The other main kind of eyepiece is the positive eyepiece with a diaphragm below its lenses, commonly known as the Ramsden eyepiece, as illustrated in Figure 2 (on the left). This eyepiece has an eye lens and field lens that are also plano-convex, but the field lens is mounted with the curved surface facing towards the eye lens. The front focal plane of this eyepiece lies just below the field lens, at the level of the eyepiece diaphragm, making this eyepiece readily adaptable for mounting reticles. To provide better correction, the two lenses of the Ramsden eyepiece may be cemented together.

where λ is the wavelength of illumination, n is the refractive index of the imaging medium, NA is the objective numerical aperture, M is the objective lateral magnification, and e is the smallest distance that can be resolved by a detector that is placed in the image plane of the objective. Notice that the diffraction-limited depth of field (the first term on the right-hand side of the equation) shrinks inversely with the square of the numerical aperture, while the lateral limit of resolution is reduced with the first power of the numerical aperture. The result is that axial resolution and the thickness of optical sections are affected by the system numerical aperture much more than is the lateral resolution of the microscope (see Table 2).

The "range of useful magnification" for an objective/eyepiece combination is defined by the numerical aperture of the system. There is a minimum magnification necessary for the detail present in an image to be resolved, and this value is usually rather arbitrarily set as 500 times the numerical aperture (500 x NA). At the other end of the spectrum, the maximum useful magnification of an image is usually set at 1000 times the numerical aperture (1000 x NA). Magnifications higher than this value will yield no further useful information or finer resolution of image detail, and will usually lead to image degradation. Exceeding the limit of useful magnification causes the image to suffer from the phenomenon of "empty magnification", where increasing magnification through the eyepiece or intermediate tube lens only causes the image to become more magnified with no corresponding increase in detail resolution. Table 3 lists the common objective/eyepiece combinations that lie in the range of useful magnification.

The properties of several common commercially available eyepieces (manufactured by Olympus America, Inc.) are listed according to type in Table 1. The three major types of eyepieces listed in Table 1 are Finder, Wide Field, and Super Widefield. The terminology used by various manufacturers can be very confusing and careful attention should be paid to their sales brochures and microscope manuals to ensure that the correct eyepieces are being used with a specific objective. In Table 1, the abbreviations that designate wide field and super widefield eyepieces are coupled to their correction for high eye point, and are WH and SWH, respectively. The magnifications are either 10x or 15x and the Field Numbers (discussed below) range from 14 to 26.5, depending upon the application. The diopter adjustment is approximately the same for all eyepieces and many also contain either a photomask or micrometer reticle.

The simplest negative eyepiece design, often termed the Huygenian eye-piece (illustrated in Figure 2), is found on most teaching and laboratory microscopes fitted with achromatic objectives. Although the Huygenian eye and field lenses are not well corrected, their aberrations tend to cancel each other out. More highly corrected negative eyepieces have two or three lens elements cemented and combined together to make the eye lens. If an unknown eyepiece carries only the magnification inscribed on the housing, it is most likely to be a Huygenian eyepiece, best suited for use with achromatic objectives of 5x-40x magnification.

where R is the separation distance, λ is the illumination wavelength, n is the imaging medium refractive index, and θ is one-half of the objective angular aperture. In examining the equation, it becomes apparent that resolution is directly proportional to the illumination wavelength. The human eye responds to the wavelength region between 400 and 700 nanometers, which represents the visible light spectrum that is utilized for a majority of microscope observations. Resolution is also dependent upon the refractive index of the imaging medium and the objective angular aperture. Objectives are designed to image specimens either with air or a medium of higher refractive index between the front lens and the specimen. The field of view is often quite limited, and the front lens element of the objective is placed close to the specimen with which it must lie in optical contact. A gain in resolution by a factor of approximately 1.5 is attained when immersion oil is substituted for air as the imaging medium.

Three critical design characteristics of the objective set the ultimate resolution limit of the microscope. These include the wavelength of light used to illuminate the specimen, the angular aperture of the light cone captured by the objective, and the refractive index in the object space between the objective front lens and the specimen.

