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Perfect Focus System

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Nikon's premier dynamic focusing system just got better.

Vasculature of a zebrafish embryo (0:16)

Inverted research microscopes -- and their peripheral equipment -- have greatly contributed to the development and advancement of life science research throughout the years. Nikon’s Perfect Focus system has been particularly revolutionary in this area, directly addressing the challenge of focus drift, which reduces the reliability of acquired data, specifically in long-term time-lapse observations using higher magnifications and resolutions. The new Perfect Focus Systems continue to combat this issue by keeping the focus precise and making corrections on a millisecond time-scale thanks to Nikon’s proprietary optical offset method.

The new models of the third generation Perfect focus System are the TI-ND6-PFS-S Perfect Focus Unit with Motorized Nosepiece for UV-visible wavelength imaging and the TI-ND6-PFS-S Perfect Focus Unit with Motorized Nosepiece for multiphoton imaging.

The next generation PFS3 offers significant enhancements over previous models:

• Maintain focus at deeper depths • Compatible with plastic dishes and well plates
• Wider useable excitation wavelength range • Streamlined Design

Key Features

Exclusive Nikon Perfect Focus Technology

Perfect Focus Technology

For use with Nikon's Eclipse Ti-E Inverted Microscope, the Perfect Focus System (PFS) provides real time focus correction to overcome microscope focus drift caused by thermal and mechanical effects for dramatically increased quality of long-term time-lapse imaging data in live cells.

By combining this highly sensitive feedback system with the accurate Z-axis control of the Ti-E, continuous real time focus correction is easily achieved.

The PFS3 contains proprietary optical offset technology, which allows the researcher to focus at the desired height above the coverslip while simultaneously tracking the focus of the coverslip interface. This technology allows continuous real time focus correction, making the system orders of magnitude faster, and considerably less prone to error than other manufacturer's systems that must continuously shift between the coverslip interface and the focal plane of interest.


Focus Detection with Infrared Light

Focus Detection with Infrared Light

The 870nm wavelength, which does not affect fluorescence observation, is used for focus detection.

PFS3 uses an LED in the infrared range and an internal linear CMOS detector, to detect the focal point, so it does not intrude on wavelengths used for observation. This means you can carry out observation and focus maintenance at the same time, with no influence at all on captured images.

Since the wavelength of the LED does not affect the image quality, high-contrast visualization of single fluorescent molecules—normally only possible with high-S/N ratio microscopy—is possible.


More Powerful and Versatile than Ever Before

TiE-PFS3

Introduced in 2005, Nikon's Perfect Focus System set a new standard for live cell imaging. The new third generation system, available in two models, offers a host of enhancements:

Maintaining Focus at Deeper Depths- PFS3 can correct for axial drifts and maintain focus at larger distances from the objective and at deeper depths in the imaging specimen. This new capability is not only relevant for developmental biology, but other fields of research as experimental trends move toward studying the dynamics of cells in their natural environment such as thick tissues or whole animals.

Increased flexibility in imaging wavelengths for multiphoton microscopy and optical trapping- The multiphoton model of the new PFS now offers compatibility with a larger range of imaging wavelengths, increasing the flexibility of fluorophores and lasers that can be used for multiphoton imaging and other near-infrared applications such as optical trapping/tweezers. The new PFS3 Multiphoton model can correct for focus drifts even when imaging with wavelengths ranging from 880-1300nm.

Streamlined Design- The newly designed PFS systems enable easier access to objective lenses and their correction collars. In addition, all PFS functions are now controlled through the PC or external controller, eliminating the need to open and close environmental chambers which can lead to disturbances in the imaging temperature.

Suitable for Use with Plastic Dishes and Plastic Well Plates- In addition to glass bottom dishes, the plastic dish -- which is less expensive and suitable for cell culture -- can be used with the PFS3. Use of plastic dishes allows researchers to reduce running costs and simplify workflow, especially helpful for high-throughput screening applications that involve multi-well plates.


Ideal for Most Observation Methods and Applications

PFS in Action

Nikon's PFS3 system is ideal for a variety of applications including TIRF. In most commercial applications of TIRF microscopy, a laser beam undergoes total internal reflection at the interface between the specimen and the coverslip, and fluorescence is excited with resulting evanescent wave. Since this wave only illuminates ~50 nm into the sample, tight control of focus is essential for effective long-term time-lapse TIRF imaging.

Additionally, many live cell imaging applications require the addition of media or a drug solution during the course of the experiment. As demonstrated in the accompanying images of fluorescent "beads" taken at the indicated times with and without the Perfect Focus System engaged, the change in temperature caused by adding media (indicated by the arrow) causes the focus to drift if PFS is not used; while engaging the PFS system completely eliminates this problem.

Another example of an application benefiting from the TiE-PFS is multipoint imaging. As shown in the accompanying DIC images of cells taken at multiple stage positions, it is possible to change the XY position of the microscope stage without losing focus, making true 6D multipoint imaging possible and truly revolutionizing long-term live cell imaging.

Because the PFS3 can maintain focus at deeper depths in the imaging specimen, it is ideal for developmental biology and applications that require studying the dynamics of cells in their natural environment such as thick tissues or whole animals.



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