Depth of Field: Guide for Beginners - what is dof
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High content imaging is primarily concerned with the automated analysis of large cell populations where the goal is to process as many cells as possible in the fastest time with the highest resolution.
Physics and biophysics imaging encompasses a wide range of techniques used to interrogate physical phenomena using high tech imaging systems.
A microscope C-mount or F-mount adaptor is needed to connect a scientific camera to the microscope camera port. The mount threading is standardized which means that a C-mount adaptor will connect to all scientific cameras that connect via C-mount. However, the adaptors are microscope specific which means that although any C-mount camera will connect to a C-mount adaptor, the adaptor will only fit microscopes of the matching brand.
The QImaging CCD family of scientific cameras are designed with solutions for electrophysiology, long stare, color imaging, documentation and live cell imaging.
Adapters can have lenses in them to magnify or demagnify the image before it reaches the camera. This can be used to better match the camera FOV to the microscope FOV. For example, if the camera has an 11 mm diagonal FOV but the microscope is capable of an 18 mm FOV, a 0.67x adaptor would demagnify the image and allow it to be displayed on the 11 mm camera. However, this increase in FOV comes at the cost of reduced resolution.
By recognizing that FOV requirements can be highly variable, we are able to better serve the needs of our customers and offer a broad range of camera FOV options.
Supplying custom cameras to instrument designers for most of our 40 year history, we will work with you every step of the way.
It’s usually possible to find the maximum FOV of the microscope by referring to the field number (FN) displayed on the eyepieces and on some objective lenses. The field number is simply the maximum FOV of measured as a diameter the objective or eyepiece in millimetres, so an objective lens with a field number of 18 would have a maximum FOV of 18 mm. However, the field number always assumes no magnification so to calculate the actual FOV, the field number should be divided by the objective magnification:
The QImaging CCD family of scientific cameras are designed with solutions for electrophysiology, long stare, color imaging, documentation and live cell imaging.
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In microscopy, it is vital to have some form of contrast or stain that gives areas of the sample color and makes it possible to image. Advanced fluorescence microscopy techniques take advantage of this.
Biochip, genomics and microarray detection represent a large mix of applications with varying needs of a scientific camera.
The Evolve family of cameras are high-resolution, back-illuminated EMCCD providing high sensitivity for the lowest light applications.
In microscopy, it is vital to have some form of contrast or stain that gives areas of the sample color and makes it possible to image. Advanced fluorescence microscopy techniques take advantage of this.
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The Prime series of 95% quantum efficient, back-illuminated sCMOS cameras are designed to support the most demanding, low-light research applications
The FOV of a microscope is ultimately limited by a number of factors, such as the objective lens, the tube-diameter of the microscope’s internal optical-system, the eyepieces, the scientific camera sensor size and the camera mounting adaptor
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The development of larger FOV microscopes and scientific cameras that can take advantage of the F-mount is relatively recent – at the time of writing only one commercially available 25 mm microscope exists. Most modern microscopes have a 19 mm or 22 mm FOV and are therefore still able to use the C-mount. The largest format spinning disk confocal systems are also limited to a 22 mm FOV.
Camera specification sheets will display the camera FOV as a diagonal measurement (usually in millimeters). Ideally, the diagonal camera FOV should match the diameter of the microscope FOV to capture as much of the available image as possible. However, this does mean that the horizontal and vertical FOV of the camera will be less than the microscope diameter.
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If the goal is simply to attach the camera to the microscope, a 1x adaptor contains no additional lenses and provides no additional magnification or demagnification. This is often the preferred method as it introduces no additional lenses into the system. Every extra lens reduces the number of photons reaching the camera by 3-4% so many researchers will try to avoid this.
The maximum field of view of the microscope is affected by the objective lens, the tube-diameter of the microscope’s internal optical-system, the eyepieces, the scientific camera sensor size, and the camera mounting adaptor. For optimal imaging performance, it’s best to match the microscope FOV to the scientific camera FOV to capture as much information as possible and avoid vignetting. Teledyne Photometrics cameras are designed to match these specifications to offer the maximum field of view possible.
