Mirrored pyramidal wells for simultaneous multiple vantage point microscopy.
We report a novel method for obtaining simultaneous images from multiple vantage points of a microscopic specimen using size-matched microscopic mirrors created from anisotropically etched silicon. The resulting pyramidal wells enable bright-field and fluorescent side-view images, and when combined with z-sectioning, provide additional information for 3D reconstructions of the specimen. We have demonstrated the 3D localization and tracking over time of the centrosome of a live Dictyostelium discoideum. The simultaneous acquisition of images from multiple perspectives also provides a five-fold increase in the theoretical collection efficiency of emitted photons, a property which may be useful for low-light imaging modalities such as bioluminescence, or low abundance surface-marker labelling.
Seale
K T
KT
Department of Biological Sciences, Vanderbilt University, VU Station B 351807, Nashville, TN 37235-1807, USA.
Reiserer
R S
RS
Markov
D A
DA
Ges
I A
IA
Wright
C
C
Janetopoulos
C
C
Wikswo
J P
JP
D000818:Animals; D004023:Dictyostelium / Q000648:ultrastructure; D021621:Imaging, Three-Dimensional / Q000379:methods; D008853:Microscopy / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods*; D015388:Organelles / Q000648:ultrastructure; D013048:Specimen Handling / Q000379:methods*