Olympus LV200 Bioluminescence imaging system


Location: Maurice Shock Building, room 388.
Contact:
Mr Matthew Barker (mcb8/3085)
Dr Kees Straatman (krs5/7085/2263)

Bioluminescence is a chemical reaction in which light is emitted when the enzyme luciferase interacts with its substrate luciferin. This process requires oxygen and ATP and when introduced into a cell system only active and intact cells will produce a signal. As this is a chemical reaction with no excitation light it results in a low background signal which allows detection of very weak signals with precise measurement of small changes.

Bioluminescence can be used in whole animal plant systems (see the IVIS Spectrum bioluminescence/ fluorescence scanner in the Pre-Clinical Imaging Facility and the Fujifilm LAS4000 luminescence imager in the AIF), in cell systems as well as in a cell extract, after adding ATP. Bioluminescence microscopy can replace fluorescence microscopy if phototoxicity, photobleaching, low signal, autofluorescence or increased signal in dying cells become critical or mask the signal of interest. Historically, bioluminescence imaging is used to study circadian rhythms of gene expression, measured over several days.

Recently, Promega has introduced NanoLuc® Luciferase, a luciferases that is reported to be up to 150x brighter than the most used firefly luciferase allowing for shorter exposure times or imaging of faster cellular processes like calcium signaling.

Specifications for the LV200
This system is specifically developed for bioluminescence imaging. A built-in system for temperature control, humidity, and gas flow helps to keep the cultured cells or tissue slices healthy for long time (days) experiments. The system can handle small samples in 35 mm dishes.

LV200 with closed condenser (left) and with upwards turned condenser (right) revealing the culture chamber.

An Hamamatsu C9100-13 EM-CCD camera is used for image acquisition. This allows fast imaging at low light levels (using EM-CCD mode) as well as long exposures times at low light levels (using most likely CCD mode). Pixel size of the camera is 16 x 16 µm with an image size of 512 x 512 pixels.

Objectives available:
UPLSAPO 10x NA=0.4
UPLSAPO 20x NA=0.75
UPLSAPO 60x Oil NA=1.35

The system has a high NA tube lens optimized for luminescence imaging with a 0.2 x magnification, all objectives have therefore an effective magnification of magnification x 0.2 (2x, 4x and 12x).

 

Some publications using the LV200:

Hirobe et. al. (2014) Synergetic cytotoxic activity toward breast cancer cells enhanced by the combination of Antp-TPR hybrid peptide targeting Hsp90 and Hsp70-targeted peptide. BMC Cancer. 14: 615-626.

Lambrechts et al. (2014) A causal relation between bioluminescence and oxygen to quantify the cell niche. PLoS ONE  9: e97572.

Han et al. (2013) Theranostic protein targeting ErbB2 for bioluminescence imaging and therapy for cancer. PLoS ONE  8: e75288.

Saini et. al. (2012) Simulated body temperature rhythms reveal the phase-shifting behavior and plasticity of mammalian circadian oscillators. Genes Dev. (2012) 26: 56-580.

Myung et.al. (2012) Period Coding of Bmal1 Oscillators in the Suprachiasmatic Nucleus. J Neurosci 32: 8900-8918.

Fluegge et. al. (2012) Mitochondrial Ca2+ mobilization is a key element in olfactory signaling. Nat Neurosci. 15: 754-62.

Bolinger et. al. (2011) Circadian Clocks in Mouse and Human CD4+ T Cells. PLoS ONE  6:12.

Hirsch et al. (2011) A novel fry1 allele reveals the existence of a mutant phenotype unrelated to 5’->3’ exoribonuclease (XRN) activities in Arabidopsis thaliana roots. PLoS ONE. 6:e16724.

Guilding et al. (2010) Circadian oscillators in the epithalamus. Neuroscience. 169:1630-9.

Yagitaa et al. (2010) Development of the circadian oscillator during differentiation of mouse embryonic stem cells in vitro. PNAS 107: 3846-3851.

Guilding et al. (2009) A riot of rhythms: neuronal and glial circadian oscillators in the mediobasal hypothalamus. Mol Brain. 2: 28-47.

Kammerloher (2008) Advertising Feature; Bioluminescence microscopy for cellular level circadian analysis in the suprachiasmatic nucleus. Application note. Nature Methods 5, V-VI.

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