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Experience Documented.....

ORIGINAL ARTICLE

Year : 2014  |  Volume : 6  |  Issue : 1  |  Page : 46-52

Use of blog as a supplementary study material resource in dentistry: An Indian experience

Thavarajah Rooban, Arunachalam Mohandoss Anusa
Marundeeshwara Oral Pathology Services and Analytics, Tiruvanmiyur, Chennai, Tamil Nadu, India

Date of Web Publication

15-May-2014

Correspondence Address:
Thavarajah Rooban
Marundeeshwara Oral Pathology Services and Analytics, CS4, Bay Breeze Duraisamy Apartments, 119, East Coast Road, Tiruvanmiyur, Chennai - 600 041, Tamil Nadu
India

DOI: 10.4103/0975-8844.132586

Abstract

Objective: Social networking sites (SNS) are emerging as an alternate teaching resource. The reach and access characteristics of SNS for a noninstitutional, academic blog in an Indian setting has not been documented and this manuscript aim to address this lacunae.Materials and Methods: A blog for oral histology, an integral basic dental subject and its Facebook promotional page was created. The access characteristics were observed using Google analytics. The Facebook promotional pages of the blog access characteristics are presented. Results: A total of 582 people visited the blog during the study period. Majority of them used Google Chrome from desktop/laptop to access the blog. There were 2723 page visits in all. Visitors from 36 countries and 99 cities across the globe accessed the blog. In all through Facebook, the promotional page reached 36,543 people. The total number of people engaged through Facebook promotion page was 10,757. Conclusion: Access characteristics of the noninstitutional, academic blog have been described for the first time in dentistry. The lessons learnt through this exercise would be helpful in designing e-mentoring courses as well promotional pages of such events in the future. The necessity of making the mentors and students to adapt to e-learning and digital learning resources before drawing such programs is highlighted.

Keywords: E-learning and Facebook, Google, Indian blog, oral histology and tooth morphology, oral histology dental blog

Read the full manuscript @ HERE

Know your tool - Microscope

Image distance and object distance. With respect to the principal planes of a lens, the image-to-lens and object-to-lens distances, as predicted by the lens equation in geometrical optics. See also Lens equation.

Immunofluorescence microscopy. A mode of fluorescence microscopy in which a certain molecular species in a specimen is labeled with a specific fluorescent antibody. Fluorescence emission from excited antibodies is collected by the objective lens to form an image of the specimen. Antibodies can be made fluorescent by labeling them directly with a fluorescent dye (direct immunofluorescence) or with a second fluorescent antibody that recognizes epitopes on the primary antibody (indirect immunofluorescence).

Incandescent lamp. A bulb containing an inert gas and metal filament that emits photons as the filament becomes excited during passage of electric current. The spectrum of visible wavelengths emitted by the filament shifts to increasingly shorter wavelengths as the amount of excitation is increased. The output of incandescent lamps is very high at red and infrared wavelengths.

Infinity corrected optics. The latest optical design for microscope objective lenses in which the specimen is placed at the focal length of the lens. Used by itself, the image rays emerge from the lens parallel to the optic axis and the image plane is located at infinity. In practice, a tube lens or Telan lens located in the body of the microscope acts together with the objective to form an image in the real intermediate image plane. This optical design relaxes constraints on the manufacture of the objective lens itself and allows for placement of bulky accessory equipment such as fluorescence filter cubes in the space between the objective and the tube lens.

Intensity of light. Qualitatively, the brightness or flux of light energy perceived by the eye. By universal agreement, the term intensity, meaning the flow of energy per unit area per unit time, is being replaced by the word irradiance, a radiometric term indicating the average energy (photon flux) per unit area per unit time, or watts/meter2. As a term describing the strength of light, intensity is proportional to the square of the amplitude of an electromagnetic wave.

Interference. The sum of two or more interacting electromagnetic waves. Two waves can interfere only if a component of the E vector of one wave vibrates in the plane of the other wave. Resultant waves with amplitudes greater or less than the constituent waves are said to represent constructive and destructive interference, respectively.

Interference color. The color that results from removal of a band of visible wavelengths from a source of white light.

Interference filter. A filter made from alternating layers of different dielectric materials or layers of a dielectric material and thin metal film that transmits a specific band of wavelengths. The spacings between the layers of one-quarter or one-half wavelength allow constructive interference and reinforce propagation through the filter of a particular wavelength λ. All other wavelengths give destructive interference and are absorbed or reflected and do not propagate through the filter.