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For many years, objective lenses designed for biological applications from most manufacturers all conformed to an international standard of parfocal distance. Thus, a majority of objectives had a parfocal distance of 45.0 millimeters and were considered interchangeable. With the migration to infinity-corrected tube lengths, a new set of design criteria emerged to correct for aberrations in the objective and tube lenses. Coupled to an increased demand for greater flexibility to accommodate the need for ever-greater working distances with higher numerical apertures and field sizes, interchangeability between objective lenses from different manufacturers disappeared. This transition is exemplified by the modern Nikon CFI-60 optical system that features "Chrome Free" objectives, tube lenses, and eyepieces. Each component in the CFI-60 system is separately corrected without one being utilized to achieve correction for another. The tube length is set to infinity (parallel light path) using a tube lens, and the parfocal distance has been increased to 60 millimeters. Even the objective mounting thread size has been altered from 20.32 to 25 millimeters to meet new requirements of the optical system.

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What iseyepiecein microscope

Manufacturers often produce specialized eyepieces, often termed photo eyepieces, that are designed to be used with photomicrography. These eyepieces are usually negative (Huygenian type) and are not capable of being used visually. For this reason, they are typically called projection lenses. A typical projection lens is illustrated in Figure 7 below.

When a manufacturer's set of matched objectives, e.g. all achromatic objectives of various magnifications (a single subset of the objectives listed in Table 1), are mounted on the nosepiece, they are usually designed to project an image to approximately the same plane in the body tube. Thus, changing objectives by rotating the nosepiece usually requires only minimal use of the fine adjustment knob to re-establish sharp focus. Such a set of objectives is described as being parfocal, a useful convenience and safety feature. Matched sets of objectives are also designed to be parcentric, so that a specimen centered in the field of view for one objective remains centered when the nosepiece is rotated to bring another objective into use.

Eyepiecelensfunction

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Modern microscopes feature vastly improved plan-corrected objectives in which the primary image has much less curvature of field than older objectives. In addition, most microscopes now feature much wider body tubes that have greatly increased the size of intermediate images. To address these new features, manufacturers now produce wide-eyefield eyepieces (illustrated in Figure 1) that increase the viewable area of the specimen by as much as 40 percent. Because the strategies of eyepiece-objective correction techniques vary from manufacturer to manufacturer, it is very important (as stated above) to use only eyepieces recommended by a specific manufacturer for use with their objectives.

Diopter adjustmentfunction

There are two major types of eyepieces that are grouped according to lens and diaphragm arrangement: the negative eyepieces with an internal diaphragm and positive eyepieces that have a diaphragm below the lenses of the eyepiece. Negative eyepieces have two lenses: the upper lens, which is closest to the observer's eye, is called the eye-lens and the lower lens (beneath the diaphragm) is often termed the field lens. In their simplest form, both lenses are plano-convex, with convex sides "facing" the specimen. Approximately mid-way between these lenses there is a fixed circular opening or internal diaphragm which, by its size, defines the circular field of view that is observed in looking into the microscope.

The focal length of a lens system is defined as the distance from the lens center to a point where parallel rays are focused on the optical axis (often termed the principal focal point). An imaginary plane perpendicular to the principal focal point is called the focal plane of the lens system. Every lens has two principal focal points for light entering each side, one in front and one at the rear. By convention, the objective focal plane that is nearer to the front lens element is known as the front focal plane and the focal plane located behind the objective is termed the rear focal plane (see Figure 4). The actual position of the rear focal plane varies with objective construction, but is generally situated somewhere inside the objective barrel for high magnification objectives. Objectives of lower magnification often have a rear focal plane that is exterior to the barrel, located in the thread area or within the microscope nosepiece.

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Just as the brightness of illumination in a microscope is governed by the square of the working numerical aperture of the condenser, the brightness of an image produced by the objective is determined by the square of its numerical aperture. In addition, objective magnification also plays a role in determining image brightness, which is inversely proportional to the square of the lateral magnification. The square of the numerical aperture/magnification ratio expresses the light-gathering power of the objective when utilized with transmitted illumination. Because high numerical aperture objectives are often better corrected for aberration, they also collect more light and produce a brighter, more corrected image that is highly resolved. It should be noted that image brightness decreases rapidly as the magnification increases. In cases where the light level is a limiting factor, choose an objective with the highest numerical aperture, but having the lowest magnification factor capable of producing adequate resolution.

