A photograph taken through a compound microscope is also known as a photomicrograph. Some call it photomicrography, others microphotography, or you may call it photography through the microscope what matters is the normal daylight or flash photography to which many are more familiar. In photomicrography, light sources are plays an important role for color balance. Light microscopy relies on the bending, or refraction, of light rays. Light rays pass straight through the center of a curved lens. The farther they are from the center, the more they bend. The illumination is decreased in contrast to daylight or flash, so reciprocity failure of film is an issue. This is true especially in the case of fluorescence photography in which the backdrop is nearly black. If you wish to observe a living cell, it must be small or thin enough for light to pass through. Structures in the side cells can be seen only if they differ in color and density from their surroundings-but most are nearly colorless and optically uniform in density.
Setting up microscope illumination to get the best image possible to capture a good photomicrograph, photographic principles should be considered carefully. In making sure, that illumination is set up properly it can be possibly done through transmission of light images. If there is not enough detail in an image, to get the best resolution, a higher numerical aperture lens should be considered. To decrease glare as well as bring out the fine feature of an image, the intermediate plane aperture should be stopped down. With fluorescence microscopy, one should use the maximum numerical aperture available for the magnification desired, especially if the staining is good. A better numerical aperture will draw in more illumination to the lens. It is highly encourage that exposures should be bracketed. Bracketing the exposures will improve the possibility to get at least one very good exposure of a particular image. Two main good reason for bracketing. First, it is not easy as normal daylight or flash photography is a photomicrography. Here the exposure period is usually longer reciprocity failure of films should be greatly considered. Next, to come back again and photograph an image taken in too light or dark exposure is a waste of time and energy. Time is more precious than using a couple of rolls of film. In doing fluorescence, it is very possible that your image taken will have faded and be that good enough like the first time it was photographed.
What is reciprocity failure and why should it be given an importance. Photographic films have a feature in which many microscopists do not like. It is because the more that they are exposed to light they lose its ability to absorb light. This is what is called reciprocity failure. It happens to a very common available photographic film for exposure period, more than eight of a second. With microscopes, exposure time often exceeds the normal time. Reciprocity failure can in most cases be resolved by adding the length of the exposure more than what an exposure meter normally would indicate for a given exposure period.
For fluorescence photomicrography reciprocity failure becomes even more important because of the longer period of exposures required. Reciprocity failure differs with the kind of film tested through trials or details should be taken from the film manufacturer itself.
For microscope mounted with camera, for fluorescence microscopy, it is best to do bracketed manual timed exposures.
For normal daylight exposure meters in those cameras to obtain a useful low light exposure reading, light levels should be too low. For microscope photographic systems, which have no reciprocity failure possible remedy, it is probably best to just to use a film with low reciprocity failure and to bracket exposures.
