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ZEISS Axioscan 7 offers reliably reproducible image quality, no matter whether you repeat your imaging task after a day, a week, a month or even on a different machine.

Introducing Transfer of Intensity Equation (TIE), the new contrast method for contrast generation in transparent samples. Now you can record the interaction of a narrow cone of light with your sample’s structures in three images: one in focus and two out of focus above and below the focal plane. From these three images, the phase information for the central plane is automatically extracted. Continuous acquisition in the z dimension, combined with flash illumination and GPU-based fast image processing, enables very fast delivery of the final contrast images. Present this as either phase contrast or DIC-like relief contrast: it’s your choice.

TIE contrast is an excellent tool that supports your experiments when working with sensitive fluorescent dyes:

• Detect transparent tissues with little to no contrast in regular brightfield mode.
• Speed up the subsequent fluorescence imaging process with very fast flash-based focusing.
• Protect your sensitive dyes from bleaching during focusing by using the lowest light doses.
• Bring your fluorescent labels into context easily by applying additional contrast information.

As part of the definition of factory pre-defined unmixing spectra for the Axioscan 7 for Spatial Biology, all fluorophores of the vendor are used for calibration and quantitative validation. In the example shown, serial tissue sections were stained for cytokeratin in all channels except DAPI. Longer exposure times are used to reveal residual unwanted signals:

• Horizontal structures represent bleed‑through and cross‑talk between channels.
• Vertical structures correspond to tissue autofluorescence.

Even without unmixing, these unwanted signals typically do not exceed 10% of the true staining intensity. With careful titration of marker concentrations, this can be further improved. Alternatively, and with less effort, spectral unmixing significantly reduces both bleed‑through and autofluorescence, providing:

• Cleaner separation of markers across channels
• Higher contrast images, even at higher exposure times
• More reliable data as input for downstream quantification and spatial analysis

Esophagus FFPE tissue sections stained with different chromogenic pathology staining

Bone Marrow FFPE tissue sections stained with different chromogenic pathology staining (first two of the images showing the normal bone marrow, last two Plasmacytoma infiltrated Bone morrow)

Use different forms of polarization illumination to highlight different features. Plane polarized light (PPL) shows the overall crystal color, habit and pleochroism. Crossed polarized light (XPL) at multiple orientations lets you assess the extinction angle and birefringence. Circular polarization allows maximum birefringence to be observed regardless of grain orientation. All channels are aligned using powerful computational algorithms during acquisition, making the resulting data perfect for subsequent segmentation and analysis.