More advanced eyepiece designs resulted in the Periplan eyepiece that is illustrated in Figure 4 above. This eyepiece contains seven lens elements that are cemented into a single doublet, a single triplet, and two individual lenses. Design improvements in periplan eyepieces lead to better correction for residual lateral chromatic aberration, increased flatness of field, and a general overall better performance when used with higher power objectives.

Illustrated in Figure 3 is a schematic drawing of light waves reflecting and/or passing through a lens element coated with two antireflection layers. The incident wave strikes the first layer (Layer A in Figure 3) at an angle, resulting in part of the light being reflected (R(o)) and part being transmitted through the first layer. Upon encountering the second antireflection layer (Layer B), another portion of the light is reflected at the same angle and interferes with light reflected from the first layer. Some of the remaining light waves continue on to the glass surface where they are again both reflected and transmitted. Light reflected from the glass surface interferes (both constructively and destructively) with light reflected from the antireflection layers. The refractive indices of the antireflection layers vary from that of the glass and the surrounding medium (air). As the light waves pass through the antireflection layers and glass surface, a majority of the light (depending upon the incident angle--usual normal to the lens in optical microscopy) is ultimately transmitted through the glass and focused to form an image.

Eyepiecediagram

Light rays emanating from the eyepiece intersect at the exit pupil or eye point, often referred to as the Ramsden disc, where the pupil of the microscopists eye should be placed in order for her to see the entire field of view (usually 8-10 mm from the eye lens). By increasing the magnification of the eyepiece, the eye point is drawn closer to the upper surface of the eye lens, making it much more difficult for the microscopist to use, especially if they are wearing eyeglasses. To compensate for this, specially designed high eye point eyepieces have been manufactured that feature eye point distances approaching 20-25 mm above the surface of the eye lens. These improved eyepieces have larger diameter eye lenses that contain more optical elements and usually feature improved flatness of field. Such eyepieces are often designated with the inscription "H" somewhere on the eyepiece housing, either alone or in combination with other abbreviations, as discussed above. We should mention that high eye point eyepieces are especially useful for microscopists who wear eyeglasses to correct for near or far sightedness, but they do not correct for several other visual defects, such as astigmatism. Today, high eye point eyepieces are very popular, even with people who do not wear eyeglasses, because the large eye clearance reduces fatigue and makes viewing images through the microscope much more pleasurable.

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During assembly of the objective, lenses are first strategically spaced and lap-seated into cell mounts, then packaged into a central sleeve cylinder that is mounted internally within the objective barrel. Individual lenses are seated against a brass shoulder mount with the lens spinning in a precise lathe chuck, followed by burnishing with a thin rim of metal that locks the lens (or lens group) into place. Spherical aberration is corrected by selecting the optimum set of spacers to fit between the lower two lens mounts (the hemispherical and meniscus lens). The objective is parfocalized by translating the entire lens cluster upward or downward within the sleeve with locking nuts so that objectives housed on a multiple nosepiece can be interchanged without losing focus. Adjustment for coma is accomplished with three centering screws that can optimize the position of internal lens groups with respect to the optical axis of the objective.

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The clearance distance between the closest surface of the cover glass and the objective front lens is termed the working distance. In situations where the specimen is designed to be imaged without a cover glass, the working distance is measured at the actual surface of the specimen. Generally, working distance decreases in a series of matched objectives as the magnification and numerical aperture increase (see Table 1). Objectives intended to view specimens with air as the imaging medium should have working distances as long as possible, provided that numerical aperture requirements are satisfied. Immersion objectives, on the other hand, should have shallower working distances in order to contain the immersion liquid between the front lens and the specimen. Many objectives designed with close working distances have a spring-loaded retraction stopper that allows the front lens assembly to be retracted by pushing it into the objective body and twisting to lock it into place. Such an accessory is convenient when the objective is rotated in the nosepiece so it will not drag immersion oil across the surface of a clean slide. Twisting the retraction stopper in the opposite direction releases the lens assembly for use. In some applications (see below), a long free working distance is indispensable, and special objectives are designed for such use despite the difficulty involved in achieving large numerical apertures and the necessary degree of optical correction.