Ion arc lamp. Lamps containing an ionized gas or plasma between two electrodes that radiates visible wavelengths when excited by an electric current. Arc lamps used in light microscopy usually contain mercury vapor or xenon gas.

Irradiance of light. The radiometrically correct term for light intensity. Irradiance is the radiant flux incident per surface unit area and is given as watts/meter2. Irradiance is a measure of the concentration of power.

Isotropic. In describing the optical properties of an object or propagation medium, having identical properties in different directions.

Jablonski diagram. A diagram showing the energy levels occupied by an excited electron in an atom or molecule as steps on a vertical ladder. Singlet and triplet excited states are shown separately as ladders standing next to each other.

Koehler illumination. The principal method for illuminating specimens in the light microscope, whereby a collector lens near the light source is used to focus an image of the light source in the front aperture of the condenser. The microscope condenser  is focused to position the conjugate image of the light source in the back focal plane (diffraction plane) of the objective lens. The method provides bright, even illumination across the diameter of the specimen.

Lens equation. In geometrical optics, the equation 1/f 1/a  1/b describing the relationship between the object distance a and the image distance b for a lens of focal length f.

Light microscope. A microscope employing light as an analytic probe and optics based on glass lenses to produce a magnified image of an object specimen.

Linearly polarized light. A beam of light in which the E vectors of the constituent waves vibrate in planes that are mutually parallel. Linearly polarized light need not be coherent or monochromatic.

Long-pass filter. A colored glass or interference filter that transmits (passes) long wavelengths and blocks short ones.

Long working distance lens. An objective lens having a working distance many times greater than that of a conventional objective lens of the same magnification. A long working distance lens is sometimes easier to employ and focus, can look deeper into transparent specimens, and allows the operator greater working space for employing micropipettes or other equipment in the vicinity of the object. However, the NA and resolution are less than those for conventional lenses of comparable magnification.

Lumen. A unit of luminous flux equal to the flux through a unit solid angle (steradian) from a uniform point source of 1 candle intensity.

Lux. A unit of illumination equal to 1 lumen per square meter.

Modulation contrast microscopy (MCM). A mode of light microscope optics in which a transparent phase object is made visible by providing unilateral oblique illumination and employing a mask in the back aperture of the objective lens that blocks one sideband of diffracted light and partially attenuates the 0th-order undeviated rays. In both MCM and DIC optics, brightly illuminated and shadowed edges in the three-dimensional relief-like image correspond to optical path gradients (phase gradients) in the specimen. Although resolution and detection sensitivity are somewhat reduced compared with DIC, the MCM system produces superior images at low magnifications, allows optical sectioning, and lets you examine cells on birefringent plastic dishes.

Monochromatic. In theory, light composed of just one wavelength, but in practice, light that is composed of a narrow band of wavelengths. Owing to Heisenberg’s uncertainty principle, true monochromatic light does not exist in nature. Even the monochromatic emission from a laser or an excited atomic source has a measurable bandwidth. Therefore, while the light produced by a narrow bandpass interference filter is called monochromatic, this is just an approximation.

Multi-immersion objective lens. An objective lens whose spherical aberration is corrected for use by immersion in media of various refractive indices, including water, glycerin, and oil. A focusable lens element used to minimize spherical aberration is adjusted by rotating a focus ring on the barrel of the objective.

Multiple fluorescence filter set. A filter set for simultaneous viewing or photography of multiple fluorescent signals. The transmission profile of each filter in the set contains multiple peaks and troughs for the reflection and transmission of the appropriate excitation and emission wavelengths as in a conventional single-fluorochrome filter set. Because of constraints on the widths of bandwidths, the steepness of transmission profiles, and the inability to reject certain wavelengths, the performance is somewhat less than that of individual filter sets for specific fluorochromes.

Negative colors. Colors resulting from the removal of a certain band of visible wavelengths. Thus, white light minus blue gives the negative color yellow, because simultaneous stimulation of red and green cone cells results in this color perception. Similarly, the mixture of cyan pigment (absorbs red wavelengths) and yellow pigment (absorbs blue wavelengths) gives green, because green is the only reflected wavelength in the pigment mixture.

Negative lens. A lens that diverges a beam of parallel incident rays. A simple negative lens is thinner in the middle than at the periphery and has at least one concave surface. It does not form a real image, and when held in front of the eye, it reduces or demagnifies.

