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See Below For a Glossary Of Terms and Choosing a Camera Guide.

GLOSSARY OF TERMS

Aberration - an optical defect in the design of a lens. The lens does not bring all the rays of light to an exact focus. There are several different types of aberrations each having a contributing factor on image quality.

Achromat - A lens system that has been designed to reduce chromatic aberration. A photographic lens system is usually corrected to provide the same focal length for the red and blue wavelengths.

Achromatic - color-corrected optics used to produce true specimen color.

Achromatic Condenser - a condenser corrected for both spherical aberration. It is the most common type found on bright field microscopes.

Alignment - condition in which all optical elements are centered on the same axis.

Aperture - a fixed or adjustable opening or hole through which light may pass. Located beneath the microscope stage.

Apochromat - A lens system in which chromatic aberration is corrected for three or more colors. Apochromatic lenses are used in photoengraving applications and for telephoto lenses that have large maximum apertures.

Arm - part that serves as both the support of the body tube and its lens systems and as the held part of the microscope when it is carried.

Base - the weighted bottom of the microscope which gives it both good balance and resistance to unexpected movement , "vibration."

Body Tube - the main structure which holds the eyepiece and objectives at a precise distance to each other, and which allows this combined system to approach the stage into clear focus.

Brightfield - used to examine specimens which have contrast/color (i.e. stained histological specimens.)

Components - a brightfield condenser and brightfield objectives.

Disadvantages - cannot be used to view specimens of little or no contrast.

Brightfield Illumination - the ray of light incident upon the object is parallel to the optical axis.

CCD-Charged Coupled Device. This is the image detector in a video camera.

Chromatic Aberration - an optical defect of a lens, seen as color fringes or halos, which causes different wavelengths of light to be focused at different distances from the lens.

Coarse Adjustment - the knob which moves the specimen (or objective) rapidly, vertically for focusing.

Color Balance - a method of correcting a light source by way of filtration so that the light has optical characteristics similar to sun light.

Color Temperature - the quantitative value indicating the amount of color or colors emitted by an object.

Condenser - the lens system between the illuminator and the specimen which condenses the light and focuses on the specimen.

Confocal - an interferometric microscope utilizing a rotating aperture and a CCD image analysis.

Contrast - light and dark. To produce a good image, you must have good contrast as well as good resolving power (N.A.).

Corrected Lens - a lens or lens system which corrects for specific aberrations.

 Darkfield - used to examine specimens which cannot be distinguished from the background; (i.e. syphilitic spirochetes, silver grains in auto-radiography.)

Components - a dry darkfield condenser for low magnifications and any low magnification objectives. An oil darkfield condenser for higher magnifications. High magnification oil objectives must have an iris.

Darkfield Illumination - the ray of light incident upon the object is at an angel from the optical axis; therefore scratches and dents on the surface are illuminated while the other intact part remains dark.

Daylight Filter - a blue filter used to correct the color temperature of a light source.

Depth of Focus - range around the focal point in which the image is still clear.  The larger the N.A., the smaller the depth of focus.

Differential Interference Contrast - (Often called Nomarski)-used to examine round, multi-cellular, tubular or thick transparent specimens.

Components - DIC condenser, re-combination prism, DIC objectives.

A very specialized item and usually ordered for specific applications.

Disadvantage - price; cannot be used with plastic dishes.

DIN - Deutch Industrie Norm-international standard used in the manufacture of interchangeable objective lenses. A typical DIN type microscope utilizes 45mm objectives+160mm tube length for a 205mm system length.

Diaphragm - a rotary disk, located under the stage, whose graded opening allows a variety of angles of light to come up through the stage opening. Also available as a continuously varying diameter adjusted by a lever located on the side of the component.

Drawing Tube (Camera Lucida) - enables a person to view the specimen and the paper/pencil simultaneously for drawing.

Dual Viewing Attachment - enables two people to view the same specimen through the same microscope simultaneously.

Empty Magnification - magnification which increases the size, but does not increase the detail, due to the limitation of the resolving power of the optical system.

