Dr. Rinaldi-Ramos is assigned 1449 sq. ft of laboratory space and has access to extensive shared instrumentation facilities at the University of Florida. These laboratory facilities are described in detail below and support nanoparticle synthesis and characterization, cell culture, and magnetic characterization.
BioNanoMagnetics Lab 1: This facility occupies two contiguous laboratories (total of 1,189 sq. ft) in the Chemical Engineering Building. It is outfitted with two bench-height chemical hoods and one walk-in chemical hood, gas lines (nitrogen, air, vacuum), organic and inorganic synthesis equipment and glassware, two chillers, several shakers, a Millipore Synergy water purifier, two analytical balances, two refrigerated preparative centrifuges (Eppendorf 5430R), two pH meters, one conductivity meter, one QSonica Q700 ultrasonic liquid processor with enclosure, several ultrasonic baths, several heated plates, several heated dry baths, two glassware ovens, two vacuum ovens, several syringe pumps, several mass flow controllers, two 6-foot wide 4°C refrigerators, two -20°C freezers, one -20° flammables freezer, one -80°C freezer, flammable storage cabinets, a Buchi R-215 rotavapor system with chiller and vacuum pump, a LabConco freeze dryer and shell freezer system, and oscilloscopes, several signal generators, magnetic field probes, and power amplifiers. Major instrumentation in the laboratory includes a Brookhaven Instruments ZetaPALS/BI-90Plus dynamic light scattering and zeta potential instrument, a DynoMag AC Susceptometer, a lab-built magnetic particle susceptometer/relaxometer, Perkin-Elmer Frontier FTIR, FormLabs Form 2 SLA 3D printer, a SpectraMax M5 microplate reader, and a nanoScale Biomagnetics D5 Series G3 3.0 kW magnetic hyperthermia instrument with calorimeter, general use, and custom coils. Inside this laboratory there is a separate tissue culture room with biosafety cabinet, two incubators, inverted tissue culture microscope, cell counter, analytical balance, autoclave, and a Keyence BZ-X710 microscope.
BioNanoMagnetics Lab 2: This laboratory occupies 110 sq. ft in the Biomedical Sciences Building, which houses shared instrumentation managed by the J. Crayton Pruitt Family Department of Biomedical Engineering (see below). This laboratory is outfitted with a magnetic nanoparticle heating system consisting of an incubator box, Ambrell EASYHEAT 8310LI 10 kW induction heater, NeOptix Reflex 4-channel fiber optic thermometer with immersion probes, FLIR SC325 thermal camera, and chill water heat exchangers to control coil temperature during magnetic field application. The laboratory has a sink, mass balances, microwave oven (for research use), and temperature-controlled water baths.
BioNanoMagnetics Magnetometry Lab: Dr. Rinaldi is assigned 150 sq. ft of laboratory space in the New Physics building. This laboratory houses a Quantum Design MPMS-3 superconducting quantum interference device (SQUID) magnetometer with AC susceptibility and ultra-low field accessories. This facility takes advantage of the Physics Building’s helium recovery and re-liquefaction system, reducing operating costs.
The following equipment is available in our laboratories and enables a continuum of research on the synthesis, characterization, and application-relevant testing of magnetic nanoparticles.
Controlled Thermal Decomposition Synthesis Setup. We have assembled a suite of instruments and components to enable fully controlled inorganic nanoparticle synthesis. This includes pressure/vacuum control (500-760 Torr) through a condenser/fraction collection system using a DigiVac Model 450 proportional vacuum regulator that senses pressure at the reactor, temperature control up to 370°C, controlled addition of carrier and reactive gases using Bronkhorst/Alicat digital mass flow controllers, controlled addition of liquid precursors using dual KD Scientific (KDS220) syringe pumps, and the necessary glassware and components to enable addition or reactants and removal of sample aliquots under controlled conditions. Reaction temperature and pressure, and precursor addition rates are controlled and recorded through a computer interface.
Flash NanoPrecipitation Setup. We have assembled a suite of instruments and components to enable the rapid formulation of nanocarriers consisting of hydrophobic cores containing magnetic nanoparticles, drugs/fluorophores, and other co-core excipients coated with amphiphilic copolymers that are kinetically trapped on the nanocarrier surface and provide a dense brush of stabilizing hydrophilic polymer. We currently have two types of FNP confined impinging jet (CIJ) mixers: (i) standard two jet mixers and (ii) custom designed eight-jet mixers. Streams containing the nanoparticles, drugs/fluorophores, co-core excipients, and amphiphilic copolymers in good solvents are rapidly mixed in these devices with an antisolvent using digital infusion syringe pumps to yield nanocarriers with sizes and contents that can be controlled through process parameters (e.g., stream flow rates and compositions).
Brookhaven Instruments BI-90Plus/Zeta PALS Dynamic Light Scattering (DLS) and Zeta Potential Analyzer. This is a fixed-angle DLS instrument with integrated zeta-potential measurement capacity. Hydrodynamic radii as small as 4 nm can be determined, depending on sample preparation and scattering properties of the colloidal particles (our particles are solid ferrites, and hence very strong scatterers). This instrument can measure zeta potential in aqueous solutions with high ionic content and in organic solvents, making it suitable for nanoparticle characterization in these environments.
