Recently, functionalized superparamagnetic iron oxide nanoparticles (SPIONs) have been utilized for protein separation [1] and therapeutic delivery of DNA and drugs. [2] The development of new methods and tools for the targeting and identification of specific biomolecular interactions within living systems is of great interest in the fields of systems biology, target and drug identification, drug delivery, and diagnostics.
Magnetic separation of organelles and proteins from complex whole-cell lysates allows enrichment and elucidation of intracellular interaction partners for a specific immobilized protein or peptide on the surface of SPIONs. This was previously shown for streptavidin-coated magnetic iron oxide beads. [3,4] However, certain binding processes can be energydependent and therefore only occur within the intact cell. Our attempt to target and isolate mitochondria by using multifunctionalized nanoparticles relies on 1) efficient cellular uptake, 2) formation of an affinity complex between the multifunctional particles and the organelle import machinery, and 3) efficient cell disruption and magnetic separation.
Polymer and peptide surface derivatization (Figure 1 a) of aminopropyltriethoxysilane (APS)-coated SPIONs (APS-SPIONs) was accomplished in a fixed-bed reactor by previously described methods. [5] Coupling of the peptide(s) on the magnetic particles was accomplished with a heterobifunctional polyethylene glycol (PEG) cross-linker. Three different Figure 1. Surface derivatization and characterization of mitochondrial targeted APS-SPIONs. a) Two-step surface functionalization of APS-SPIONs. b) Photon correlation spectroscopy number-weighted hydrodynamic size distribution of subsequently functionalized SPIONs. &