Negative phase contrast. In phase contrast optics, the term applies to systems employing a negative phase plate that retards the background 0th-order light by /4 relative to the diffracted waves. Since the diffracted light from an object is retarded /4 relative to the phase of the incident light, the total amount of phase shift between background and diffracted waves is 0 and interference is constructive, causing objects to appear bright against a gray background.

Neutral density (ND) filter. A light-attenuating filter that reduces equally the amplitudes of all wavelengths across the visible spectrum. The glass substrate contains light-absorbing colloids or is coated on one surface with a thin metal film to reduce transmission. Neutral density filters are labeled according to their absorbance or fractional transmission.

Nipkow disk. In confocal microscopy, a thin opaque disk with thousands of minute pinholes, which when rotated at high speed provides parallel scanning of the specimen with thousands of minute diffraction-limited spots. The return fluorescence emission is refocused at the same pinhole in the disk, which provides the same function in rejecting out-of-focus light as does a single pinhole in a conventional confocal microscope. Nipkow disk confocal microscopes produce a real image that can be inspected visually or recorded on a high-resolution CCD camera, whereas images of single-spot scanning microscopes are reconstructed from signals from a PMT and are displayed on a computer monitor.

Numerical aperture (NA). The parameter describing the angular aperture of objective and condenser lenses. NA is defined as n sin, where n is the refractive index of the medium between the object and the lens, and , the angle of light collection, is the apparent half-angle subtended by the front aperture of the lens as seen from a point in the specimen plane.

Objective lens. The image-forming lens of the microscope responsible for forming the real intermediate image located in the front apertures of the eyepieces.

Optical path length. In wave optics, a measure of the time or distance (measured in wavelengths) defining the path taken by a wave between two points. Optical path length is defined as n  t, where n is the refractive index and t indicates the thickness or geometrical distance. A complex optical path composed of multiple domains of different refractive index and thickness is given as  n1t1  n2t2  . . . niti.

Optical path length difference. The difference in the optical path lengths of two waves that experience refractive index domains of different value and thickness. In interference optics, differences in optical path length determine the relative phase shift and thus the degree of interference between 0th-order and higher-order diffracted waves that have their origins in a point in the object.

Optovar. A built-in magnification booster lens that can be rotated into the optical path to further increase the magnification provided by the objective by a small amount.

Ordinary ray or O ray. In polarization optics, the member of a ray pair that obeys normal laws of refraction and whose velocity remains constant in different directions during transmission through a birefringent medium. See also Extraordinary ray.

Paraboloid condenser. A high numerical aperture condenser for dark-field microscopy having a reflective surface that is a segment of a figure of revolution of a parabola. The steeply pitched illumination cone produced by the condenser is suitable for darkfield examination with high-power oil immersion objectives.

Parfocal. The property of having the same distance between the specimen and the objective turret of the microscope. With parfocal lenses, one can focus an object with one lens and then switch to another lens without having to readjust the focus dial of the microscope.

Particle wave. In phase contrast and other modes of interference microscopy, the wave (P wave) that results from interference between diffracted and surround waves in the image plane, and whose amplitude is different from that of the surrounding background, allowing it to be perceived by the eye. See also Diffracted wave and Surround wave.

Phase contrast microscopy. A form of interference microscopy that transforms differences in optical path in an object to differences in amplitude in the image, making transparent phase objects appear as though they had been stained. Surround and diffracted rays from the specimen occupy different locations in the diffraction plane at the back aperture of the objective lens where their phases are differentially manipulated in order to generate a contrast image. Two special pieces of equipment are required: a condenser annulus and a modified objective lens containing a phase plate. Because the method is dependent on diffraction and scattering, phase contrast optics differentially enhance the visibility of small particles, filaments, and the edges of extended objects. The technique allows for examination of fine details in transparent specimens such as live cells.

Phase gradient. In interference microscopy, the gradient of phase shifts in an image corresponding to optical path differences in the object.

Phase object. Objects that shift the phase of light as opposed to those that absorb light (amplitude objects) as the basis for image formation. See also Amplitude object.

Phase plate. In phase contrast microscopy, a transparent plate with a semitransparent raised or depressed circular annulus located at the rear focal plane of a phase contrast objective. The annulus reduces the amplitude of background (0th order) waves and advances or retards the phase of the 0th-order component relative to diffracted waves. Its action is responsible for the phase contrast interference image.

