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What is dark field microscopy?

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

What is dark field microscopy ?-dark field microscopy is a transmitted light technique that uses oblique light to illuminate the sample. Light that does not impinge on the sample is not collected by the objective and results in a dark background. Light that interacts with the sample is scattered (refracted, reflected, and/or diffracted) and is “bent” toward the objective collection angle. This light is collected by the objective and is seen as light spots or areas (resulting from scattered light) on a dark background. Contrast is therefore generated and the sample visualized.dark field microscopy is provided to the sample by a specialized condenser. The simplest DF condenser has a Stop, or Annulus illuminating ring (A). Here, an opaque circle obscures the central portion of the condenser light path. This allows only light in a ring to illuminate the sample. The diameter of the central stop, and thus illuminating annulus, is such that the angle of light is greater than the collecting angle of the objective. Thus without a sample, no light is collected by the objective. This kind of DF stop is useful only for low magnification objectives (<20x).For higher magnification objectives, modifications of the Annular Stop are: B: Immersion paraboloid; C: immersion double mirror concentric; D: cardioid concentric. Gray cone represents the light reflected and refracted from the specimen and collected by the objective. Hatched areas represent glass. Light blocking stops (s) limit light transmission to a hollow cone. i: Immersion oil.; r: reflecting surfaces. (Ruzin,1999).

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Where need the dark field microscopy?

dark field microscopy at High Magnifications-For more precise work and blacker backgrounds, you may choose a condenser designed especially for darkfield, i.e. to transmit only oblique rays. There are several varieties: “dry” darkfield condensers with air between the top of the condenser and the underside of the slide–and immersion darkfield condensers which require the use of a drop of immersion oil (some are designed to use water instead) establishing contact between the top of the condenser and the underside of the specimen slide. The immersion darkfield condenser has internal mirrored surfaces and passes rays of great obliquity and free of chromatic aberration, producing the best results and blackest background.

Perhaps the most widely used darkfield condenser is the paraboloid, consisting of a solid piece of glass ground very accurately into the shape of a paraboloid, as illustrated in Figure 5(b). Light incident upon the reflecting surface (between the glass and condenser housing in Figure 5(b)) of a paraboloid condenser will be focused at the focal point of the reflector. Most paraboloid condensers are cut to ensure that the focal point is slightly beyond the top of the condenser so that parallel light rays will be focused at a position that maximizes illumination of the specimen. The light stop at the bottom of the glass condenser serves to block central rays from reaching the specimen. Light rays that are reflected by the condenser are angled higher than the critical angle of reflection and converge at the principal focus of the condenser. The combination of a glass slide, mounting medium, and immersion oil (between the condenser and the microscope slide) complete the optical homogeneity of the paraboloid shape.

As discussed above, the dry darkfield condenser is useful for objectives with numerical apertures below 0.75 (Figure 5(a)), while the paraboloid and cardioid immersion condensers (Figures 1 and 5(b)) can be used with objectives of very high numerical aperture (up to 1.4). Objectives with a numerical aperture above 1.2 will require some reduction of their working aperture since their maximum numerical aperture may exceed the numerical aperture of the condenser, thus allowing direct light to enter the objective. For this reason, many high numerical aperture objectives designed for use with darkfield as well as brightfield illumination are made with a built-in adjustable iris diaphragm that acts as an aperture stop. This reduction in numerical aperture also limits the resolving power of the objective as well as the intensity of light in the image. Specialized objectives designed exclusively for darkfield work are produced with a maximum numerical aperture close to the lower limit of the numerical aperture of the darkfield condenser. They do not have internal iris diaphragms, however the lens mount diameters are adjusted so at least one internal lens has the optimum diameter to perform as an aperture stop.

Table 2 lists several properties of the most common reflecting high numerical aperture darkfield condensers. This table should be used as a guide when selecting condenser/objective combinations for use with high numerical aperture darkfield applications.

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

dark field microscopy blood analysis

What is Blood and What Does it Do-Two types of blood vessels carry blood throughout our bodies: The arteries carry oxygenated blood (blood that has received oxygen from the lungs) from the heart to the rest of the body.

The blood then travels through the veins back to the heart and lungs, where it receives more oxygen. As the heart beats, you can feel blood traveling through the body at your pulse points – like the neck and the wrist – where large, blood-filled arteries run close to the surface of the skin.

