Learn how ZEISS LSM 980 with Airyscan 2 can help your research

LSM Plus improves literally any confocal experiment with ease, independent of detection mode or emission range. Its linear Wiener filter deconvolution needs next to no interaction while still ensuring a reliable quantitative result. Just as in our time-tested Airyscan super-resolution processing, the underlying optical property information is adapted automatically based on objective lens, refractive index, and emission range.

Apply LSM Plus with no extra effort and benefit from:

• Enhanced signal to noise  at high acquisition speed and low laser power – particularly useful for live cell imaging with low expression levels
• Improved resolution  of spectral data with up to 36 channels in a single scan
• More spatial information  and even greater resolution enhancement for bright samples that allow to close the pinhole of the LSM
• Integrated workflows  to combine the advantages of LSM Plus with Airyscan super-resolution imaging

_{Caption: Drosophila egg chambers stained for F-actin (Phalloidin, magenta) and DE-Cadherin (cyan). Courtesy of T. Jacobs, AG Luschnig, WWU Münster; with T. Zobel, Münster Imaging Network, Germany}

Airyscan 2 is an area detector with 32 circularly arranged detection elements. Each of these acts as a small pinhole, contributing to super-resolution information, while the complete detector area collects more light than the standard confocal setting. This produces much greater light efficiency while capturing enhanced structural information.

Expanding your spectral range into the NIR allows you to use more labels in parallel. Visualize additional structures with more dyes in your multi-color experiments, with the Quasar and NIR detectors efficiently supporting spectral multiplexing experiments. NIR fluorescent labels are less phototoxic for living samples due to the longer wavelength. This allows you to investigate living samples for longer periods of time while limiting the influence of light. Additionally, light of longer wavelength ranges is less scattered by the sample tissue, increasing penetration depth.

For any of the advantages you pursue with NIR labels, the dual-channel NIR detector combines two different detector technologies (extended red GaAsP and GaAs) for optimal sensitivity up to 900 nm.

To separate even highly overlapping signals or to remove autofluorescence, you can take a Lambda Scan using the complete detection range with up to 36 detectors, keeping both illumination and time required to a minimum. Improve spectral imaging along the complete wavelength range, including Online Fingerprinting, with LSM Plus.

The example shows a murine cremaster muscle, multi-color labeled with Hoechst (blue), Prox-1 Alexa488 (green), neutrophil cells Ly-GFP, PECAM1 Dylight549 (yellow), SMA Alexa568 (orange), VEGEF-R3 Alexa594 (red), platelets Dylight 649 (magenta). Acquired with 32-channel GaAsP detector using Online Fingerprinting.

_{Caption: Comparison of the improved SNR before and after LSM Plus processing. Courtesy of Dr. S. Volkery, MPI for Molecular Biomedicine, Münster, Germany}

Multiphoton microscopy (two-photon microscopy, non-linear optical microscopy, NLO) is a preferred method for non-invasive and deep tissue imaging of living or fixed samples, particularly in neuroscience. Multiphoton microscopy capitalizes on the fact that longer wavelengths (600 – 1300 nm) are less absorbed and less scattered by tissues, travelling deeper into the sample while still forming a focal point. The required energy to excite a fluorescent dye is provided not by one photon but by two photons with half the energy each. The probability of two photons to reach the fluorophore at the same time is only sufficient at the focal point. That is why emission light originates from the focal plane and can be efficiently detected, generating an optical section while omitting a pinhole.

_{Caption: Energy diagram of two-photon microscopy}

Combining laser point illumination, linear scanning, and detectors that can capture the signal in photon counting mode make the LSM 980 more than an imaging device:

• Raster Image Correlation Spectroscopy (RICS)
• Fluorescence Correlation Spectroscopy (FCS)
• Fluorescent Cross Correlation Spectroscopy (FCCS)
• Förster Resonance Energy Transfer (FRET)
• Fluorescence Recovery after Photobleaching (FRAP)
• Fluorescence Lifetime Imaging Microscopy (FLIM)

_{Caption:  Fluorescence Correlation Spectroscopy (FCS) principle. Trajectory of a fluorescent particle through the detection volume}