microscopy

Examples of microscopy in the following topics:

• Interference Microscopy

  • Interference microscopy is a variation of phase-contrast microscopy that uses a prism to split a light beam in two.
  • Interference microscopy uses a prism to split light into two slightly diverging beams that then pass through the specimen.
  • There are three types of interference microscopy: classical, differential contrast, and fluorescence contrast.
  • Since its introduction in the late 1960s differential interference contrast microscopy (DIC) has been popular in biomedical research because it produces high-resolution images of fine structures by enhancing the contrasted interfaces.
  • Fluorescence differential interference contrast (FLIC) microscopy was developed by combining fluorescence microscopy with DIC to minimize the effects of photobleaching on fluorochromes bound to the stained specimen.

• Enhancement of Microscopy

  • Microscopy helps us view objects that cannot be seen with the naked eye.
  • In this section we will discuss both optical and electron microscopy.
  • In optical microscopy, light reflected from an object passes through the microscope's lenses; this magnifies the light.
  • Although this type of microscopy has many limitations, there are several techniques that use properties of light and optics to enhance the magnified image:
  • Electron microscopes use electron beams to achieve higher resolutions than are possible in optical microscopy.

• Scanned-Probe Microscopy

  • Scanned-probe microscopy uses a fine probe rather than a light-beam or electrons to scan the surface of a specimen and produce a 3D image.
  • Scanned-probe microscopy (SPM) produces highly magnified and three-dimensional-shaped images of specimens in real time.
  • Scanning tunneling microscopy (STM) measures a weak electrical current flowing between tip and sample as they are held apart.
  • Near-field scanning optical microscopy (NSOM) scans a very small light source very close to the sample.
  • Describe the different types of scanning probe techniques and their advantages over other types of microscopy

• Confocal Microscopy

  • Confocal microscopy is a non-invasive fluorescent imaging technique that uses lasers of various colors to scan across a specimen with the aid of scanning mirrors.
  • The biological sample to be studied is stained with antibodies chemically bound to fluorescent dyes similar to the method employed in fluorescence microscopy .
  • Unlike in conventional fluorescence microscopy where the fluorescence is emitted along the entire illuminated cone creating a hazy image, in confocal microscopy the pinhole is added to allow passing of light that comes from a specific focal point on the sample and not the other.
  • Confocal microscopy has multiple applications in microbiology such as the study of biofilms and antibiotic-resistant strains of bacteria.
  • Tetrahymena cell, visualized using GFP-labeled anti-beta tubulin antibodies under confocal microscopy.

• Phase-Contrast Microscopy

  • Phase-contrast microscopy visualizes differences in the refractive indexes of different parts of a specimen relative to unaltered light.
  • Phase-contrast microscopy is a method of manipulating light paths through the use of strategically placed rings in order to illuminate transparent objects.
  • In phase-contrast microscopy, parallel beams of light are passed through objects of different densities.
  • Phase-contrast microscopy allows the visualization of living cells in their natural state with high contrast and high resolution .
  • One disadvantage of phase-contrast microscopy is halo formation called halo-light ring.

• Dark-Field Microscopy

  • Dark-field microscopy is ideally used to illuminate unstained samples causing them to appear brightly lit against a dark background.
  • The entire field appears dark when there is no sample on the microscope stage; thus the name dark-field microscopy.
  • Samples observed under dark-field microscopy should be carefully prepared since dust and other particles also scatter the light and are easily detected.
  • Dark-field microscopy has many applications in microbiology.

• Electron Microscopy

  • Electron microscopy uses magnetic coils to direct a beam of electrons from a tungsten filament through a specimen and onto a monitor.
  • Electron microscopy uses a beam of electrons as an energy source.
  • Describe the technique employed for electron microscopy, distinguishing between different types

• Fluorescence Microscopy

  • Fluorescence microscopy is used to study specimens that are chemically manipulated to emit light.
  • The fluorescent microscope uses a high-pressure mercury, halogen, or xenon vapor lamp that emits a shorter wavelength than that emitted by traditional brightfield microscopy.
  • Fluorescence microscopy does not allow examination of live microorganisms as it requires them to be fixed and permeabilized for the antibody to penetrate inside the cells.

• Urinalysis

  • A urinalysis (UA), also known as routine and microscopy (R&M), is an array of tests performed on urine, and one of the most common methods of medical diagnosis.
  • Another method is light microscopy of urine samples.
  • When doctors order a urinalysis, they will request either a routine urinalysis or a routine and microscopy (R&M) urinalysis; the difference being that a routine urinalysis does not include microscopy or culture.
  • Microscopy can identify casts in urine and use them to diagnose kidney diseases, by characterizing symptoms such as:

• Diagnosing Microbial Diseases

  • The methods used to diagnose microbial disease include microbial culture, microscopy, biochemical tests, and molecular diagnostics.
  • An additional tool utilized for microbial disease diagnosis is microscopy.
  • To ensure proper identification of a pathogen, microscopy, in combination with biochemical staining techniques, is often used to ensure definitive identification.
  • Compare and contrast the various methods used to diagnose microbial diseases: microbial culture, microscopy, biochemical tests and molecular diagnostics