Eyepiece - the lens system closest to the eye. Magnifies the image produced by the objective. So, with a 10X objective and 10X eyepiece, the total magnification is 100X.

Eyepiece tube - a smaller diameter extension of the body tube which receives the eyepiece.

Eye Point - the location of the eye when using a visual instrument which allows for the best possible viewing.

Eye relief - the distance from the eye lens of the microscope to the eye point.

Fluorescence - use to locate fluorescently tagged material (protein, enzyme, gene) by illuminating the material with one wavelength of light in hopes that the fluorescence tag will appear by emitting this light at a different wavelength; i.e. FITC labeled materials in tissue when illuminated with blue light will emit with green light.

Components - Epi fluorescence attachment, a mercury (or other) light source, power supply, specialized filter systems, each one for a different fluorescent tag.

Disadvantage - Specimen is usually fragile, unstable and cannot be used for long periods or stored.

Fluorite - an objective corrected for two wavelengths and therefore, with a higher resolving power than an Achromat. (There are exceptions) Manufacturers call them Fluor, Neo-Fluor, Fluotar.

Field - the actual diameter of the viewing area, usually expressed in millimeters. As magnifying power is increased, the field of view is decreased.

Filter - a colored transparent material placed in the path of illumination to vary the conditions of viewing.

Filter mount - an existing slot on the microscope which can hold one of a set of filters in the path of illumination.

Fine Adjustment - the knob which moves the specimen (or objective) slowly, vertically for focusing.

Finity Correction System - an optical system in which the image is formed only by an objective lens.

Focal Length - distance between a principal point and a focal point.   f1 is a focal length of objective, f2 is a focal length of tube lens.  Magnification is determined by the ratio of objective lens focal length and tube lens focal length. (for Infinity correction system)

Focus Rack - part of the assembly (plus pinion) which specifically allows the distance from the objective lens to the specimen to vary, and therefore permit required coarse and fine focus adjustments.

Fully (Anti-reflective) Coated - coating of a lens which improves its ability to transmit light more efficiently. 

Hoffman Modulation Contrast - called the "poor man's" Nomarski. Used to examine the same types of specimens as DIC/Nomarski.

Components - Hoffman condenser, polarizer and Hoffman objectives.

Disadvantages- Loss of some resolving power.

Huygenian Eyepiece - an eyepiece which effects a certain amount of correction for chromatic difference of magnification in the achromatic objective lens.

Illuminating Mirror - usually a two-sided mirror on a rotating axis located below the stage. It reflects light up into the stage opening and onto the specimen.

Illumination - the supply of light onto the object under the objective lens.

Illuminator - the source of light which illuminates the object or specimen to be observed. May be fixed intensity, or variable.

Image Plane - a plane which is at right angles to the optical axis at the image point.

Inclination Joint - located at the base of the arm of some microscopes allowing the viewer to tilt the microscope at a more comfortable angle to look at dry slides.

Incline(30-45 degrees) - the eyepiece tube is manufactured at an angle to the horizontal to allow more comfortable posture during long periods of observation.

Infinity Correction System - an optical system in which the image is formed by an objective lens and a tube lens.

Interchangeable Eyepiece - a lens system which fits within the eyepiece tube and which is usually replaced by another lens of equal mounting diameter but of different magnification power.

Interchangeable Objective - a lens system threaded into the nosepiece turret which can be removed and which is usually replaced by another lens of equal mounting diameter and threading, but different magnification power. 

JIS Standard-Japanese Industrial Standard -international standard used in the manufacture of interchangeable objective lenses. A Typical JIS type microscope utilizes 36mm objectives+170mm tube length for a 206mm system.

Lens - optical glass which has two polished surfaces and is used to converge or diverge light rays.

Light - electromagnetic radiation.

Long Working Distance - usually an objective with a greater than normal working distance.

Used in micro-manipulation, inverted microscopy, producing micro instruments. A normal 40X objective will have a working distance of 0.06mm. A long working distance 40X will have 2.4mm; an extra long working distance 40X will have 10mm; a super long working distance 40X will have 15mm. The longer the working distance, the lower the resolving power..

