Confocal microscopy is an optical imaging technique that uses point-by-point illumination and spatial pinhole modulation to remove scattered light from the non-focal plane of the sample, allowing for improved optical resolution and visual contrast compared to traditional imaging methods. Alfa Chemistry, the imaging experts, can provide you with Laser Scanning Confocal Microscopy (LSCM)-based confocal microscopy imaging services.
Laser Scanning Confocal Microscopy (LSCM)
Current instrumentation
The current LSCM at the AAC is a BioRad Radiance 2000 with the following configuration:
Laser configuration
1. Argon ion - 488 nm(14mW); 514 nm(11mW)
2. Green HeNe - 543 nm(1.5mW)
3. Red laser diode - 638 nm(5mW)
Emission Filters
Detector (photomultiplier - PMT) Emission filters (wavelength in nm)
1. Open; 488/10; 500LP; 515/30; 530/60;
2. Open; 515/30; 530/60; 570LP; 590/70; 600/50; 600LP
3. Open; 660LP
It is based around the light optics of a Nikon E600 upright microscope with tungsten transmitted and reflected light sources and a mercury lamp. Current lenses available are; 10, 20 and 40× (cover slip corrected); 50 and 100× (air); 60× oil immersion and 40× (water immersion).
LSCM is used in many applications for imaging biological specimens. Although it cannot improve on magnifications available to conventional fluorescence microscopy the sharper, clearer images obtained greatly improving image quality. Also, as the system acquires images digitally, timed images may be automatically generated. In samples that are relatively transparent to the lasers the ability to create sequential images through the depth of a specimen ("z-series or stacks) produce 3-dimensional information provides real spatial context to the features of interest. Opaque materials can also be imaged this way, essentially recording reflected light. Thus it is also a useful technique in material sciences for modelling material in 3D.
Sample requirements
For fluorescence imaging specimens need by fixed and stained using fluorphores that will not only attach (label) to the relevant feature of interest but are excited by and emit at wavelengths appropriate to the configuration of lasers and detector filters available.
Laser scanning confocal microscopy (LSCM)is a system used in epifluorescence and reflected light imaging. A finely focused beam of laser light is scanned across a sample and the resultant light emitted passed through a pinhole aperture to exclude any out of focus light. The image thus produced includes only the plane of focus for a given objective lens (hence a clearer image). Additionally, by varying the stage height, a series of sequential images through the thickness of a single sample can be collected and used to project a three-dimensional image.
A LSCM will typically include at least 3 different wavelength lasers which can be used simultaneously or sequentially. For imaging fluorescence in a sample this means multiple stains (fluorophores) can be used to highlight specific features of interest.
Alfa Chemistry' Laser Scanning Confocal Microscopy (LSCM)-based confocal microscopy service is an ideal partner for cutting-edge biomedical research, providing precise 3D imaging, as well as accurate imaging of cell structure and dynamic processes, all in one package. If you are interested, please feel free to contact us.
For research use only, not intended for any clinical use.
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