Some eyepieces have a movable "pointer" located within the eyepiece and positioned so that it appears as a silhouette in the image plane. This pointer is useful when indicating certain features of a specimen, especially when a microscopist is teaching students about specific features. Most eyepiece pointers can be rotated in a 360 degree angle around the specimen and more advanced versions can translate across the viewfield.

Function of eyepiecein microscope

Best results in microscopy require that objectives be used in combination with eyepieces that are appropriate to the correction and type of objective. The basic anatomy of a typical modern eyepiece is illustrated in Figure 1. Inscriptions on the side of the eyepiece describe its particular characteristics and function.

Care should be taken in choosing eyepiece/objective combinations to ensure the optimal magnification of specimen detail without adding unnecessary artifacts. For instance, to achieve a magnification of 250x, the microscopist could choose a 25x eyepiece coupled to a 10x objective. An alternative choice for the same magnification would be a 10x eyepiece with a 25x objective. Because the 25x objective has a higher numerical aperture (approximately 0.65) than does the 10x objective (approximately 0.25), and considering that numerical aperture values define an objective's resolution, it is clear that the latter choice would be the best. If photomicrographs of the same viewfield were made with each objective/eyepiece combination described above, it would be obvious that the 10x eyepiece/25x objective duo would produce photomicrographs that excelled in specimen detail and clarity when compared to the alternative combination.

The field diameter in an optical microscope is expressed by the field-of-view number or simply field number, which is the diameter of the viewfield expressed in millimeters and measured at the intermediate image plane. The field diameter in the object (specimen) plane becomes the field number divided by the magnification of the objective. Although the field number is often limited by the magnification and diameter of the ocular (eyepiece) field diaphragm, there is clearly a limit that is also imposed by the design of the objective. In early microscope objectives, the maximum usable field diameter was limited to about 18 millimeters (or considerably less for high magnification eyepieces), but modern plan apochromats and other specialized flat-field objectives often have a usable field that can range between 22 and 28 millimeters or more when combined with wide-field eyepieces. Unfortunately, the maximum useful field number is not generally engraved on the objective barrel and is also not commonly listed in microscope catalogs.

Compensating eyepieces play a crucial role in helping to eliminate residual chromatic aberrations inherent in the design of highly corrected objectives. Hence, it is preferable that the microscopist uses the compensating eyepieces designed by a particular manufacturer to accompany that manufacturer's higher-corrected objectives. Use of an incorrect eyepiece with an apochromatic objective designed for a finite (160 or 170 millimeter) tube length application results in dramatically increased contrast with red fringes on the outer diameters and blue fringes on the inner diameters of specimen detail. Additional problems arise from a limited flatness of the viewfield in simple eyepieces, even those corrected with eye-lens doublets.

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The axial range through which an objective can be focused without any appreciable change in image sharpness is referred to as the depth of field. This value varies radically from low to high numerical aperture objectives, usually decreasing with increasing numerical aperture (see Table 2 and Figure 2). At high numerical apertures, the depth of field is determined primarily by wave optics, while at lower numerical apertures, the geometrical optical "circle of confusion" dominates. The total depth of field is given by the sum of the wave and geometrical optical depths of field as:

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A modified version of the Ramsden eyepiece is known as the Kellner eyepiece, as illustrated on the left in Figure 3. These improved eyepieces contain a doublet of eye-lens elements cemented together and feature a higher eye point than either the Ramsden or Huygenian eyepiece as well as a much larger field of view. A modified version of the simple Huygenian eyepiece is also illustrated in Figure 3, on the right. While these modified eyepieces perform better than their simple one-lens counterparts, they are still only useful with low-power achromat objectives.

Table 1 lists working distance and numerical aperture as a function of magnification for the four most common classes of objectives: achromats, plan achromats, plan fluorites, and plan apochromats. Note that dry objectives all have a numerical aperture value of less than 1.0 and only objectives designed for liquid immersion media have a numerical aperture that exceeds this value.