Supplying custom cameras to instrument designers for most of our 40 year history, we will work with you every step of the way.
It’s possible to use a camera with a larger diagonal FOV than the microscope to capture the entire microscope FOV (Figure 4). However, this is not optimal as there will be substantial vignetting at the corners of the image. Ideally, when choosing a scientific camera, it should have a diagonal FOV that matches the specifications of the microscope it will be used with.
The Evolve family of cameras are high-resolution, back-illuminated EMCCD providing high sensitivity for the lowest light applications.
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All cameras are controllable with the PVCAM driver and supported in Ocular software. The PVCAM driver SDK can also be used integrate into other software packages.
A 20x objective with a field number of 18 would actually have a FOV of 0.9 mm. Likewise, a 100x objective with a field number of 18 would have a FOV of 0.18 mm. The more an object is magnified, the smaller the field of view will be. Therefore, when looking to increase FOV, one of the first considerations should always be whether it’s possible to decrease magnification (Figure 2).
The Iris family of sCMOS cameras deliver up to a 15 megapixel sensor with a 25 millimetre field of view for high-resolution imaging over a large imaging area.
Adaptors can also affect the microscope and camera FOV depending on the type of adaptor used. A C-mount adaptor is the most popular microscope camera adaptor and is restricted to a maximum 22 mm FOV. The F-mount adaptor is a larger format adaptor capable of reaching >30 mm FOV.
Using the field number to calculate microscope FOV works well when imaging using the eyepieces but not when imaging using a scientific camera. Like most digital cameras, scientific cameras use square or rectangular sensors. This means that a scientific camera cannot capture the whole, circular FOV that the microscope is capable of. Instead, the camera FOV must fit inside the microscope FOV (Figure 3).
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The Prime series of 95% quantum efficient, back-illuminated sCMOS cameras are designed to support the most demanding, low-light research applications
Microscope field of view (FOV) is the maximum area visible when looking through the microscope eyepiece (eyepiece FOV) or scientific camera (camera FOV), usually quoted as a diameter measurement (Figure 1). Maximizing FOV is desirable for many applications because the increased throughput results in more data collected which gives a better statistical measurement for detecting subtle effects and also decreases time needed at the microscope.
Biochip, genomics and microarray detection represent a large mix of applications with varying needs of a scientific camera.
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See what others are doing. Stories and images from scientists using our high-performance sCMOS, EMCCD and CCD cameras to advance their research.
The brand new Kinetix family of back-illuminated sCMOS cameras delivers a framerate and field of view unmatched by any other sCMOS camera.
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All cameras are controllable with the PVCAM driver and supported in Ocular software. The PVCAM driver SDK can also be used integrate into other software packages.
CMOS made scientific. The Moment is a true global shutter CMOS camera with an ultra-compact form factor, powered through USB 3.2 Gen 2.
High content imaging is primarily concerned with the automated analysis of large cell populations where the goal is to process as many cells as possible in the fastest time with the highest resolution.
The brand new Kinetix family of back-illuminated sCMOS cameras delivers a framerate and field of view unmatched by any other sCMOS camera.
Physics and biophysics imaging encompasses a wide range of techniques used to interrogate physical phenomena using high tech imaging systems.
At Teledyne Photometrics, we aim to create cameras that can optimally match the FOV of all modern microscopes (Table 1). For this reason, the Prime 95B Series comprises a 19 mm camera, a 22 mm camera and a 25 mm camera. Additionally, the Prime BSI and Iris 9 both fit a 19 mm microscope FOV and the Iris 15 fits a 25 mm microscope FOV. The Kinetix is our largest format sensor which is able to be used to get the maximum FOV out of any system up to 29 mm.
The Iris family of sCMOS cameras deliver up to a 15 megapixel sensor with a 25 millimetre field of view for high-resolution imaging over a large imaging area.
Cooled, low-noise CMOS cameras designed for integration. With unprecedented thermal control, Retiga E cameras are capable of exposures over an hour!