuick serum reaction size is based on using 250 µl serum for exosome isolation and ExoQuick-LP is based on a 100 µl starting serum sample.
The ExoQuick-TC reagent provides enough for either 10 reactions (EXOTC10A-1) or 50 reactions (EXOTC50A-1) from 5 ml Media or Urine for exosome isolation.
Exo-Flow™: Purify Exosomes using Immunoaffinity beads – Selectively capture exosomes on magnetic beads for flow sorting to quantify and purify distinct subpopulations of exosomes.
Exo-FBS exosome-depleted FBS media supplement – Study exosomes from cultured cells and not from fetal bovine serum.
ExoAB and ExoELISA™ Kits – Detect and quantitate exosomes with CD63, CD9, CD81 and Hsp70 antibodies.
SeraMir™ Exosome RNA Amplification Kit – Isolate and convert exosomal RNAs including microRNAs into cDNA for qPCR profiling, Microarrays and NextGen Sequencing applications.
|MCF-7 Human breast cancer, noninvasive cell line: >1×10^6 frozen exosomes||50 ug||EXOP-100A-1|
|MDA-MB-231 Human breast cancer, aggressive/invasive/metastatic cell line: >1×10^6 frozen exosomes||50 ug||EXOP-105A-1|
|HEK293 Human embryonic kidney cell line: >1×10^6 frozen exosomes||50 ug||EXOP-110A-1|
|PC-3 Human prostate cancer cells derived from metastatic cancer cell line: >1×10^6 frozen exosomes||50 ug||EXOP-115A-1|
|A549 Human non-small cell lung cancer cell line: >1×10^6 frozen exosomes||50 ug||EXOP-120A-1|
|H841 Human small cell lung cancer cell line: >1×10^6 frozen exosomes||50 ug||EXOP-125A-1|
|H196 Human small cell lung cancer cell line: >1×10^6 frozen exosomes||50 ug||EXOP-130A-1|
|DMS114 Human small cell lung cancer cell line: >1×10^6 frozen exosomes||50 ug||EXOP-135A-1|
|PCS-500-011 Human pre-adipose derived mesenchymal stem cell: >1×10^6 frozen exosomes||50 ug||EXOP-140A-1|
|PCS-500-012 Human bone marrow-derived mesenchymal stem cell line: >1×10^6 frozen exosomes||50 ug||EXOP-145A-1|
|BC-3 Human B lymphocyte cell line: >1×10^6 frozen exosomes||50 ug||EXOP-150A-1|
|Jurkat Clone E6-1 Human T lymphocyte cell line: >1×10^6 frozen exosomes||50 ug||EXOP-155A-1|
|JAWSII Mouse bone marrow immature dendritic cell line: >1×10^6 frozen dexosomes||50 ug||EXOP-200A-1|
|Human pooled serum: >1×10^6 frozen exosomes||50 ug||EXOP-300A-1|