Question

Discuss the pros and cons of light microscopy with and without staining.

Answer 1

Advantages

• Brightfield microscopy is very simple to use with fewer adjustments needed to be made to view specimens.
• Some specimens can be viewed without staining and the optics used in the brightfield technique don’t alter the color of the specimen.
• The advantage of using stains to look at cells is that stains reveal these details and more. The arrangement of cells within a tissue reveals the health of that tissue. Multiple stains can used simultaneously to mark different cells by different colors.
• It is adaptable with new technology and optional pieces of equipment can be implemented with brightfield illumination to give versatility in the tasks it can perform.

Disadvantages

Certain disadvantages are inherent in any optical imaging technique.

• By using an aperture diaphragm for contrast, past a certain point, greater contrast adds distortion. However, employing an iris diaphragm will help compensate for this problem.
• Brightfield microscopy can’t be used to observe living specimens of bacteria, although when using fixed specimens, bacteria have an optimum viewing magnification of 1000x.
• Light microscopes can show a useful magnification only up to 1000-2000 times. This is a physical limit imposed by the wavelength of the light.
• Brightfield microscopy has very low contrast and most cells absolutely have to be stained to be seen; staining may introduce extraneous details into the specimen that should not be present.
• Also, the user will need to be knowledgeable in proper staining techniques.
• Lastly, this method requires a strong light source for high magnification applications and intense lighting can produce heat that will damage specimens or kill living microorganisms.
• A disadvantage of staining a specimen is that the stain can kill off the live specimen fairly.