Staining and Aseptic Technique

The microbiology laboratory is unique and different from the laboratory of other biological and medical sciences in that it has some fundamental techniques which the microbiologists use to manipulate microorganisms for improved productivity. Some of these techniques (as shall be discussed in this section) are what distinguishes the microbiologist from other scientists; and these techniques must be dutifully followed in order to achieve better research results. It is also important that students and tutors of microbiology acquaint themselves of these techniques so as to always be on top in their chosen discipline. Microbiology like the other biological sciences is a practical discipline, and it cannot do without some modus operandi which is universally acceptable and central to any research performed in the microbiology laboratory. These microbiology techniques including isolation, inoculation, culturing, incubation and microscopy among others shall be discussed in this section. Students of microbiology are advised to ensure continues practice of these techniques in order to be familiar with them.

STAINING TECHNIQUE

Staining is any microbiological process which increases the contrast of organisms when certain dyes or stains are applied to them prior to their examination under the microscope. It is generally the process of colouring specimens and microorganisms so that they can be easily observed and distinguished under the microscope. Staining in the microbiology laboratory is usually carried out on dead microbial cells which must have undergone a series of smearing during smear preparation. In some scenarios, microbial cells are only stained once but staining can involve a series of process in which bacterial cells are stained several times using different types of dyes or counterstains meant to reveal certain features of the organism during examination under the microscope. Some commonly used stains in the microbiology laboratory include crystal violet, methylene blue, lactophenol cotton blue, Giemsa stain, eosin stain, carbol fuchsin, acridine orange and fluorescent dyes among others. Gram staining, negative staining and acid fast staining are some of the commonly encountered staining techniques used in the microbiology laboratory. Staining techniques generally helps microbiologist to observe microbial cells under the microscope and describe them appropriately based on their ability to react with certain stains or dyes.

ASEPTIC TECHNIQUE

Aseptic technique refers to all the quality control and precautionary measures taken by microbiologists in the laboratory in order to ensure that all working apparatuses are germ-free. It involves numerous processes taken to reduce or prevent the contamination of microbial cultures, reagents and culture media. Aseptic techniques also include all other precautionary measures taken to protect the laboratory personnel from direct or indirect contamination from harmful microorganisms. Working inside the biological safety cabinet or hood (Figure 1), flaming the inoculating loop prior to the transfer of microbes, disinfecting the work bench with antimicrobial agents, sterilizing or autoclaving used plates and cultures before disposing them, flaming the neck of reagent bottles prior to dispensing and flaming the surfaces of forceps and other working tools among others are some of the important aseptic techniques observed in the microbiology laboratory. The mouth or neck of reagent bottles and tubes or flask containing culture media should be properly flamed over a Bunsen burner flame (Figure 2) prior to dispensing in order to prevent contamination of the medium. In summary, the main aim of ensuring aseptic technique in the microbiology laboratory is to prevent contamination.

Figure 1:  A microbiologists performing cell/tissue culture experiment in a biological safety cabinet or hood. A clean environment as provided by the hood is critical for microbiological investigations in the laboratory, as this method will help to prevent contamination of the personnel and working materials.

Figure 2: An illustration of blue flame (arrow) from a Bunsen burner. Passing the neck of culture media flasks or vessels through the flame from a Bunsen burner is a routine aseptic technique performed in the microbiology laboratory.

REFERENCES

Atlas R.M (2010). Handbook of Microbiological Media. Fourth edition. American Society of Microbiology Press, USA.

Balows A, Hausler W, Herrmann K.L, Isenberg H.D and Shadomy H.J (1991). Manual of clinical microbiology. 5th ed. American Society of Microbiology Press, USA.

Basic laboratory procedures in clinical bacteriology. World Health Organization (WHO), 1991. Available from WHO publications, 1211 Geneva, 27-Switzerland.

Black, J.G. (2008). Microbiology:  Principles and Explorations (7th ed.). Hoboken, NJ: J. Wiley & Sons.

Garcia L.S (2010). Clinical Microbiology Procedures Handbook. Third edition. American Society of Microbiology Press, USA.

Garcia L.S (2014). Clinical Laboratory Management. First edition. American Society of Microbiology Press, USA.

Ira R (1995). Bacteriology, Standard Operative procedure manual for microbiology laboratories, National Institute of Biologicals.  Pp. 73-97.

Madigan M.T., Martinko J.M., Dunlap P.V and Clark D.P (2009). Brock Biology of Microorganisms, 12th edition. Pearson Benjamin Cummings Inc, USA.

Woods GL and Washington JA (1995). The Clinician and the Microbiology Laboratory. Mandell GL, Bennett JE, Dolin R (eds): Principles and Practice of Infectious Diseases. 4th ed. Churchill Livingstone, New York.

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