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Flash Photomicrography.
Electronic Flash as a Light Source for  
Photography through the Microscope. 
5 of 7
Introduction Simple Setup Köhler lamp Lamp Optics Exposure Camera Support Fibre Optics

Determining Exposure in Flash Photomicrography.

The Ramsden Circle.

The diagram below introduces the concept of the Ramsden circle, and is a key to understanding the problems of film exposure in light micrography.
The Ramsden circle, also called the exit-pupil of the microscope, is a circular area through which pass all the image-forming rays leaving the microscope. It is itself an image of the back-lens of the microscope objective, demagnified in proportion to the power of the eyepiece, and is usually located a few millimetres above the eye lens of the eyepiece.

Diagram showing Ramsden circle.
Formation of the Ramsden circle in a positive eyepiece.

In spite of the fact that microscope objectives work at extremely high apertures (very small f number) the image they project is highly magnified, and the intensity of the image is reduced by a factor proportional to the square of the magnification.
If magnification is doubled, the intensity of the resultant image is one quarter that of the original image. This is because at twice the magnification, for any given lens, the same amount of image-forming light now has to cover four times the original area.

In the case of an oil-immersion objective of x100, the magnification of the primary image at the plane of the eyepiece diaphragm is x100, and the intensity of the magnified image is therefore one ten thousandth of the illumination intensity at the plane of the specimen.
The primary image is further magnified by the eyepiece, which projects it (with or without a focus achromat) an appropriate distance to fill the film gate of the camera in use. Since this distance is also subject to the inverse square relationship between projection distance and image intensity, it is not surprising that it takes a much longer time to expose a 10" x 8" photomicrographic plate than a frame of 16mm film.

  f Number and the Microscope.

The familiar concept of f number is as useful in this context as it is in routine photography. From the point of view of the film, all of the image-forming rays are emanating from a small circle in a plane lying a few millimetres above the eyepiece -- the Ramsden circle. The diameter of this circle, together with its distance from the image on the film, provides the figures required to calculate the f number of the microscope with respect to the film format in use.
As in normal photography:

f number = distance between lens aperture and image ÷ aperture diameter

In the case of the x100 OI objective and 35mm film, the Ramsden circle will have a diameter of 1mm or so (depending on the eyepiece in use), and the projection distance to the film (with a 125mm focus achromat placed at the Ramsden circle) is 125mm . This gives an f number for the system of around f 125.
This will at once explain to those familiar with normal photography why intense illuminants are required to give reasonably short exposure times in photomicrography.

  f Number and Exposure.

This calculation for the f number of the system is of practical use for exposure determination in the simple case of a low-power photomicrographic system used to image objects illuminated by incident flash. An example of such a system is illustrated in the article on photographing feeding mosquitoes.

If a photographic flash meter is placed at the point where the specimen will be, and the flash units fired to give a reading, adequate exposure of the film is assured if the meter gives a reading equal to the f number calculated for the microscope.
The intensity of the illumination can be varied by varying the distance of the flash units from the meter. Some allowance should also be made for light losses due to reflection and absorption in the optics of the imaging system. In the example system, this is in the region of half a photographic stop.

Other methods must be used for transmitted light setups including brightfield, darkfield, phase contrast, Nomarski etc. Notes on determining flash exposure for these systems will be added soon.

Watson's Laboratory Camera.
Watson's Laboratory Camera. c. 1910.

In the meantime, just as an exercise, calculate the approximate f number for this photomicrographic setup of almost a hundred years ago.

The next section deals with ways of supporting the camera.