Eyepieces can be adapted for measurement purposes by adding a small circular disk-shaped glass reticle (sometimes referred to as a graticule or reticule) at the plane of the field diaphragm of the eyepiece. Reticles usually have markings, such as a measuring rule or grid, etched onto the surface. Because the reticle lies in the same plane as the field diaphragm, it appears in sharp focus superimposed over the image of the specimen. Eyepieces using reticles must contain a focusing mechanism (usually a helical screw or slider) that allows the image of the reticle to be brought into focus. Several typical reticles are illustrated in Figure 5 below.

Camera systems have become an integral part of the microscope and most manufacturers provide photomicrographic attachment cameras as an optional accessory. These advanced camera systems often feature motorized black boxes that store and automatically step through film frame-by-frame as photomicrographs are taken. A common feature of these integral camera systems is a beamsplitter focusing telescopic eyepiece (see Figure 8) that allows the microscopist to view, focus, and frame samples for photomicrography. This telescope contains a photomicrography reticle, similar to the one illustrated in Figure 5(a) that is inscribed with a rectangular element that circumscribes the area captured with 35 mm film, and also corner brackets for larger format films. For convenience in scanning and photographing samples, the microscopist can adjust the telescopic eyepiece so that it is parfocal with the ocular eyepieces to make it easier to frame and take photomicrographs.

At one time, eyepieces were available in a wide spectrum of magnifications ranging from 6.3x to 25x and sometimes even higher for special applications. These eyepieces are very useful for observation and photomicrography with low-power objectives. Unfortunately, with higher power objectives, the problem of empty magnification becomes important when using very high magnification eyepieces and these should be avoided. Today most manufacturers restrict their eyepiece offerings to those in the 10x to 20x range. The diameter of the viewfield in an eyepiece is expressed as a "field-of-view number" or field number (FN), as discussed above. Information about the field number of an eyepiece can yield the real diameter of the object viewfield using the formula:

Projection lenses must be carefully corrected so that they will produce flat-field images, a definite "must" for accurate photomicrography. They are generally also color-corrected to ensure true reproduction of color in color photomicrography. Magnification factors in photomicrography projection lenses range from 1x to about 5x, and these can be interchanged to adjust the size of the final image in the photomicrograph.

Arm microscopefunction

As light rays pass through an objective, they are restricted by the rear aperture or exit pupil of the objective, as illustrated in Figure 4. The diameter of this aperture varies between 12 millimeters for low magnification objectives down to around 5 millimeters for the highest power apochromatic objectives. Aperture size is extremely critical for epi-illumination applications that rely on the objective to act as both an imaging system and condenser, where the exit pupil also becomes an entrance pupil. The image of the light source must completely fill the objective rear aperture to produce even illumination across the viewfield. If the light source image is smaller than the aperture, the viewfield will experience vignetting from uneven illumination. On the other hand, if the light source image is larger than the rear aperture, some light does not enter the objective and the intensity of illumination is reduced.

is known as the numerical aperture (abbreviated NA), and provides a convenient indicator of the resolution for any particular objective. Numerical aperture is generally the most important design criteria (other than magnification) to consider when selecting a microscope objective. Values range from 0.1 for very low magnification objectives (1x to 4x) to as much as 1.6 for high-performance objectives utilizing specialized immersion oils. As numerical aperture values increase for a series of objectives of the same magnification, we generally observe a greater light-gathering ability and increase in resolution. The microscopist should carefully choose the numerical aperture of an objective to match the magnification produced in the final image. Under the best circumstances, detail that is just resolved should be enlarged sufficiently to be viewed with comfort, but not to the point that empty magnification hampers observation of fine specimen detail.

World-class Nikon objectives, including renowned CFI60 infinity optics, deliver brilliant images of breathtaking sharpness and clarity, from ultra-low to the highest magnifications.

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Simple eyepieces such as the Huygenian and Ramsden and their achromatized counterparts will not correct for residual chromatic difference of magnification in the intermediate image, especially when used in combination with high magnification achromatic objectives as well as any fluorite or apochromatic objectives. To remedy this, manufacturers produce compensating eyepieces that introduce an equal, but opposite, chromatic error in the lens elements. Compensating eyepieces may be either of the positive or negative type, and must be used at all magnifications with fluorite, apochromatic and all variations of plan objectives (they can also be used to advantage with achromatic objectives of 40x and higher). In recent years, modern microscope objectives have their correction for chromatic difference of magnification either built into the objectives themselves (Olympus and Nikon) or corrected in the tube lens (Leica and Zeiss).