The blood that flows through this network of veins and arteries is called whole blood. Whole blood contains three types of blood cells:

Red Blood Cells
White Blood Cells
Platelets

These blood cells are mostly manufactured in the bone marrow (the soft tissue inside our bones), especially in the bone marrow of the vertebrae (the bones that make up the spine), ribs, pelvis, skull, and sternum (breastbone). These cells travel through the circulatory system suspended in a yellowish fluid called plasma (pronounced: plaz-muh). Plasma is 90% water and contains nutrients, proteins, hormones, and waste products. Whole blood is a mixture of blood cells and plasma.
Red Blood Cells

Red blood cells (RBCs, and also called erythrocytes, pronounced: ih-rith-ruh-sytes) are shaped like slightly indented, flattened disks. Red blood cells contain an iron-rich protein called hemoglobin (pronounced: hee-muh-glow-bun). Blood gets its bright red color when the hemoglobin in RBCs picks up oxygen in the lungs. As the blood travels through the body, the hemoglobin releases oxygen to the tissues. The body contains more RBCs than any other type of cell, and each has a life span of about 4 months. Each day, the body produces new RBCs to replace those that die or are lost from the body.

White Blood Cells

White blood cells (WBCs, and also called leukocytes, pronounced: loo-kuh-sytes) are a key part of the body’s system for defending itself against infection. They can move in and out of the bloodstream to reach affected tissues. The blood contains far fewer white blood cells than red cells, although the body can increase production of WBCs to fight infection. There are several types of white blood cells, and their life spans vary from a few days to months. New cells are constantly being formed in the bone marrow.

Several different parts of blood are involved in fighting infection. White blood cells called granulocytes (pronounced: gran-yuh-low-sytes) and lymphocytes (pronounced: lim-fuh-sytes) travel along the walls of blood vessels. They fight germs such as bacteria and viruses and may also attempt to destroy cells that have become infected or have changed into cancer cells.

Certain types of WBCs produce antibodies, special proteins that recognize foreign materials and help the body destroy or neutralize them. Someone with an infection will often have a higher white cell count than when he or she is well because more WBCs are being produced or are entering the bloodstream to battle the infection. After the body has been challenged by some infections, lymphocytes “remember” how to make the specific antibodies that will quickly attack the same germ if it enters the body again.
Platelets

Platelets (also called thrombocytes, pronounced: throm-buh-sytes) are tiny oval-shaped cells made in the bone marrow. They help in the clotting process. When a blood vessel breaks, platelets gather in the area and help seal off the leak. Platelets survive only about 9 days in the bloodstream and are constantly being replaced by new cells.

Drop of Blood

Blood also contains important proteins called clotting factors, which are critical to the clotting process. Although platelets alone can plug small blood vessel leaks and temporarily stop or slow bleeding, the action of clotting factors is needed to produce a strong, stable clot.

Platelets and clotting factors work together to form solid lumps to seal leaks, wounds, cuts, and scratches and to prevent bleeding inside and on the surfaces of our bodies. The process of clotting is like a puzzle with interlocking parts. When the last part is in place, the clot happens – but if only one piece is missing, the final pieces can’t come together.

When large blood vessels are severed (or cut), the body may not be able to repair itself through clotting alone. In these cases, dressings or stitches are used to help control bleeding.

In addition to the cells and clotting factors, blood contains other important substances, such as nutrients from the food that has been processed by the digestive system. Blood also carries hormones released by the endocrine glands and carries them to the body parts that need them.

What you can expect from Dark Field Blood Analysis-Thus, the dark field blood analysis provides information about the function and structure of blood cells and plasma endobionts, as well as developing bacterial and fungal precursors. The dark field examination also indicates changes within the cell through hormonal and mineral deficiencies. It is particularly beneficial for the evaluation of patients with chronic diseases, and children with susceptibility to infection, or recurrent bacterial problems, such as Candida or other fungal diseases. Dark field blood analysis is crucial in answering questions related to any chronic, or toxic problems.

Dark field microscopy is also an important tool in biological therapies. It can be used to test the effect of certain medications, by adding the medication to a blood sample and analyzing the reaction that is produced. This investigation is extremely motivating for the patient, by allowing him to directly experience the diagnosis.

The effects of dark field microscopy can not be replaced by any other blood test, especially not by normal laboratory microscopic blood tests, sent in fixed samples, as the blood changes its function due to environmental changes, so the blood must be tested while it is still fresh. It is also important to evaluate the degeneration tendency of blood samples, especially when considering the behavior of tumors.

The presence of bacteria precursors, which are not disease-inducing, but increase the risk of future disease development, can also be found in the dark field examination. Therefore, this dark field examination of the blood is a valuable and necessary preventative measure.

 

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