Buchi R-215 Rotavapor System. This rotary evaporator has fully integrated temperature/pressure controller, vacuum pump, and chiller, allowing for programmed runs and automated solvent recovery.
Molecular Devices SpectraMax M5 Microplate Reader. This is a temperature controlled (4°C above ambient to 50°C) plate reader that can measure fluorescence, luminescence, absorbance, time-resolved fluorescence (TRF) and fluorescence polarization (TRP). It has dual monochromators that allows users to determine optimal excitation and emission settings for both fluorescence and absorbance. This instrument can handle 6-, 96-, and 384- well plates and allows for end point, kinetic, spectral scanning, and well-area scanning modes.
Keyence Epi-Fluorescence microscope BZ-X710. This compact sized microscope supports fluorescence, brightfield, and phase contrast imaging, with full plate imaging, multi-point image capture, advanced image stitching, and optical sectioning capabilities. This microscope is equipped with 4x and 20x long working distance objectives and 20x, 40x, and 100x short working distance objectives. Filter sets are available for DAPI, GFP, Texas Red, Cy5, and Cy 7. The microscope has image acquisition and processing software that enables automating capture of high-resolution images and process data for cell counting.
PerkinElmer Frontier FTIR. This is a flexible, easy to use FTIR with Ge coated KBr optics, scant range of 8,300 to 350 cm-1 with 0.4 cm-1 resolution, and 10,000/1 peak-to-peak noise. It is equipped with a diamond/ZnSe attenuated total reflectance measurement apparatus with a top plate and pressure arm.
FormLabs Form 2 SLA 3D printer. This is a benchtop stereolithography 3D printer capable of printing using commercial clear and colored resins and biocompatible/autoclaveable resins. It has a build volume of 145 mm x 145 mm x 175 mm, laser spot size of 140 μm, and adjustable layer height of 25-100 μm. Also available are automater rinsing (Form Wash) and photocuring (Form Cure) stations.
Quantum Design SQUID-VSM Magnetometer. This is the state-of-the-art SQUID-based magnetometer by Quantum Design. It is an extremely sensitive (~10-8 emu) magnetometer capable of measuring sample magnetization up to 300 emu with controllable temperature range from 1.9 – 400 K. Cooling and heating rates of up to 30 K/min are possible above 10 K. This fully programmable instrument is equipped with an AC susceptometer (±10 Oe applied field amplitude and 0.1 – 1,000 Hz applied field frequency range) and ultra-low field option. The instrument is housed in the Physics building, benefitting from the Helium recovery and re-liquefaction facilities available there.
DynoMag AC Susceptometer. This is a portable AC susceptometer with frequency range from 1 Hz to 500 kHz with magnetic moment resolution of 10-11 Am2 at 1 kHz. AC susceptibility measurements allow determination of magnetic nanoparticle relaxation mechanism and characteristic times.
MPI Susceptometer/Relaxometer. This is a custom-built instrument capable of applying alternating magnetic fields at selectable frequencies from 3 kHz to 25 kHz and amplitudes up to 50 mT peak-to-peak, while also applying constant and ramped bias fields in the range of ±100 mT to a sample up to 500 μl. The sample magnetization is measured using a balanced gradiometer coil and a 10 MHz sampling rate data acquisition card and processed through in-house software to obtain derivative of the magnetization as a function of time, harmonic spectra, and point spread function information useful in characterizing the magnetic particle imaging properties of magnetic nanoparticles.
10 kW Induction Heating System. This is a combination of an induction heater high frequency magnetic field source (Ambrell EASYHEAT 8130LI 10 kW unit with custom coils), an incubator enclosure to maintain sample temperature and environmental CO2 and humidity mimicking an incubator, fiberoptic temperature probes (NeOptix Reflex), thermal imaging camera (FLIR SC325), and fiberoptic fluorescence probe (Ocean Optics USB4000) to monitor release of fluorophores under alternating magnetic fields. Depending on coil geometry magnetic fields of 300 kHz and up to 80 kA/m can be generated. We use this suite of instruments to carry out experiments in which nanoparticles are exposed to alternating magnetic fields. We are capable of monitoring temperature simultaneously during application of an AMF. A magnet rig is available to generate a field free line selection magnetic field gradient with tunable gradient strength. This device permits spatial selection of magnetic nanoparticle heating with sub-centimeter resolution.
3 kW Induction Heating System. This is a nanoScale Biomagnetics D5 series G3 alternating magnetic field driver for advanced magnetic hyperthermia and nanoparticle heating research. It is equipped with a calorimeter coil for accurate determination of nanoparticle specific absorption rate (SAR), a Double-H Helmholtz coil with heated pad, and a 56 mm internal diameter general purpose coil. The driver is computer controlled and can run automated experimental sequences. Temperature is monitored using fiberoptic temperature probes and is monitored by the driver software and used in performing automated experiments. Alternating magnetic fields (AMF) can be generated in a range of amplitudes at 14 selected frequencies for each coil.
