Darkfield Microscopy Imaging Techniques

Darkfield microscopy imaging techniques provide a method to view features in blood not normally shown with phase contrast and brightfield illumination. This method illuminates the specimen from the side, not from behind, so the background is black. Minute features of the blood are shown in a glowing light as it reflects off their surfaces.

In live blood cell analysis, darkfield microscopy is best used to understand and view the cause of improper nutrition and problems in the body. Some evaluations are done solely using phase contrast or even using only brightfield, but they are missing a whole spectrum of enlightning information that can only be viewed in true darkfield.

Light enters the microscope for illumination of the sample. The condenser lens focuses the light towards the sample. A specially sized disc, the patch stop blocks some light from the sample, leaving an outer ring of illumination. The light enters the sample. Most is directly transmitted, while some is scattered from the sample. The scattered light enters the objective lens, while the directly transmitted light simply misses the lens and is not collected. Only the scattered light goes on to produce the image, while the directly transmitted light is omitted.

Dark field microscopy is a very simple yet effective technique and well suited for uses involving live and unstained biological samples, such as a smear from a tissue culture or individual water borne organisms with single cell. Considering the simplicity of the setup, the quality of images obtained from this technique is impressive.

The main limitation of dark field microscopy is the low light levels seen in the final image. This means the sample must be very strongly illuminated, and can cause damage to the sample.

To view a specimen in dark field, an opaque disc is placed underneath the condenser lens, so that only light that is scattered by objects on the slide can reach the eye. Instead of coming up through the specimen, the light is reflected by particles on the slide. Everything is visible regardless of color, usually bright white against a dark background.

Pigmented objects are often seen in false colors, that is, the reflected light is of a color different than the color of the object. Better resolution can be obtained using dark as opposed to bright field viewing.

Sophisticated equipment is not necessary to get a dark field effect, but you do need a higher intensity light, since you are seeing only reflected light. At low magnification any decent optical instrument can be set up so that light is reflected toward the viewer rather than passing through the object directly toward the viewer.

Dark field microscopy techniques are normally used by scientists to, due to the simplicity of the setup, are almost entirely free of artifacts. However the interpretation of dark field images must be done with care as common dark features of bright field microscopy images may be invisible, and vice versa.

Dark Field Microscopy thus allows a health professional to evaluate the shapes and other properties of individual blood cells, indicating nutritional conditions which can be adversely affecting a health. The advantage of this analysis over standard blood tests, which detect chemical changes in the blood, is the ability of dark field microscopy to detect nutritional disorders sooner, when the problem is in its infancy stages. By monitoring the condition of the blood, a health professional can assist in balancing the blood by giving dietary and lifestyle recommendations which can enhance health.

In darkfield microscopy, one is therefore able to observe live blood. Unlike the techniques of electron microscopy, no fixative is used so the picture is one of mobility rather than fixity. With stains and fixatives, the picture reveals a moment in time rather than a continuum. What one sees in the mobile situation are the usual red blood cells, white blood cells, plasma and what is floating in the plasma. Microbial activity, undigested food, fungi, and crystals are all apparent as is the capacity of the red blood cells to circulate and the white blood cells to devour morbid matter.

As we know, red blood cells transport oxygen to the tissues of the body. Without oxygen, we are devitalized, and according to some theories, the tissues go into a morbid state in which they can survive on fermentation rather than oxygenation. This is what is referred to as anaerobic and it is believed, by such persons as Nobel laureate Prof. Otto Warburg, that cancer thrives in such oxygen deficient conditions.

With darkfield microscopy, one often sees a condition called rouleau in which the red blood cells are stacked together as shown below. Some people believe it is because of the stress on the body of poor metabolism and others believe it is due to this as well as pH or acid and alkaline balance, wrong dietary choices or the presence of excessively high levels of free radicals. In any event, it is usually correctable.

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