Magnification - an enlargement of an object by an optical instrument. It is the ratio of the size of the image to the actual size of the object under observation.

Magnifying Power - a measure of the ability of a lens or combination of lenses to make an object appear larger. It is the number of times the image seen through the microscope is larger than the object would appear to the unaided eye.

Micro-manipulator - used to inject or extract substances from the specimen. Also used to measure electric current produced by the specimen.

Microscope - a high precision optical instrument which uses light to observe objects. It is capable of high magnification and resolution and is used for making minute details visible.

Motorized Scanning Stages - used to allow a computer to control movement of the specimen. Highly specialized and rare at the moment but soon will become the "in" thing in combination with CCTV.

Multi-viewing Attachment - enables 2-8 people to view the same specimen through the same microscope simultaneously. 

N.A. - Numerical Aperture.  N.A determines resolving power, focal depth, and luminosity of the image.  The larger N.A., is, the higher resolving power and smaller focal depth are.

Nosepiece - a rotary turret mounting for the set of objectives.

Numerical Aperture (N.A.) - a number to represent resolving power of an objective.

Objective Lens - the compound lens system in a microscope which receives light from the field of view and forms the first image. The lens system closest to the specimen. Produces the primary magnification; i.e. 2X, 4X, 10X.

Ocular Micrometer - inserted in the eyepiece, it enables the person to measure the specimen.

Oil Immersion - a technique of placing special oil between positioned 100X objective and the glass slide on the stage to improve the resolving quality at high magnification. Also placed between the condenser and the glass slide.

Parallax - an imaging error introduced when using a stage micrometer of significant thickness. This is when the scale and the specimen appear to be in the same plane, thus causing incorrect measurements.

Parfocal - the ability to rotate the objective turret with minimal refocusing.

Parfocal Length - distance between the surface of the specimen and objective lens mounting position when in focus.

Phase Contrast - used to examine live, unstained specimens.

Components - Phase condenser and phase objectives.

Disadvantages- produces an optical artifact called the phase halo. It is, therefore, completely unusable for round, multi-cellular or tubular specimens. i.e. eggs, worms (nematodes), neurons.]

 

Photo-Micrographic Camera - Can only be attached to a trinocular tube.

Provides a clear focus, minimum to no vibration and great flexibility in interchangeability of film holders; i.e.-35mm. Polaroid 3 x 4 or Polaroid 4 x 5. Vary greatly in complexity and degree of sensitivity; (i.e. manual, semi-automatic or fully automatic.)

Photomicrography - the process of documenting images on film as seen through a microscope.

Plan Objective - an objective corrected for flatness of field so that when you view the specimen it is in focus all across the field. We then have Plan Achromats, Plan Fluorites, and Plan Apochromats.

Polarizing Components - can be added to an existing basic microscope to locate birefringent materials (i.e. crystals, etc.). 

Rack and Pinion - the intermeshing of a geared wheel and matching vertical grooved rack to develop tight accurate focusing.

Ramsden Eyepiece - an eyepiece similar to the Huygenian eyepiece. It differs in that it has its focal plane either on or just outside the surface of the collective lenses.

Real Field of View - range (diameter) of specimen observable with a microscope.

Resolving Power - the ability resolve two points at a given distance.

Rotational Viewing (360 degrees) - a microscope set up in such a way that the eyepiece and eyepiece tube can rotate around horizontally. Several people gathered around the same instrument can thus view the same specimen without moving, or having to move the microscope.

Scanning Electron Microscope - a microscope which uses a beam of electrons to impact the specimen. Secondary electrons emitted at the point of impact are imaged by high tech scanners.

Spherical Aberration - an optical defect in which the lens fails to form a sharp image. Rays of light which pass through a lens near its edge are converged to a point nearer the lens than those rays passing through the center of the lens.

Stage - the platform which hold the specimen.

Mechanical Stage - moves the specimen East to West and North to South.

Rotating Stage - usually rotates 360 degrees or, in the case of a rotating mechanical stage, as much as 270 degrees.

