Multiplane imaging and three dimensional nanoscale particle tracking in biological microscopy.
A conventional microscope produces a sharp image from just a single object-plane. This is often a limitation, notably in cell biology. We present a microscope attachment which records sharp images from several object-planes simultaneously. The key concept is to introduce a distorted diffraction grating into the optical system, establishing an order-dependent focussing power in order to generate several images, each arising from a different object-plane. We exploit this multiplane imaging not just for bio-imaging but also for nano-particle tracking, achieving approximately 10 nm z position resolution by parameterising the images with an image sharpness metric.
Dalgarno
Paul A
PA
School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, UK. P.A.Dalgarno@hw.ac.uk
Dalgarno
Heather I C
HI
Putoud
Aurélie
A
Lambert
Robert
R
Paterson
Lynn
L
Logan
David C
DC
Towers
David P
DP
Warburton
Richard J
RJ
Greenaway
Alan H
AH
D004867:Equipment Design; D019544:Equipment Failure Analysis; D007089:Image Enhancement / Q000295:instrumentation*; D021621:Imaging, Three-Dimensional / Q000295:instrumentation*; D008853:Microscopy / Q000295:instrumentation*; D053758:Nanoparticles / Q000648:ultrastructure*; D036103:Nanotechnology / Q000295:instrumentation*; D012031:Refractometry / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity