Servicio de Microscopia Optica Avanzada del Centro Nacional de Biotecnologia

Laser Scanning Confocal Microscopy

Employs a pinhole aperture and focused laser beams to collect serial optical sections, allowing for high-contrast 3D reconstruction.

Wide Field Fluorescence Microscopy

Conventional fluorescence imaging using broadband sources (LED, Metal-Halide, or Hg lamps) for full-field excitation.

Photobleaching Techniques/ FRAP-Fluorescence Recovery After...

Photobleaching techniques used to measure the mobility, diffusion coefficients, and exchange rates of tagged molecules.

Brightfield & other Transmission Microscopy Contrasts

Basic trans-illumination where contrast arises from light absorption by the specimen; typically used for fixed, stained tissues.

FRET- Förster Resonance Energy Transfer

Distance-dependent energy transfer used as a molecular ruler (1–10 nm) to monitor protein interactions and conformational changes.

Total Internal Reflection Fluorescence Microscopy (TIRF)

Utilizes an evanescent field (100–200 nm depth) to selectively excite fluorophores near the coverglass, drastically reducing background noise.

Super Resolution/ Stochastic Optical Reconstruction microsco...

Stochastic activation and localization of single molecules over thousands of frames to reconstruct an image with ~20 nm precision.

FCS- Fluorescence Correlation Spectroscopy

Analyzes intensity fluctuations in a tiny volume to determine molecular concentration, diffusion, and binding kinetics.

FLIM- Fluorescence Lifetime Imaging Microscopy

Measures the fluorescence decay rate, providing information on the molecular microenvironment (pH, Ca2+, quenching) independent of intensity.

Super Resolution/ Stimulated Emission Depletion microscopy (...

Uses a doughnut-shaped depletion laser to quench peripheral fluorescence, effectively narrowing the focal spot to sub-50 nm levels.

Fluorescence Cross-Correlation Spectroscopy (FCCS)

Measures correlated fluctuations of two fluorescent species within a tiny volume to study molecular interactions and complex formation.

Darkfield Microscopy (DFM)

Contrast-enhancing technique that uses oblique illumination to block the central beam, detecting only light scattered by the sample.

Phase Contrast Microscopy (PCM)

Converts phase shifts induced by transparent samples into intensity differences, enhancing contrast without labeling.

Differential Interference Contrast (DIC / Nomarski)

Enhances contrast in transparent samples by converting optical path differences into intensity gradients using polarized light and prisms.

Single-molecule Förster Resonance Energy Transfer (smFRET)

FRET applied at the single-molecule level for real-time monitoring of molecular conformational changes.

Single Particle Tracking (SPT)

High-speed imaging and localization of individual molecules or organelles to study their trajectories and diffusion modes in live cells.

Photoactivation / Photoconversion

Targeted light usage to "switch on" or change the emission color of fluorophores for tracking or pulse-chase experiments.

Image Analysis

Computational extraction of quantitative biological parameters (size, intensity, shape) from raw image data.

Image Restoration

Computational algorithms (e.g., deconvolution) used to reverse optical blurring and enhance image contrast and resolution.

Microscopio Digital Light Sheet

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