Laser Scanning Multiphoton Microscopes
novel scan technology and detection hardware for additional uses includ- ing opto-genetics, are included in the design. Overall, the MPX is engi- neered to be a powerful multimodal imaging tool making LSMM simple and turnkey for all researchers and clinicians, even in a remote non- laboratory environment.
Motivation For a Highly Flexible and Advanced LSMM Space requirements for vari-
Figure 2: Multimodal imaging with the MPX-1040 yields complementary data without moving the sample. The sample shown is a 3μm FFPE section of human skin imaged with A) widefield fluorescence, B) multi-photon: two- photon fluorescence (green, yellow) and SHG (blue); and C) widefield FLIM with a lifetime-color code ranging from 1–4 ns. Scale bar = 500 μm.
of having accessibility to multiple imaging modes. Te flexibil- ity of the MPX provides a long working distance in an upright, inverted, or any oblique imaging angle for addressing various experimental setups. Tese features work to expand functional- ity with limitless degrees of freedom and multimodality.
Adapt the Microscope to Your Experiment and Not Your Experiment to the Microscope Progress in biomedical research is rapidly advancing as
medical applications and benefits grow. Researchers demand an industry-ready, multimodal, and easy-to-use microscope that can manage various tasks. With this concept in mind, the MPX was engineered to be suitable for label-free, whole-slide imaging (WSI) as well as imaging live animals, 2D sections, z-stack 3D and deep tissue samples, immunohistochemistry, immunocy- tochemistry, digital pathology, and more. It also covers a broad range of other applications in materials research. A highly flex- ible front-end is a key design feature that differentiates the MPX from similar microscopes [6]. Te optics and electronics are integrated and alignment-free. Te scan head can be completely rotated around a single axis, optimizing the orientation of the imaging plane with respect to the sample surface, from com- pletely upright to inverted. Auto-adjustable scanning area for macroscopic (whole slide) to microscopic (single-cell) imaging, and even longer working distances necessary for imaging surgi- cally prepared live animals, are possible (Figure 4). Tis allows investigators to adapt the microscope to an experiment and not an experiment to the microscope. Even though the architecture is enclosed, experimental expansion and implementation of future technology, such as a
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ous multiphoton microscopes are diverse. In general, a heavy optical table having dimensions larger than 2 × 3 m is required. Te laser system and beam optics consume most of the space, with the multiphoton lasers oſten requiring a water-cool- ing system, a room temperature stabilization unit, and vibration isolation. For decades commer- cial LSMMs used femtosecond Ti:Sapphire lasers as the main light engine. Tese systems provide excel- lent specifications for studying bio- logical samples. Total average power
is typically a few Watts with pulses < 150 fs at the gain maxi- mum of around 800 nm. Such lasers have a large tuning range of > 300 nm and operate at a high repetition rate of typically 80 MHz (that is, Coherent and Spectra lasers). For experiments involving tissue, only a small fraction of the total power is used before photobleaching or damaging of the sample. As a matter of fact, high resolution of shallow features has been reported as using only 10–100 mW at the sample [7]. In traditional systems Ti-Sapphire lasers consume a con-
siderable portion of the complete system cost, both upfront and over time. Required water cooling also adds complexity and cost. Also, these lasers are non-transportable and difficult to set up outside of the laboratory, for example, in a clinical envi- ronment, and are intimidating for many that are not experts in laser use to operate. Due to the size and sophistication, the laser is not built into the microscope and is usually provided by a manufacturer different than the microscope manufac- turer. Also, further constraining the usability are safety con- siderations due to a Class 4 free-space output, which may be hazardous to the eyes and skin. Terefore, intimate usability and access to the microscope is restricted to well-trained tech- nicians and users. Fiber lasers do reduce the risk of injury and mitigate the concerns mentioned as they provide long-term per- manent alignment and reliability. Te scan head of the MPX allows full 360 degrees rota-
tional freedom and large x, y, z travel. It can be mounted on various motion (manual and motorized) solutions and configu- rations spanning from a simple z-slide to a sophisticated x, y, z motorized linear stage solution with gimbal capability, rotation, and sub-micron precision. Tis design allows many different
www.microscopy-today.com • 2022 May
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