84 MICROSCOPY
a
Suzuki, Y., Kajitani, K. and Ohde, H. (2015). Method for observing phase objects without halos or directional shadows. Optics letters; Vol. 40, No. 5. pp 812-815 2
REFERENCES: 1
The Nobel Assembly at Karolinska Institutet, (2012). [online] Available at:
http://www.nobelprize.org/nobel_prizes/
medicine/laureates/2012/
press.html [Accessed 22 Oct. 2015]. 3
F. A. Ross, Phase Contrast and Interference Microscopy for
Cell Biologists (Edward Arnold, 1967) 4
R. Hoffman, J. Microsc. 110, 205 (1977)
5. K. Okita, T. Ichisaka, and S. Yamanaka, Nature 448, 313 (2007)
b
detect changes in morphology suggesting cellular health or differentiation. Significantly, the IVC method has been applied to visualise the outline and structure of mouse iPS cells (iPS-MEF- Ng-20D-17, Kyoto University5
),
capturing the 3D nature of the cells when compared to brightfield and phase contrast (Fig. 2).1
Interestingly, IVC has c
Fig. 2. Comparing visualisation methods to observe mouse iPS cell colonies. Inversion contrast observation (a) provides greater contrast when compared to brightfield imaging (b) and standard phase contrast (c)
annular aperture is shifted slightly from the position of the pupil of the objective, and the area of the light flux passing through the objective increases. Te increased light forms an image devoid of halos, and with the concentric arrangement of annular aperture and objective, the setup is independent of
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phase shift direction, eliminating directional shadows. Te contrast is also inverted across the focal plane and the depth of field is reduced, facilitating the morphological analysis of 3D cells such as iPS cell colonies. Clearly observing iPS cell colonies, it is possible to
been found to be particularly ideal for distinguishing iPS cells from the surrounding feeder cells, as the contrast of the feeder cells is lower due to their flatter morphology, highlighting the iPS cells against the background. Removing the interference of halos or directional shadows that appear when using standard phase contrast illumination, the novel IVC technique presents many benefits for observing and documenting cell cultures. Continuing to drive recent advances in the areas of regenerative medicine and stem cell research, this method will become part of a complete range of cell culture technologies centred on Olympus’ optical expertise – including the CKX53 cell culture microscope – that are set to play an important role in tomorrow’s discoveries.
For more information ✔ at
www.scientistlive.com/eurolab
Wolfgang Hempell & Dr Joachim Kirschner are with Olympus Europe.
www.olympus-lifescience.com
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