Phosphorescence: The relatively slow (9 s) emission of photons after excitation of a material by light or other radiation source.

Polar: The common term applied to a sheet of linear polarizing film (dichroic filter or Polaroid filter) and particularly to its use as a polarizer or analyzer in producing and analyzing polarized light.

Polarizability. In polarization optics, a property describing the strength of interaction of light with molecules in a manner that depends on the orientation of atomic bonds. Light waves interact more strongly with molecules when their E vectors are oriented parallel to the axis defining light-deformable (polarizable) covalent bonds such as the axes of long-chain hydrocarbon polymers like polyvinyl alcohol, cellulose, and collagen. This geometry is supported when an incident light ray is perpendicular to the long axis of the polymer. Interaction of light with molecules along their polarizable axis retards wave propagation and accounts for the direction-dependent variability in their refractive index, a property known as birefringence.

Polarization cross. In polarization microscopy, the appearance of a dark upright cross in the back aperture of the objective lens under conditions of extinction with two crossed polars. Ideally, the back aperture is uniformly dark under this condition, but the depolarization of light by the curved lens surfaces of the condenser and objective lenses causes brightenings in four quadrants and hence the appearance of a cross.

Polarization microscopy. A mode of light microscopy based on the unique ability of polarized light to interact with polarizable bonds of ordered molecules in a directionsensitive manner. Perturbations to waves of polarized light from aligned molecules in an object result in phase retardations between sampling beams, which in turn allow interference-dependent changes in amplitude in the image plane. Typically the microscope contains a polarizer and analyzer, and a retardation plate or compensator.  Image formation depends critically on the existence of ordered molecular arrangements and a property known as double refraction or birefringence.

Polarized light. Light waves whose E vectors vibrate in plane-parallel orientation at any point along the axis of propagation. Polarized light can be linearly polarized (vibrations at all locations are plane parallel) or elliptically or circularly polarized (vibration axis varies depending on location along the propagation axis). Polarized light need not be monochromatic or coherent.

Polarizer. A device that receives random light and transmits linearly polarized light. In microscopy, polarizers are made from sheets of oriented dichroic molecules (Polaroid filter) or from slabs of birefringent crystalline materials.

Positive colors. Colors that result from mixing different wavelengths of light. The equal mixture of red and green wavelengths results in the perception of yellow, a positive color.

Positive lens. A lens that converges a beam of parallel incident rays. A simple positive lens is thicker in the middle than at the periphery, and has at least one convex surface. A positive lens forms a real image and enlarges or magnifies when held in front of the eye.

Positive phase contrast. In phase contrast optics, the term applies to systems employing a positive phase plate that advances the background wave by /4 relative to the diffracted wave. Since the diffracted light from an object is retarded /4 relative to the phase of the incident light, the total phase shift between background and diffracted waves is /2 and interference is destructive, causing objects to appear dark against a gray background.

Principal plane. For a simple thin lens, the plane within the lens and perpendicular to the optic axis from which the focal length is determined. Thick simple lenses have two principal planes separated by an intervening distance. Complex compound lenses may have multiple principal planes.

Rayleigh criterion for spatial resolution. The criterion commonly used to define spatial resolution in a lens-based imaging device. Two point sources of light are considered to be just barely resolved when the diffraction spot image of one point lies in the first-order minimum of the diffraction pattern of the second point. In microscopy, the resolution limit d is defined, d m/1.22 λ/(NAobjective  NAcondenser), where λ is the wavelength of light and NA is the numerical aperture of the objective lens and of the condenser.

Real image. An image that can be viewed when projected on a screen or recorded on a piece of film.

Real intermediate image. The real image focused by the objective lens in the vicinity of the oculars of the microscope.

Refraction. The change in direction of propagation (bending) experienced by a beam of light that passes from a medium of one refractive index into another medium of different refractive index when the direction of propagation is not perpendicular to the interface of the second medium.

Refractive index ellipsoid and wavefront ellipsoid. An ellipsoid is the figure of revolution of an ellipse. When rotated about its major axis, the surface of the ellipsoid is used to describe the surface wavefront locations of E waves propagating outward from a central point through a birefringent material. The same kind of figure is used to describe the orientation and magnitude of the two extreme refractive index values that exist in birefringent uniaxial crystals and ordered biological materials.

Relative retardation. In polarization optics, the relative shift in phase between two waves expressed in fractions of a wavelength.

