NIEF is a core research facility that serves the academia and industry of Puerto Rico, Latin American, and the Caribbean by providing advanced technology and training in confocal microscopy and electrophysiology.
This technology makes possible the acquisition of fluorescence-based images of cellular structures and molecular events from biological samples and nanomaterials. For example, cellular organelle structures and neuronal action potentials, as well as intracellular calcium oscillations and drug delivery of nanoparticles, can be visualized with high temporal and spatial resolution.
Patch Clamp Electrophysiology
It is a versatile instrument to measure membrane action potentials and ion channel currents as well as the activity of single ion channels from any isolated cells or cellular tissues.
Imaging and Electrophysiology
The coupling of confocal microscopy with patch-clamp electrophysiology enables simultaneous acquisition of fluorescence images and bioelectrical signals from biological cells and tissues.
NIEF’s technologies can be used in a series of applications, for example:
- In vivo ultra-deep and high-speed imaging. For example, biochemical reactions, dynamics of biological processes, and cell interactions in deep areas within living organisms can be visualized at great speed and resolution.
- Simultaneous photostimulation and image acquisition at high-speed can be realized such as photoactivation, photoconversion, FRAP, FLIP, and photo caged-compounds.
- Simultaneous IR excitation imaging. Two different probes can be simultaneous excited with IR light and visualized.
- Digital videos of many important cellular functions such as muscle contraction, cell motility, cell division, and cytokinesis.
- Nanomedicine research. The drug delivery of therapeutic nanomaterials into cells can be visually monitored.
- Ion channel research. Ionic currents from ligand- and voltage-gated ion channels can be recorded with high fidelity as well as optical action potentials from any excitable cells.
- Neuroscience research. The structure and function of neuronal circuits in rats can be simultaneous analyzed for example the action potential firing of multiple neurons in a brain tissue.
Types of Samples
NIEF’s equipment can be used in:
- Primary & recombinant cells (e.g. neurons, cardiomyocytes, yeasts, bacteria, HEK, CHO)
- Cellular tissues (e.g. brain and heart slices, diaphragm, nephron, pancreas)
- Small animal models (e.g. zebrafish, mouse, rat, guinea pig)
- Nanomaterials (e.g. carbon fiber, nanoparticle)
- Epi-Fluorescence Microscopy: Commonly known as widefield microscopy. Emission light excite the whole sample at the same time and digital pictures of the emission light may be capture with a Charge-Couple Device (CCD Camera) or a complementary metal oxide semiconductor (cMOS Camera). This technique maximizes the intensity of sample illumination.
- Confocal Laser Scanning Microscopy: Excitation laser reaches sample over the scanned area. Uses a spatial pinhole to block out-of-focus light that provides the ability to create optical sections of the sample. Axial resolution allows for a 3D reconstruction. Increases signal to noise ratio, reduce phototoxicity compare to Epi-fluorescence but only captures about 5% of all the emitted light.
- Spinning Disc Super Resolution Confocal Microscope: This technique combines the ability of blocking out-of-focus light from confocal with the high sensitivity of the Epi-fluorescence. Excitation is performed just as in Epi-fluorescence but collection of emitted light is made through a pinhole. Ideal for live cell imaging. Spatial resolution is improved to 120nm by deconvolution.
- Total Internal Reflection Fluorescence Microscope: Commonly known as TIRF. This technique restricts excitation of the sample to a thin region, typically near the coverslip (up to 200nm). Improves signal to noise ratio and therefore resolution. Ideal for studies in cell membrane and single molecule.
- Multiphoton Microscope: Also referred as Two-Photon Microscopy. Excitation comes from a tuning laser, wavelengths range from 700nm to 1000nm. Allows for deep tissue penetration and optical sectioning. Pototoxicity is reduce since excitation only occurs on focal plane. Ideal for whole brain samples, thick tissue slides, whole animal and development.