Q-PHASE

TESCAN USA

The Q-PHASE is a unique instrument for quantitative phase imaging (QPI) based on patented technology of Coherence-controlled holographic microscopy. The Q-PHASE is purposely designed to observe living cells in vitro.

TESCAN proudly introduces the Q-PHASE, a multimodal holographic microscope (MHM).  With this instrument TESCAN expands into the field of advanced light microscopy. The Q-PHASE is a unique instrument for quantitative phase imaging (QPI) based on patented technology of Coherence-controlled holographic microscopy. This technology uses incoherent light sources (halogen lamp, LED) providing QPI with the highest quality, without any compromises and it is the only QPI technique enabling imaging of samples in scattering media. The Q-PHASE is purposely designed to observe living cells in vitro. It is based on a robust inverted transmission microscope platform. The whole system is situated in a microscope incubator. The full motorization fulfills even the highest demands regarding experiment automation. Furthermore, this system includes multiple imaging modes with fully integrated Fluorescence Module, simulated DIC and brightfield imaging options. All these features make of Q-PHASE a valuable research tool for biological and biotechnical applications such as testing reactions of cells to a specific treatment - even with scattering non-transparent substances, monitoring cell’s life cycle including mitosis, distinguishing between different forms of cell deaths, analyzing cell growth, motility or morphology changes, imaging cells in extracellular matrices.

Key features and advantages

  • ‚‚No image artifacts such as halo effect (as opposed to techniques based on Zernike phase contrast illumination)
  • ‚‚Enables very precise detection of cell boundaries
  • ‚‚Strong suppression of coherent noise (speckles) & parasitic interferences (as opposed to laser-based approaches)
  • ‚‚Label-free – no staining is needed, simple sample preparation, observation of live cells in their native environment, no photobleaching problems
  • Low phototoxicity – low light power density (107× lower than fluorescence microscopy) allows long-term observations (for days)
  • Coherence-gating effect – Q-PHASE special feature enabling to observe samples even in scattering media (phospholipid emulsions, extracellular matrices, etc.)
  • Multimodality – fully integrated fluorescence module, simulated DIC and brightfield which enables automatic multimodal imaging of the sample
  • High-quality QPI – unique Q-PHASE’s optical setup allows using incoherent illumination which provides extraordinary imaging quality without any compromises
  • ‚‚Lateral resolution of conventional microscopes (up to 2× better when compared to common laser-based approaches or pinhole spatial filtering based techniques)
  • Fast acquisition – the use of off-axis holographic approach makes Q-PHASE a single-shot instrument, thus enabling imaging of very fast cell dynamics
  • Full motorization – focusing, sample stage, objective exchange, fluorescence filters
  • Automated multidimensional acquisition – time-lapse, channel, position, Z-stack
  • Simple image segmentation and processing – comparable to fluorescence data processing
  • Quantitative – phase values can be recalculated e.g. to cell dry-mass density (pg/μm2) or direct topography with nanometer sensitivity (usually non-biological samples with homogeneous refractive index distribution)
  • High phase detection sensitivity – enables to detect even the smallest changes in axial direction, very sensitive detection of morphology or position changes
ccQPI of non-metastatic fibroblast-like LW13K2 (K2) cells QPI of rat sarcoma cells, cell tracking QPI of human adipocyte
QPI of non-metastatic fibroblast-like LW13K2 (K2) cells QPI of rat sarcoma cells, cell tracking QPI of human adipocytes

 

Working principle

Working principle - 01
  • The illumination beam is divided into two beams (object and reference beam) forming two arms of the microscope

 

  • The sample is put into the object arm

 

  • The reference sample (blank) is put into the reference arm

 

   
 Working principle - 02  
  • The beams pass through the sample and the reference sample
  • The reference beam is spectrally decomposed on diffraction grating
   
 Working principle - 03  
  • Beams from both arms are focused onto the CCD camera
 
  • The beams interfere and form a hologram
 
  • The hologram is recorded and further processed on PC to produce quantitative phase images
 Working principle

Intrinsic imaging modes

Complementary image contrast can be obtained just by numerical processing of the acquired phase images. In this way simulated DIC images can be produced with adjustable shear and displayed in real time. Another possibility is the brightfield imaging which can be simply achieved by closing the reference arm of the microscope. In summary, the Q-PHASE offers multiple imaging modes widely used in biological research such as fluorescence or DIC integrated in a single instrument and supported by the Q-PHASE’s software allowing fully automated multimodal imaging.

