05 Jun 2003
Researchers describe an all-optical technique for studying the structure of brain tissue at this week’s CLEO.
Delegates at this year’s CLEO in Baltimore, US, have heard a US collaboration describe an all-optical technique for studying the structure of brain tissue. The method, called serial histology, uses femtosecond laser pulses to examine consecutive slices of tissue and build up a three-dimensional image.
Histology, studying the make-up of tissue, typically involves staining the sample before it is examined by optical microscopy. Traditionally, the tissue is sliced manually but this is both labor intensive and may unduly affect the structures. Philbert Tsai and colleagues have now developed an automated optical process that uses femtosecond laser pulses to both slice and image the sample.
The iterative process involves three steps: stain, image and ablate. “We used femtosecond pulses to both image and ablate tissue for the purpose of serial sectioning,” report the authors. “The region of the tissue that has been imaged is subsequently removed by laser ablation with amplified femtosecond pulses.” The process is repeated until the complete sample has been studied.
Tsai and co-workers have reconstructed a 250 x 375 x 400 µm region of the neocortex of a mouse brain using just four successive cutting and imaging cycles. The team used a 850-nm laser in a two-photon scanning microscope to collect emission from a fluorescent protein in the brain cells. Laser pulses with an energy of between 0.4 to 1.7 µJ were then scanned across the sample at 5 mm/s to ablate a series of 10-micron-thick layers before imaging took place once again.
According to the authors, their approach does not require the tissue to be pre-frozen and requires no realignment of the imaged sections. “It is ideally suited for the analysis of transgenic animals with intrinsic fluorescent markers,” they say. “An entire mouse brain can be transformed into approximately 5 terapixels of data that shows the distribution of a label at the diffraction limit of spatial resolution.”
Jacqueline Hewett is news reporter on Optics.org and Opto & Laser Europe magazine.