Anton Paar MCR301 Rheometer. This rheometer is capable of a variety of rheological tests in both oscillatory and rotational modes for testing viscoelastic properties of biomaterials. It is possible to control and measure temperature throughout the experiment. In addition, the modular format enables experimental customization.
Shared Major Instrumentation in the UF J. Crayton Pruitt Family Department of Biomedical Engineering
The J. Crayton Pruitt Family Department of Biomedical Engineering manages an extensive suite of shared instrumentation to facilitate research in biomedical engineering. The following instruments are some of the ones most relevant to our research.
Magnetic Insight MomentumTM Imager. This is a fully integrated and self-shielded pre-clinical magnetic particle imaging (MPI) system. It utilizes a high-field gradient main magnet geometry and proprietary x-space reconstruction process to produce high sensitivity and resolution MPI images. Image acquisition is fully automated and computer controlled. It has a 40 mm imaging bore. It applies a 45 kHz, 16 mT excitation field and selection field gradient of 3 to 6.1 T/m to obtain images with < 1 mm resolution using commercial nanoparticle tracers (VivoTraxTM). It has a sensitivity of 10 ng Fe using commercial nanoparticle tracers (VivoTraxTM). Projection scans can be obtained in < 10s/image. Tomographic scans can be obtained in 10-25 minutes per image. It is also equipped with a beta version of the RelaxTM module for MPI tracer characterization.
Perkin Elmer IVIS Spectrum CT In Vivo Imaging System. This is an in vivo imaging system to facilitate non-invasive longitudinal monitoring of disease progression, cell trafficking and gene expression patterns. It also offers single-view 3D tomography for both fluorescent and bioluminescent reporters that can be analyzed in an anatomical context. It offers High resolution (to 20 microns) with 3.9 cm field of view, with twenty-eight high efficiency filters spanning 430 – 850 nm. This instrument can do 3D diffuse tomographic reconstruction for both fluorescence and bioluminescence.
Zeiss LSM 880 Confocal Microscope with airy scan. This device provides high resolution, sensitivity and speed that increases productivity when performing quantitative imaging. Its unique Airyscan detector enables superresolution imaging (120 nm resolution in XY, 400 nm in Z). It includes laser lines405, 458, 488, 514, 561, 633 nm.
BD FACS Celesta Flow Cytometer. This is a multicolor flow cytometer that comes with 3 lasers (blue, red and violet) and can be employed in cell counting and biomarker detection.
Guava 8HT Flow Cytometer. This is a multicolor flow cytometer that comes with 2 lasers (blue and red ) and can be employed in cell counting and biomarker detection.
Other Relevant Shared Instrumentation Facilities
The Herbert Wertheim College of Engineering’s Research Service Centers (RSC) support and enhance the research, education, and public service missions of the University of Florida by providing access to characterization and process instrumentation. Expert staff provides the assistance and guidance necessary so that students, faculty, and industry get the most effective and appropriate use of the center’s facilities. A wide range of instrumentation is available to enable 3D printing, micro- and nano-fabrication, photolithography, scanning and transmission electron microscopy with microanalysis, metrology, particle analysis and characterization, spectroscopy (UV, FTIR, Raman, XPS), and x-ray analysis. Access to an aberration corrected S/TEM with high-angle annular dark field scanning transmission electron microscopy (HAADF-STEM) and elemental mapping capabilities available at Florida State University is provided at internal user rates through the National High Magnetic Field Laboratory and coordinated by HWCOE RSC.
The ICBR Electron Microscopy core supports researchers in visualizing structures of microscopic samples by carrying out imaging projects and user training to facilitate solving research questions and promote acquisition of grants. Major service categories include: transmission and scanning electron microscopy, confocal laser scanning and epiflourescence microscopy, preparation of samples for microscopic analyses, and training users to operate microscopes and ancillary equipment.
The ICBR Proteomics and Mass Spectroscopy core provides services related to protein fractionation, qualitative and quantitative mass spectrometry analysis, de novo protein sequencing and protein database searching. Protein sequencing and identification are routing procedures of the core. The core has a suite of state-of-the-art mass spectrometers and software available for detailed characterization of proteins and peptides, including post-translational modification analysis and accurate molecular weight determination. To ensure success and maximize productivity, the core offers education, consultation, data processing and reporting, and support for grant applications.
The ICBR Cytometry core provides a variety of tools and expertise for cellular measurements. The laboratory has numerous flow cytometers, from entry-level devices to high-end 5-laser, 16-parameter instruments to analyze and sort cells. The laboratory also offers laser scanning confocal microscopy instrumentation. Instruments can be operated by core staff or trained project personnel.
The College of Medicine Electron Microscopy Core Facility supports researchers by providing consultation, service, teaching, and access to equipment for ultrastructural studies of tissue samples using electron microscopy.