Relay lens. An intermediate magnifying lens in an imaging system placed between the objective and the real intermediate image. In video, so-called TV lenses increase the magnification of the image projected on the camera 2- to 8-fold.

Short-pass filter. A colored-glass or interference filter that transmits (passes) short wavelengths and blocks long ones.

Simple lens. A lens consisting of a single lens element and distinct from a compound lens having multiple lens elements.

Spatial filter. A filter that selectively manipulates a location in an image such as an aperture in a field plane of a microscope or a sharpening or blurring filter in image processing.

Spatial frequency. The reciprocal of the distance between two objects (periods/ distance).

Spatial frequency filter. A filter that selectively manipulates a location in the diffraction plane in a microscope (aperture plane masks in modulation contrast microscopy) or a mask applied to Fourier transforms to manipulate low and high spatial frequency information in image processing.

Spatial resolution. The resolution of component features in an image. In optical systems, resolution is directly proportional to the wavelength and inversely proportional to the angular aperture. The practical limits on wavelength and angular aperture determine the limit of spatial resolution, which is approximately one-half the wavelength of light.

Spectral range. The range of wavelengths, or bandwidth, under consideration.

Spherical aberration. A lens aberration typical of lenses with spherical surfaces that causes paraxial rays incident on the center and periphery of a lens to be focused at different locations in the image plane. The degree of aberration increases with the decreasing focal ratio of the lens. The aberration can be corrected in simple lenses by creating aspherical surfaces.

Stokes shift. The distance in nanometers between the peak excitation and peak emission wavelengths of a fluorescent dye.

Thin lens. A lens whose thickness is small compared to its focal length. A line through the center of the lens (a plane representing the two coincident principal planes of the lens) provides a reasonably accurate reference plane for refraction and object and lens distance measurements. Lenses are assumed to be thin when demonstrating the principles of graphical ray tracing.

Tube lens or Telan lens. An auxiliary lens in the body of the microscope, which in conjunction with an infinity focus objective lens forms the real intermediate image. The Telan lens provides some of the correction for chromatic aberration, which lessens constraints on the manufacture of the objective lens.

Uniaxial crystal. A birefringent crystal characterized by having a single optic axis.

Virtual image. An image that can be perceived by the eye or imaged by a converging lens, but that cannot be focused on screen or recorded on film as can be done for a real image. The image perceived by the eye when looking in a microscope is a virtual image.

Wavelength. The distance of one beat cycle of an electromagnetic wave. Also, the distance between two successive points at which the phase is the same on a periodic wave. The wavelength of light is designated λ and is given in nanometers.

Wollaston prism. In interference microscopy, a beam splitter made of two wedgeshaped slabs of birefringent crystal such as quartz. In differential interference contrast (DIC) microscopy, specimens are probed by pairs of closely spaced rays of linearly polarized light that are generated by a Wollaston prism acting as a beam splitter. An important feature of the prism is its interference plane, which lies inside the prism (outside the prism in the case of modified Wollaston prism designs).

Working distance. The space between the front lens surface of the objective lens and the coverslip. Lenses with high NAs typically have short working distances (60–100 micro m). Lenses with longer working distances allow you to obtain focused views deep within a specimen. 

Definitions in Microscopy

Diffraction grating. A transparent or reflective substrate containing an array of parallel lines having the form of alternating grooves and ridges with spacings close to the wavelength of light. Light that is reflected by or transmitted through such a grating becomes strongly diffracted. Depending on the geometry of illumination and wavelength, a grating can generate color spectra and patterns of diffraction spots.

Diffraction plane. One of the aperture planes of the light microscope containing the focused diffraction image of the object. Under conditions of Koehler illumination, the diffraction plane is located in or near the back focal plane of the objective lens.

Distortion. An aberration of lenses, where the magnification factor describing an image varies continuously between the central and peripheral portions of the image. Depending on whether the magnification is greater at the center or at the periphery, the distortion can be of the barrel or the pincushion type, respectively.

Double refraction. In polarization optics, the splitting of light into distinct O and E rays in a birefringent material. When a birefringent crystal of calcite is placed on a page  of printed words, the effects of double refraction are clearly observed as an overlapping, double image of the text.

Emission filter. In fluorescence microscopy, the final element in a fluorescence filter cube, which transmits fluorescence emission wavelengths while blocking residual excitation wavelengths. Commonly called a barrier filter. Emission filters are colored glass or interference filters and have the transmission properties of a bandpass or long-pass filter.

