Light microscopy can be done quickly, but accuracy depends on the experience of the microscopist and quality of equipment. Regulations often limit physicians’ use of microscopy for diagnostic purposes outside a certified laboratory. Show
Microscopic examination of tissue may be required to distinguish invasive disease from surface colonization—a distinction not easily achieved by culture methods. Most specimens are treated with stains that color pathogens, causing them to stand out from the background, although of unstained samples can be used to detect fungi and certain other pathogens. The clinician orders a stain based on the likely pathogens. However, no stain is 100% specific (ie, different organisms may stain similarly). Most samples are treated with and, if mycobacteria are suspected, with an . However, some pathogens are not easily visible using these stains; if these pathogens are suspected, different stains or other identification methods are required. Because microscopic detection usually requires a microbe concentration of at least about 1 ×104-5/mL, most body fluid specimens (eg, cerebrospinal fluid) are concentrated (eg, by centrifugation) before examination. Gram stainThe Gram stain does the following:
Such characteristics can direct antibiotic therapy pending definitive identification. Finding a mixture of microorganisms with multiple morphologies and staining characteristics on Gram stain suggests a contaminated specimen or a polymicrobial bacterial infection. Finding many squamous cells in a sputum specimen suggests that the specimen is contaminated with saliva and thus is of limited diagnostic usefulness. To do a Gram stain, technicians heat-fix specimen material to a slide and stain it by sequential exposure to Gram crystal violet, iodine, decolorizer, and counterstain (typically safranin). Acid-fast and modified acid-fast stainsThese stains are used to identify the following:
Although detection of mycobacteria in sputum requires at least 10,000 organisms/mL, mycobacteria are often present in lower levels, so sensitivity is limited. Usually, several mL of sputum are decontaminated with sodium hydroxide and concentrated by centrifugation for acid-fast staining. Specificity is better, although some moderately acid-fast organisms are difficult to distinguish from mycobacteria. Fluorescent stainsFluorescent stains allow detection at lower concentrations (< 1 × 104 cells/mL). Examples are
Coupling a fluorescent dye to an antibody directed at a pathogen (direct or indirect immunofluorescence) should theoretically increase sensitivity and specificity. However, these tests are difficult to read and interpret, and few (eg, Pneumocystis and Legionella direct fluorescent antibody tests) are commercially available and commonly used. Wet mountsWet mounts of unstained samples can be used to detect the following via darkfield microscopy:
Visibility of fungi can be increased by applying 10% potassium hydroxide (KOH) to dissolve surrounding tissues and nonfungal organisms. India ink (colloidal carbon) stainIndia ink stain is used to detect mainly Cryptococcus neoformans Cryptococcosis Cryptococcosis is a pulmonary or disseminated infection acquired by inhalation of soil contaminated with the encapsulated yeasts Cryptococcus neoformans or C. gattii. Symptoms... read more Warthin-Starry stain and Dieterle stainThese silver stains are used to visualize bacteria such as
Wright stain and Giemsa stainThese stains are used for detection of the following:
Trichrome stain (Gomori-Wheatley stain) and iron hematoxylin stainThese stains are used to detect intestinal protozoa. The Gomori-Wheatley stain is used to detect microsporidia. It may miss helminth eggs and larvae and does not reliably identify Cryptosporidium. Fungi and human cells take up the stain. The iron hematoxylin stain differentially stains cells, cell inclusions, and nuclei. Helminth eggs may stain too dark to permit identification. What are the 2 types of microscopes that are commonly used in the laboratory?Applications of laboratory microscopes
The goal of any laboratory microscope is to produce clear, high-quality images, whether an optical microscope, which uses light to generate the image, a scanning or transmission electron microscope (using electrons), or a scanning probe microscope (using a probe).
Which microscope is most used in the clinical laboratory?The light microscope is perhaps the most well-known and well-used tool in the laboratory.
What are 2 of the most common types of microscopes?Today, we still use light microscopes to see objects that are too small to see with the naked eye, but we also use other types of microscopes, like electron microscopes and fluorescence microscopes, to see things that you can't see even with a light microscope!
What is the most commonly used microscopy technique?Optical Microscopy
This is the most common and well-known type of microscopy, and involves magnifying the image of the object by passing light through or reflecting light off it, and then examining this light through a single or multiple lenses.
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