Stage Clips - fasteners located on the stage and placed over slides to hold them securely in place while viewing.

Stage Micrometer - used to calibrate the ocular micrometer.

Stand - the basic component of the microscope which holds all of the other components. Usually contains the light source (illuminator).

Stop Screw - an adjustable screw located at the base of the focus rack which, when adjusted properly, prevents the body tube from lowering too far and potentially causing damage to both the highest power objective and the specimen. 

Tube Length - the optical distance from the objective to the eyepiece. Governs interchangeability of optical components; i.e. a microscope objective corrected for 160mm. tube length cannot be used on a microscope corrected for infinity.

Turret-mounted - all objectives are attached to one common rotating nosepiece to allow for quick and accurate objective positioning during viewing.

Ultra-violet - the portion of the spectrum where the wavelengths are below the visible spectrum.

Video Microscope - enables the specimen to be viewed on a video screen. Image can also be analyzed by a computer.

Components - "C" Mount (interface between closed circuit TV cameras and microscope), CCTV Camera, monitor and possibly an image processor.

Disadvantages - Expense in the case of the more specialized systems only.

Virtual Image - an image that does not converge in real space.

Wavelength - light travels in waves varying in length. Measurement is from the top of one wave tot he top of the next one and is usually measured in units of nanometers (nm) or Angstoms (A).

Wide Field Eyepiece - an eyepiece having a large field of view with a high eye point.

Working Distance - the distance from the front element of the objective to the specimen.

CHOOSING A CAMERA / IMAGING SYSTEM

Optekusa.com can assist you in choosing a camera / imaging system. We can supply almost any combination that you may require. We manufacture and sell most all interconnect devices necessary to a successful installation. We can supply computers with imaging adapter cards and software or install and configure your system. Contact us for your needs. 

When choosing a camera or imaging system, your first priority should be to look at your application requirements. You should ask yourself and answer the following questions:

Will you need special filtering or lighting techniques to provide a usable image of your specimen or object? Keep in mind the material that you are viewing and the surface finish. Highly reflective objects may appear ok in the eyepiece but a camera may interpret the image differently.
What field of view does the application require? This can be measured as a percent of the field of view as seen through the eyepieces. You can use a stage micrometer or measuring system to determine this. 
Do you feel comfortable with installing image processing boards in to your computer? If you have not had experience doing this, you should work with an expert who has imaging experience or consider purchasing a camera that uses an existing interface (Serial, Parallel, or if newer USB or IEEE 1394).
Do you really need a digital camera? There are still some applications where film is best especially if you require special lighting. Remember that you can post-process a photo image through a high quality scanner to get digital results.
What about the price/performance issue in digital cameras? As with any product, you get what you pay for. A lower price means some sacrifice in performance. These days several really great cameras are priced at around $1000 and provide user resolution or 3.0 - 3.5 Mpixel resolution. Lower priced cameras may lack features, resolution or both. Keep in mind that you will need to spend some money to provide yourself with printing capability that is in line with the resolution that you choose. More on this later.

Now that you have an idea of what your requirements are, make a list of these requirements and use it to select your camera. Stay in line with your list. The following descriptions provide some overview in specific areas to consider when composing your list.

When selecting a camera for microscope imaging, you have the following choices:

Traditional Film Cameras
High Quality Video Cameras
Digital Imaging Cameras
Video Hybrid Cameras

Traditional Film Cameras

Whether standard format, or Polaroid formats, film cameras provide the necessary high resolution and color fidelity required by microscopy. 

There are several drawbacks to film cameras:

  1. You can not preview the image you are saving to film.
  2. Processing film as prints or slides is laborious and expensive even if outside services are used.
  3. The learning curve for successful application of film technology is extensive.
  4. No direct computer interface with a film camera.

Films advantages are:

  1. A high degree of light sensitivity makes film ideal for low light and high speed applications.
  2. Large formats provide unequaled resolution.