Emission spectrum. In fluorescence, the spectrum of wavelengths emitted by an atom or molecule after excitation by a light or other radiation source. Typically, the emission spectrum of a dye covers a spectrum of wavelengths longer than the corresponding excitation spectrum.

Epi-illumination. A common method of illumination in fluorescence microscopy, where the illuminator is placed on the same side of the specimen as the objective lens, and the objective performs a dual role as both a condenser and an objective. A dichroic mirror is placed in the light path to reflect excitatory light from the lamp toward the specimen and transmit emitted fluorescent wavelengths to the eye or camera.

Excitation filter. In fluorescence microscopy, the first element in a fluorescence filter cube and the filter that produces the exciting band of wavelengths from a broadband light source such as a mercury or xenon arc lamp. Commonly the excitation filter is a high-quality bandpass interference filter.

Excitation spectrum. In fluorescence, the spectrum of wavelengths capable of exciting an atom or a molecule to exhibit fluorescence. Typically the excitation spectrum covers a range of wavelengths shorter than the corresponding  fluorescence emission spectrum.

Eyepiece or ocular. The second magnifying lens of the microscope used to focus a real magnified image on the retina of the real intermediate image produced by the objective. The added magnification provided by the eyepiece increases the angular magnification of the virtual image perceived by the eye. The typical range of eyepiece magnifications is 5–25.

Eyepiece telescope. See Bertrand lens.

Field diaphragm. A variable diaphragm located in or near the aperture plane of the light source that is used to reduce the amount of stray light in the object image. Since the edge of the diaphragm is conjugate with the object plane under conditions of Koehler illumination, the field diaphragm is used as an aid in centering and focusing the condenser lens.

Field planes. That set of conjugate focal planes representing the field diaphragm, the object, the real intermediate image, and the retina.

Flat-field correction. In image processing, the procedure used to obtain a photometrically accurate image from a raw image. A so-called dark frame containing bias and thermal counts is subtracted from the raw image and from a “flat” or “background” image. The dark-subtracted raw image is then divided by the dark-subtracted flatfield image to produce the corrected image. With operation, all optical faults are removed. The photometric relation of pixel values to photoelectron count is also lost during division, although the relative amplitudes of pixel values within an image are retained. See also Dark frame and Flat-field frame.

Fluorescence. The process by which a suitable molecule, transiently excited by absorption of external radiation (including light) of the proper energy, releases the energy as a longer-wavelength photon. This process usually takes less than a nanosecond.

Fluorescence microscopy. A mode of light microscopy whereby the wavelengths of fluo-rescence emission from an excited fluorescent specimen are used to form an image.

Fluorite or semiapochromat lens. Objective lenses made of fluorite or Ca2F, a highly transparent material of low color dispersion. The excellent color correction afforded by simple fluorite elements accounts for their alternative designation as semiapochromats.  The maximum numerical aperture is usually limited at 1.3.

Fluorochrome. A dye or molecule capable of exhibiting fluorescence.

Fluorophore. The specific region or structural domain of a molecule capable of exhibiting fluorescence. Examples include the fluorescein moiety in a fluoresceinconjugated protein and the tetrapyrrole ring in chlorophyll.

Focal length. The distance along the optic axis between the principal plane of a lens and its focal plane. For a simple converging (positive) lens illuminated by an infinitely distant point source of light, the image of the point lies precisely one focal length away from the principal plane.

Focal ratio or f-number. The ratio of the focal length of a lens to the diameter of its aperture.

Fovea. A 0.2–0.3 mm diameter spot in the center of the macula on the retina that lies on the optic axis of the eye and contains a high concentration of cone cell photoreceptors for color vision and visual acuity in bright light conditions.

Frame averaging or Kalman averaging. In electronic imaging, the method of averaging a number of raw image frames to reduce noise and improve the signal-to-noise ratio. The signal-to-noise ratio varies as the square root of the number of frames averaged.

Halo. In phase contrast microscopy, characteristic contrast patterns of light or dark gradients flanking the edges of objects in a phase contrast image. Halos are caused by the phase contrast optical design that requires that the image of the condenser annulus and objective phase plate annulus have slightly different dimensions in the back focal plane of the objective.