Multimodal Imaging of human malignant melanoma cells
Quantitative phase image of human malignant melanoma cells Simulated DIC of human malignant melanoma cells Brighfield image of human malignant melanoma cells
Quantitative phase image Simulated DIC Brightfield

Imaging in scattering media

A special feature of Q-PHASE is the coherence-gating, a well-known effect in optical coherence tomography which enables observations of samples even in scattering media. This effect is induced by using incoherent light in the unique patented setup of Q-PHASE. Its transmitted-light configuration enables to effectively suppress the light which was scattered by the environment in defocused planes and to only use unscattered light for imaging. In this way, cells can be observed even in moderately scattering non- transparent substances such as an active phospholipid emulsion.

QPI of reaction of human colorectal cancer cells to treatment QPI of reaction of human colorectal cancer cells to treatment QPI of reaction of human colorectal cancer cells to treatment
QPI of reaction of human colorectal cancer cells to treatment by 0.15% active phospholipid emulsion (scattering medium)

 

Imaging in extracellular matrices

The coherence-gating effect can also be beneficial when imaging cells in extracellular matrices such as collagen gel. Extracellular matrices mimic in vivo situation making the study of the cell’s dynamic reactions to its surroundings more realistic. Usually it is used as a biological test for cancer cell invasivity and ability to metastasis. The Q-PHASE microscope enables to record mechanism of cell motion and interactions between extracellular matrix fibers and cells with high contrast and without any additional staining.

QPI of human sarcoma cell motion mechanism in collagen gel QPI of human sarcoma cell motion mechanism in collagen gel
QPI of human sarcoma cell motion mechanism in collagen gel (inverted LUT)

The Q-PHASE’s software is an integral part of this microscope. It runs on the 64-bit WindowsTM edition. It is divided into two modes, Live and Data-set, which provide all the necessary functionality for experimental measurements, image processing and analysis. Simple graphical interface with tabs allows easy orientation. Exporting images to standard formats allowing to process and analyze the images with third party software. The implemented GPU acceleration gives the highest performance.

  • Full microscope control (focusing, sample stage, objective exchange, fluorescence channels, light-source, shutters, camera settings)
  • Multidimensional image acquisition (time-lapse, channel, position, Z-stack)
  • GPU accelerated real-time holographic image reconstruction
  • Multidimensional dataset viewer
  • Image processing and analysis
  • Microscope control with control panel
  • User management
  • Microscope alignment wizard with automatic procedures
  • Microscope incubator control

Fluorescence module

The Q-PHASE can combine holographic microscopy with the fluorescence microscopy. This powerful combination provides the possibility to verify structures or processes observed in QPI with fluorescence microscopy in the same field of view using a single instrument. For example, morphological and position changes prior to cell death can be observed in QPI with following fluorescence verification of cell death types. This approach greatly reduces the phototoxicity and photobleaching problems of fluorescence imaging and it allows long-term observations.The focus plane in both methods is located at the same position. This allows easy and fast switching between the two imaging methods at the same conditions and time points. Multiple fluorescence channels are possible with motorized channel exchange for automated multidimensional measurements.The illumination can be implemented by using liquid light guide coupled solid state light sources or a xenon arc lamp. Multidimensional image acquisition combining holography and fluorescence is fully integrated in the Q-PHASE’s software. A fluorescence module is attached to the side port of the Q-PHASE, which can alternatively be used for other imaging techniques.

Fluorescence module attached to the side port

Microscope incubator

Maintains constant temperature – suitable conditions for long-term live cell imaging. Computer temperature setting and temperature data logging.

 

 

Piezo-focusing insert for precise focusing (500 µm travel range)

 

Control panel with multifunctional touchscreen, sample stage joystick and rotary knobs

 

Stage top incubator for precise control of temperature, humidity and CO2 concentrations

 

 

 

 

TESCAN USA

QPI of Rat healthy cell

TESCAN USA

Extracellular Matrix

TESCAN USA

Fluorescence image of human prostate cancer cells

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TESCAN USA

TESCAN proudly introduces the Q-PHASE, a multimodal holographic microscope (MHM).  With this instrument TESCAN expands into the field of advanced light microscopy.

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