Photosynthesis Research

, Volume 102, Issue 2, pp 111–141

Optical microscopy in photosynthesis

  • Richard Cisek
  • Leigh Spencer
  • Nicole Prent
  • Donatas Zigmantas
  • George S. Espie
  • Virginijus Barzda
Review

DOI: 10.1007/s11120-009-9500-9

Cite this article as:
Cisek, R., Spencer, L., Prent, N. et al. Photosynth Res (2009) 102: 111. doi:10.1007/s11120-009-9500-9

Abstract

Emerging as well as the most frequently used optical microscopy techniques are reviewed and image contrast generation methods in a microscope are presented, focusing on the nonlinear contrasts such as harmonic generation and multiphoton excitation fluorescence. Nonlinear microscopy presents numerous advantages over linear microscopy techniques including improved deep tissue imaging, optical sectioning, and imaging of live unstained samples. Nonetheless, with the exception of multiphoton excitation fluorescence, nonlinear microscopy is in its infancy, lacking protocols, users and applications; hence, this review focuses on the potential of nonlinear microscopy for studying photosynthetic organisms. Examples of nonlinear microscopic imaging are presented including isolated light-harvesting antenna complexes from higher plants, starch granules, chloroplasts, unicellular alga Chlamydomonas reinhardtii, and cyanobacteria Leptolyngbya sp. and Anabaena sp. While focusing on nonlinear microscopy techniques, second and third harmonic generation and multiphoton excitation fluorescence microscopy, other emerging nonlinear imaging modalities are described and several linear optical microscopy techniques are reviewed in order to clearly describe their capabilities and to highlight the advantages of nonlinear microscopy.

Keywords

Nonlinear microscopySecond harmonic generationThird harmonic generationChlamydomonasChloroplastsCyanobacteria

Abbreviations

3D

Three-dimensional

AOM

Acousto-optic microscopy

CARS

Coherent anti-Stokes Raman scattering

CCD

Charge-coupled device

CD

Circular dichroism

DIC

Differential interference contrast

F0

Minimum fluorescence yield

Fm

Maximum fluorescence yield

Fv

Variable fluorescence yield

FLIM

Fluorescence lifetime imaging microscopy

FPGA

Field-programmable gate array

GFP

Green fluorescent protein

IR

Infrared

LH2

Light-harvesting complex 2

LHCII

Light-harvesting pigment–protein complex of photosystem II

MPF

Multiphoton excitation fluorescence

mRNA

Messenger ribonucleic acid

NA

Numerical aperture

NSOM

Near-field scanning optical microscopy

PAM

Pulse amplitude modulation

PSI

Photosystem I

PSII

Photosystem II

PT

Photothermal

SHG

Second harmonic generation

SFG

Sum frequency generation

SLIM

Spectrally resolved fluorescence lifetime imaging microscopy

SNR

Signal to noise ratio

STED

Stimulated emission depletion

TCSPC

Time-correlated single photon counting

THG

Third harmonic generation

TIRF

Total internal reflection fluorescence

TR-PT

Time-resolved photothermal

Copyright information

© Springer Science+Business Media B.V. 2009

Authors and Affiliations

  • Richard Cisek
    • 1
  • Leigh Spencer
    • 1
  • Nicole Prent
    • 1
  • Donatas Zigmantas
    • 2
  • George S. Espie
    • 3
  • Virginijus Barzda
    • 1
  1. 1.Department of Chemical and Physical Sciences, Department of Physics, and Institute for Optical SciencesUniversity of TorontoMississaugaCanada
  2. 2.Department of Chemical PhysicsLund UniversityLundSweden
  3. 3.Department of BiologyUniversity of TorontoMississaugaCanada