Huygens’ principle. A geometrical method used to show the successive locations occupied by an advancing wavefront. An initial source or wavefront is treated as a point source or a collection of point sources of light, each of which emits a spherical wave known as a Huygens’wavelet. The surface of an imaginary envelope encompassing an entire group of wavelet profiles describes the location of the wavefront at a later time, t. Huygens’ principle is commonly used to describe the distribution of light energy in multiple interacting wavefronts as occurs during diffraction and interference. 

Inking the Specimen

INKING THE SPECIMEN

·         Various Water/organic fluids insoluable inks and colored powders can be used to mark critical points on the specimen.

·         These dyes and powders may help orient both the gross specimen and the histologic section. For example, colored tattoo powder sprinkled on the outer surface of a cystic mass can be used to distinguish between the outer and inner aspects of the cavity.

·         Similarly, India ink can be painted on the surgical margins so that they can be easily recognized at the time of histologic examination.

·         Indeed, many times the critical distinction of whether a neoplasm extends to the surgical margin depends entirely on the absence or presence of ink.

·         Given the important implications of an inked surface, these inks should be carefully and judiciously applied to the gross specimen.

·         Keep in  mind that just as the effective use of inks can facilitate the histologic interpretation, the careless and improper use of these inks can befuddle the microscopic findings.

·         The implications of sloppily applied ink that runs across a surface where it does not belong will be disastrous.

·         The following guidelines outline the proper application of inks:

o   If possible, apply ink before sectioning the specimen.

o   Do not use excessive ink. 

o   Dry the surface of the specimen with paper towels before applying ink.

o   When applied to a dry surface, ink is more likely to stick to the desired surface and less likely to run onto other areas of the specimen.

o   Allow the ink to dry before further processing the specimen.

o   Do not cut across wet ink, as the knife is likely to carry the ink onto the cut surface.

Cassette Dimensions

Inside dimensions for a screened cassette are: 2.5 x 2.0 x 0.3cm

Inside dimensions for a standard slotted cassette are: 3.0 x 2.5 x 0.3cm

Routine tissue sections submitted in standard slotted cassettes should be no larger than 2.5 x 1.5 x 0.3cm to allow for proper processing.

Screened Cassettes

• Distinct advantages to using screened cassettes with small tissue biopsies;

1.Negates the need to wrap samples, a big time saver.

2.Positive seal created when properly closed.

3.Prevents cross-contamination with other tissues during processing

Cost is a major Disadvantage

For Small Biopsies

• Number of Pieces Each Container - To the best of your ability, give an accurate count. Check the container (to include the lid!) and req. for a reference to the number of pieces submitted.

Often samples are fragmented. In this case, count the number of significant pieces, give size(s), and add the descriptor “fragmented”. Additional Descriptors for Number

• Additional descriptors for number of pieces; Multiple(>10) - Give aggregate dimensions with average size each. Do not submit more than 5 per cassette.  Myriad – too many to count (fragments), give aggregate dimensions. Filter thru screened cassette.

• State the size(s) of the tissue(s) received: Always stated in the context of mm.’s or cm.’s Do not use inches. If you start the case using mm.’s, then use mm.’s throughout. If you start the case with cm.’s, then use cm.’s throughout. Ex. 0.4cm or 4mm.  Whole cm.-Do not use decimal point and zero.

• Referencing the size(s) of the piece(s): If only one, self explanatory. If two or more of the same size, then state as: __ mm. or cm. each. If two of different sizes, then state as: __ and __ mm. or cm. each.

If three or more with different sizes, then state as: ranging from __ mm. or cm. to __mm. or cm.

Cores - • Whenever possible, give exact count of tissue cores. Not necessary to give the diameter of the cores in most cases, but always give the length of each core.

• Indicate formalin exposure times with cores.

Biopsy Tissue Configurations

Irregular /Fragmented/ Cores/Polypoid/Sessile/Pedunculated

Punch Biopsies(Derms)- • Elipses(Derms)/• Shaves (Derms)

To Cut or Not To Cut!

• Most diagnostic cases do not require additional cutting or inking.

• Polyps >5mm should have their bases inked and be bisected.

• Punch biopsies >4mm should also be bisected.

Tissue Sampling Techniques - Small Biopsies & Triaging

Tissue Sampling Techniques - Small Biopsies & Triaging

Most Important Steps

• Patient identification - Identification on the requisition must match the container(s). This includes name. Accession number must match requisition, specimen container and cassette.

Receiving/Accepting Specimens

• If you accept a specimen in the receiving area with incorrect information, it becomes the laboratories problem to get it back to the sender for correction. Better to refuse the specimen at the time of delivery.