High Quality Video Cameras

Over the last 10 years, video has emerged as a highly successful and popular alternative to film. With CCTV high resolution imaging, video provides:

  1. WYSIWYG (What you see is what you get) capability.
  2. Good resolution at high magnification, fine structure characterization.
  3. With 3 chip (RGB) camera technology, excellent color rendition.
  4. Easy digital conversion for computer input, storage and manipulation.
  5. Acceptable sensitivity for low light level applications, marginal sensitivity for high speed applications. 

Video imaging has some down sides also. These include:

  1. Complex microscope interface issues (image magnifiers, adapters, filters, etc.) all of which optekusa.com can address for you.
  2. Often complex computer interface issues (image processing board configurations, software compatibility issues).
  3. Video image display issues (RF interference, color balancing, display resolution, video tape storage resolution, video duplication image loss).

Digital Imaging Cameras

In the past 5 years, video technology has been modified to provide high quality, high resolution digital images. By creating new imaging sensors, and ignoring standard video image standards (NTSC, PAL), it is possible to produce an electronic format camera that has high resolution and a simple direct computer interface with excellent color rendition. 

The advantages of digital imaging are:

  1. High resolution with excellent field of view (where chip format is 1K x 1K pixels, or megapixel quality).
  2. Direct digital mapping of the image into a computer is eliminating the need for image processing boards (but you still may require dedicated image transfer boards).
  3. Relatively high sensitivity and image integration for low light applications such as fluorescence imaging.
  4. Some newer models have near real time image viewing.

Digital image cameras have some disadvantages as well. These may include:

  1. Slow image creation and transfer provide little or no WYSIWYG capability.
  2. Long integration times, often coupled with multiple image captures (for color imaging) make the use of these devices impractical for some high sensitivity fluorescence applications.
  3. Large physical size makes the mounting on some microscopes (especially tissue culture or other "inverted" optics microscopes) cumbersome or impossible.

Video Hybrid Cameras

These cameras feature the ease of use of a video device with a method of taking a digital "snap shot". Most transfer the digital image over an existing computer interface (serial, parallel, or USB). These cameras provide the following benefits:

  1. Complete WYSIWYG functionality at standard video rates (NTSC, PAL).
  2. High Resolution 3-CCD imaging with real time image enhancement rivals the image quality of megapixel imagers.
  3. Direct digital input without image processing boards.
  4. TWAIN compliance for direct input into image manipulation and storage programs.
  5. Excellent sensitivity and cooling provide short duration integration times for low light (fluorescence) requirements.
  6. Small remote-head design provides a simple, low mass microscope mount.

The disadvantages of the hybrid system are few:

  1. Smaller field of view when compared to megapixel digital imagers .
  2. Some color mis-registration when compared to color wheel megapixel imagers.

The table below summarizes the features of each camera type with respect to the nature of the microscope application:

Camera Application Table I: General Issues

Camera Type

Resolution

Sensitivity

How to make digital image

Input to computer

Film

Excellent

Excellent 

Scanner

Scanner

Video

Marginal

Very Good

Image Processor 

Image Processor

Digital

Very good

Very good 

Direct or board

Direct or board

Hybrid

Very Good

Very Good

Direct Parallel

Direct Parallel

Legend

 Excellent

 Able to reproduce image at or beyond the resolution of the microscope.

 Very Good

 Matches the resolution of the microscope without image enhancement.

 Marginal

 Does not match the microscope image in one or more ways.

 Scanner

 A separate hardware device which copies (digitizes) the printed film image to the computer.

 Image processor

 A board-level addition to the computer that converts analog video images to digital images in real or delayed time.

 Direct

 Permits a digital image to be stored in a computer without a scanner or image processor. This method may require a proprietary interface board and/or a parallel or serial cable, or proprietary cable.