Describe thefunction ofthe mirror

Our recommendation is to carefully choose the objective first, then purchase an eyepiece that is designed to work in conjunction with the objective. When choosing eyepieces, it is relatively easy to differentiate between simple and more highly compensated eyepieces. Simple eyepieces such as the Ramsden and Huygenian (and their more highly corrected counterparts) will appear to have a blue ring around the edge of the eyepiece diaphragm when viewed through the microscope or held up to a light source. In contrast, more highly corrected compensating eyepieces with have a yellow-red-orange ring around the diaphragm under the same circumstances.

The last, but perhaps most important, factor in determining the resolution of an objective is the angular aperture, which has a practical upper limit of about 72 degrees (with a sine value of 0.95). When combined with refractive index, the product:

The eyepieces illustrated in Figure 1 are inscribed with UW, which is an abbreviation for the Ultra Wide viewfield. Often eyepieces will also have an H designation, depending upon the manufacturer, to indicate a high eye point focal plane that allows microscopists to wear glasses while viewing samples. Other inscriptions often found on eyepieces include WF for Wide-Field; UWF for Ultra Wide-Field; SW and SWF for Super Wide-Field; HE for High Eye point; and CF for eyepieces intended for use with CF corrected objectives. Compensating eyepieces are often inscribed with K, C, or comp as well as the magnification. Eyepieces used with flat-field objectives are sometimes labeled Plan-Comp. The eyepiece magnification of the eyepieces in Figure 1 is 10x (indicated on the housing), and the inscription A/24 indicates the field number is 24, which refers to the diameter (in millimeters) of the fixed diaphragm in the eyepiece. These eyepieces also have a focus adjustment and a thumbscrew that allows their position to be fixed. Manufactures now often produce eyepieces having rubber eye-cups that serve both to position the eyes the proper distance from the front lens, and to block room light from reflecting off the lens surface and interfering with the view.

Eyepieces work in combination with microscope objectives to further magnify the intermediate image so that specimen details can be observed. Oculars is an alternative name for eyepieces that has been widely used in the literature, but to maintain consistency during this discussion we will refer to all oculars as eyepieces.

Function ofbody tube in microscope

Resolution for a diffraction-limited optical microscope can be described as the minimum detectable distance between two closely spaced specimen points:

Presented in Figure 1 is a cut-away diagram of a microscope objective being illuminated by a simple two-lens Abbe condenser. Light passing through the condenser is organized into a cone of illumination that emanates onto the specimen and is then transmitted into the objective front lens element as a reversed cone. The size and shape of the illumination cone is a function of the combined numerical apertures of the objective and condenser. The objective angular aperture is denoted by the Greek letter θ and will be discussed in detail below.

One of the most significant advances in objective design during recent years is the improvement in antireflection coating technology, which helps to reduce unwanted reflections that occur when light passes through a lens system. Each uncoated air-glass interface can reflect between four and five percent of an incident light beam normal to the surface, resulting in a transmission value of 95-96 percent at normal incidence. Application of a quarter-wavelength thick antireflection coating having the appropriate refractive index can increase this value by three to four percent. Nikon's more recent CFI Plan Apochromat Lambda Series of objective lenses utilize their proprietary Nano Crystal Coat technology, which consists of several layers of ultra-low refractive index nano-sized crystals. As objectives become more sophisticated with an ever-increasing number of lens elements, the need to eliminate internal reflections grows correspondingly. Some modern objective lenses with a high degree of correction can contain as many as 15 lens elements having many air-glass interfaces. If the lenses were uncoated, the reflection losses of axial rays alone would drop transmittance values to around 50 percent. The single-layer lens coatings once utilized to reduce glare and improve transmission have now been supplanted by multilayer coatings that produce transmission values exceeding 99.9 percent in the visible spectral range.