• Never process a specimen without a patient name. Never label the container yourself with patient name or specimen source/type.

GROSS BENCH RULES

• Never have more than one specimen out at a time.

• Close containers when leaving the area.

• Don’t leave small biopsies on the cutting board or on paper towels.

• Keep cutting area neat, clean and organized.

• Keep sharps in clear view, not under toweling etc. Clear cutting area of sharps when leaving, and disinfect the cutting board and countertop.

• Beware of “carry-over” from case to case

Specimens can be subclassified

Depending on utility

Diagnostics specimen

Routines – Small and Large

Dermatology

Resections

Others

Depending on Nature of handling:

·         Specimens only requiring transfer from container to tissue cassette.

o   All small biopsies

o   Bone marrow & Aspirates.

o   Punch biopsies.

o   Needle biopsies

o   Any biopsies not requiring dissection

·         Specimens requiring transfer, but with standard sampling, counting, weighing or slicing.

o   Sebaceous cysts.

o   Small lipomas.

o   Unremarkable tonsils.

o   Unremarkable nasal polyps.

o   Temporal arteries.

o   Thyroglossal cysts.

o   Lymph nodes.

·         Simple dissection required with sampling needing a low level of diagnostic assessment and/or preparation.

o   Salivary gland – non-tumour.

o   Cone biopsy.

o   Small soft tissue tumours.

o   Skin biopsies – benign – requiring dissection.

o   Simple small benign biopsies.

·         Dissection and sampling required needing a moderate level of assessment.

o   Salivary gland – tumours.

o   Pigmented skin lesions.

o   Complex (non-neoplastic) gastrointestinal resections.

·         Specimens requiring complex dissection and sampling methods.

o   Bone tumours.

o   Neck dissection.

o   Mandibulectomy.

Diagnostic

• Diagnostic Cases: Small tissues being submitted to establish a diagnosis or monitor status.

Typical features of biopsy tissues;

Small in size (minute to ~1cm)

Do not require orientation

Require counting when possible

Often are submitted in “toto”

Currettings

Currettage specimens come as multiple tissue fragments admixed with; Blood or Blood Clot /Mucous

Sample EMC dictation - “Specimen consists of multiple fragments of pink/tan irregular soft tissue admixed with mucous and blood having aggregate dimensions of _____ x ______ x _____cm which are submitted in toto in a single cassette, levels are requested.

·         Small specimens should never be forcibly squeezed between the ends of a forceps or the tips of the fingers. Instead, small specimens should be gently lifted from the specimen container using the end of a wooden applicator stick or pickups. Alternatively, small specimens can be filtered directly into a tissue bag, avoiding instrumentation altogether.

·         Small specimens should be quickly placed in fixative. Ideally, most small specimens (i.e., less than 1 cm) should reach the surgical pathology laboratory already in fixative.

·         This requires that physician offices, biopsy suites, and operating rooms be supplied with appropriate fixatives, and that all personnel involved be instructed as to their proper use. Sometimes delays in fixation are necessary, as when a frozen section is required or when special tissue processing is indicated. In these instances, the tissue should be kept damp in  saline-soaked gauze.

·         Never leave small tissue fragments exposed to the air on the cutting table, and never place these small fragments directly on a dry paper towel. These practices are sure to hasten tissue desiccation.

·         For extremely small specimens, the journey from specimen container to histologic slide is a treacherous one, and they may be lost at any point along the way. For this reason, it is a wise practice to identify these small tissue fragments first and then mark the fragments so that they can be found more easily by the histotechnologist.

·         Before the specimen container is even opened, check its contents for the size and number of tissue fragments, and record these in the gross description.

·         If no tissue is seen or if inconsistencies with the requisition form are noted, carefully open the specimen container and thoroughly examine its surfaces (including the undersurface of the lid) for adherent tissue fragments. If no tissue is found or if discrepancies persist, the submitting physician should be notified immediately, and the outcome of this investigation should be documented in the surgical pathology report.

·         Once all of the tissue is identified in the specimen container, efforts should be taken to ensure that it safely reaches the histology laboratory and that it is easily identified for embedding and sectioning. Minute tissue fragments should be wrapped in porous paper or layered between porous foam pads before they are placed in the tissue cassette.

·         Before these fragments are submitted  to the histology laboratory, they can be marked with eosin or mercurochrome so that they are easier for the histotechnologist to see.