 

Camera Application Table II: Low Light Applications

Camera Type

Fluorescence 

WYSIWYG

Color Generation

Film

Seconds to minutes

No

Various emulsions 

Video

Seconds to minutes

Yes

NTSC, PAL

Digital

Seconds to minutes

No

Matrix CCD or Filter

Hybrid

Seconds to minutes

Yes

NTSC, PAL

Microscope imaging is often plagued by uneven and or low illumination levels. Some hybrid cameras provide the easiest-to-use solution for low light imaging. With high speed image integration and Peltier cooling, low light level fluorescence images may be produced in a short time (seconds), with a single exposure, and viewed live before they are stored. With some models 3-CCD true RGB imaging, color correction and digital image enhancement allow the user to control color appearance and image detail in real time.

Digital cameras use color wheels or color matrix CCDs to capture color information in the image. What are the pros and cons of these image acquisition solutions?

Mosaic CCD Digital Imaging Cameras

Most imaging "chip" manufacturers (primarily SONY and Kodak) provide color versions of their megapixel imagers. All of the lower cost digital imaging cameras use this form of sensor. Color is provided by segregating the color information into a mosaic on the chip surface. This is accomplished by taking a group of four pixels and assigning a color frequency to each in a specific pattern. There are usually single red, single blue and two green pixels in the array.

This array is replicated over the entire chip surface to create a color mosaic on to which the incoming color image is mapped. The chip is "read", and the color image is reconstructed using software or dedicated firmware.

What is gained and what is lost in this process? There is an obvious gain in speed. A color image is possible with a single exposure. Single pass color imaging has the further advantage of permitting the frame and focus functions of the camera to be in color as well (often at 12 images/second or more). What is lost is resolution. The color and image definition produced by this method will not tolerate the same degree of enlargement as will an image produced by a multi-pass imaging camera. Color fidelity and registration will not be as good, since each pixel in the array is not exposed to each color of the image. What you get is an "averaged" image, with the resultant compromises.

Color Wheel & Tunable Filter Digital Imaging Cameras

Color wheel and "tunable" filter imaging cameras require multiple subject exposure (at least three; red, green and blue). Often, a fourth exposure is taken, a "dark" exposure, which may be used to subtract image noise from the result. The four exposure method, although longer, generally provides superior image results, with superior color rendition and minimal noise artifact. The down side is that the additive exposure and subsequent image transfer and computer color reconstruction can create a total image acquisition time of a minute or more in cameras with slow data transfer rates and/or long integration times.

In addition, there are distinct differences between color wheel and tunable filter technologies that are significant to long exposure (fluorescent) applications:

  1. Color wheels add a mechanical component to the camera design that increases size and adds an additional service component. The good news is that you maintain the high sensitivity of the imager. The fact that all pixels read the RGB component of the image results in better color fidelity when compared to mosaic imagers.
  2. Tunable filters greatly reduce camera sensitivity (by as much as 60% when compared to color wheel cameras). Reduced sensitivity extends exposure time (more integration: often to critical exposure levels with highly light sensitive fluorescent preparations). The good news is that the camera package can be made smaller and less expensive (there are fewer mechanical and electronic components).
Camera Application Table III: Ease of Use

Camera Type

Time to Acquire Image Time to Store Image Learning Curve

Film

Depends on scanner speed and resolution, generally minutes. Depends on storage program used, generally 5-15 seconds.  Long, knowledge of developing required or use of outside services is required.

Video

1/30 second Up to 1/30 second Depends on image processing components. 

Digital

Seconds to minutes depending on need for color and light level Seconds to minutes depending on camera model  Shorter than film, but similar. Trial and error exposure may require many exposures.

Hybrid

1/30 second Up to 15 seconds  Very short, easiest of all to use.

In summary, I would recommend the following (assuming that your choice meets your application requirements):

  1. If you want the ultimate in ease of use, excellent image quality, "real-time" image display and you are not concerned with a large field of view, I would recommend a hybrid camera.
  2. If you want the ultimate in image quality and format size, go with the multi-pass digital camera.
  3. If you want a large format image, with digital image quality and cost is a factor, go with a mosaic digital imaging camera.
  4. If you need real time, and the ability to record strings of images (motion studies, etc.), go with a high quality hybrid or video camera and an appropriate video digitizing board.
  5. If you have lots of time, and you enjoy darkroom photography, film is still an option

     

 

 
       

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