Magnesium fluoride is one of many materials utilized in thin-layer optical antireflection coatings, but most microscope manufacturers now produce their own proprietary formulations. The general result is a dramatic improvement in contrast and transmission of visible wavelengths with a concurrent destructive interference in harmonically-related frequencies lying outside the transmission band. These specialized coatings can be easily damaged by mis-handling and the microscopist should be aware of this vulnerability. Multilayer antireflection coatings have a slightly greenish tint, as opposed to the purplish tint of single-layer coatings, an observation that can be employed to distinguish between coatings. The surface layer of antireflection coatings used on internal lenses is often much softer than corresponding coatings designed to protect external lens surfaces. Great care should be taken when cleaning optical surfaces that have been coated with thin films, especially if the microscope has been disassembled and the internal lens elements are subject to scrutiny.

There is a wealth of information inscribed on the objective barrel. Briefly, each objective has inscribed on it the magnification (e.g. 10x, 20x or 40x etc.); the tube length for which the objective was designed to give its finest images (usually 160 millimeters or the Greek infinity symbol); and the thickness of cover glass protecting the specimen, which was assumed to have a constant value by the designer in correcting for spherical aberration (usually 0.17 millimeters). If the objective is designed to operate with a drop of oil between it and the specimen, the objective will be engraved OIL or OEL or HI (homogeneous immersion). In cases where these latter designations are not engraved on the objective, the objective is meant to be used dry, with air between the lowest part of the objective and the specimen. Objectives also always carry the engraving for the numerical aperture (NA) value. This may vary from 0.04 for low power objectives to 1.3 or 1.4 for high power oil-immersion apochromatic objectives. If the objective carries no designation of higher correction, one can usually assume it is an achromatic objective. More highly corrected objectives have inscriptions such as apochromat or apo, plan, FL, fluor, etc. Older objectives often have the focal length (lens-to-image distance) engraved on the barrel, which is a measure of the magnification. In modern microscopes, the objective is designed for a particular optical tube length, so including both the focal length and magnification on the barrel becomes somewhat redundant.

For highly accurate measurements a Filar Micrometer similar to the one illustrated in Figure 6 is used. This micrometer replaces the conventional eyepiece and contains several improvements over conventional reticles. In the filar micrometer, a reticle with a measuring scale (there are many variations in scale types) and a very fine wire is brought into focus with the specimen (Figure 6(b)). The wire is mounted so that it can be slowly moved across the viewfield by the calibrated thumbscrew located on the side of the micrometer (Figure 6(a)). One complete turn of the thumbscrew (divided into 100 equal divisions) equals the distance between two adjacent reticle marks. By slowly moving the wire from one position on the specimen image to another and taking note of the changes in thumb screw numbers, the microscopist has a much more accurate measurement of distance. Filar micrometers (and other simple reticles) must be calibrated with a stage micrometer for each objective with which it will be used.

where FN is the field number in millimeters, M(O) is the objective magnification, and M(T) is the tube lens magnification factor (if any). Applying this formula to the Super Widefield eyepiece listed in Table 1, we arrive at the following for a 40x objective with a tube lens magnification of 1.25: FN = 26.5 / M(O) = 40 x M(T) = 1.25 = a viewfield diameter of 0.53 mm. Table 2 lists the viewfield sizes over the common range of objectives that would occur using this eyepiece.

The reticle in Figure 5(a) is a common element of eyepieces intended to "frame" viewfields for photomicrography. The small rectangular element circumscribes the area that will be captured on film using 35 mm format. Other film formats (120 mm and 4 x 5 inch) are delineated by sets of "corners" within the larger 35mm rectangle. In the center of the reticle is a series of circles surrounded by four sets of parallel lines arranged in an "X" pattern. These lines are used to focus the reticle and image to be parfocal with the film plane in a camera back attached to the microscope. The reticle in Figure 5(b) is a linear micrometer that can be used to measure image distances, and the crossed micrometer in 5(c) is used with polarizing microscopes to locate the alignment of samples with respect to the polarizer and analyzer. The grid illustrated in Figure 5(d) is used to partition a section of the viewfield for counting. There are many other variations of eyepiece reticles, and the reader should consult the many manufacturers of microscopes and optical accessories to determine the types and availability of these useful measuring devices.