McFarland Standards serve as a reference for adjusting the turbidity of liquid or bacterial suspensions in vials or tubes within microbiology laboratories.
They are used to help maintain and ensure that the number of bacteria remains within a specific range, thereby standardizing microbial testing.
The McFarland standard can be prepared in varying concentrations, typically ranging from 0.5 to 4, with the cell count density differing according to the concentration used.
Among these, the 0.5 McFarland standard is the most commonly applied in microbiological laboratories.
The 0.5 concentration is particularly used for antimicrobial susceptibility testing and for evaluating the performance of culture media.
Principle of McFarland Standards
A McFarland Standard is a chemical solution made from barium chloride and sulfuric acid.
The reaction between these two chemicals produces a fine precipitate of barium sulfate.
When shaken well, the turbidity of the McFarland Standard can be visually compared to a bacterial suspension of a known concentration.
McFarland turbidity standards are prepared by mixing specific volumes of 1% sulfuric acid and 1% barium chloride to achieve solutions with defined optical densities.
By adjusting the proportions of these two reagents, McFarland standards with different degrees of turbidity can be produced, each representing a particular bacterial density or cell count.
The 0.5 McFarland turbidity standard has an optical density equivalent to a bacterial suspension containing approximately 1.5 × 10^8 colony-forming units (CFU/ml).
McFarland Standard
1% BaCl2 (mL)
1% H2SO4 (mL)
Approximate Cell Count Density (×108 cells)
0.5
0.05
9.95
1.5 × 108
1.0
0.10
9.90
3.0 × 108
2.0
0.20
9.80
6.0 × 108
3.0
0.30
9.70
9.0 × 108
4.0
0.40
9.60
12.0 × 108
Preparation of McFarland Standards
Prepare a 1% solution of anhydrous barium chloride (BaCl₂) and a 1% solution of sulfuric acid (H₂SO₄).
Combine and thoroughly mix the barium chloride and sulfuric acid solutions to produce a turbid suspension.
Transfer the resulting mixture into a foil-covered screw-cap tube.
Store the McFarland standard at room temperature (25 °C) when not in use.
Be aware that the McFarland standard density solution will precipitate and form clumps over time, requiring vigorous agitation such as vortexing or shaking before each use.
Mark the tube to indicate the liquid level and check it before use to ensure that evaporation has not occurred.
Under good lighting conditions, visually compare the turbidity by holding both the bacterial sample and the McFarland Standard tubes against the black and white bars printed on the provided card.
If the bacterial suspension shows heavy turbidity, adjust it during the log growth phase by adding broth or saline via a sterile pipette until the turbidity matches that of the known McFarland Equivalence Standard.
If the test suspension appears too light, add more organisms or incubate the tubes until the turbidity is equivalent to that of the standard.
Uses of McFarland Standards
It is used in antimicrobial susceptibility testing, where the bacterial suspension is compared to the McFarland Standard before swabbing on Mueller-Hinton Agar (MHA) media.
It serves as part of quality control to check and adjust the density of bacterial suspensions for use in identification and susceptibility testing procedures.
Limitations of McFarland Standards
When using colored media, the McFarland Equivalence Standards may not provide adequate contrast, potentially leading to incorrect results or inaccurate density measurements.
Older bacterial cultures, particularly those over 24 hours old, may not correspond to the expected bacterial counts when compared to the standard.
McFarland standards are calibrated using spectrophotometer analysis, and the use of any other type of instrumentation may produce unreliable results during testing.
When using a latex standard, the suspension tubes should have the same diameter as the McFarland Latex Standard tube to ensure accuracy.
During storage, exposure of the McFarland standard to light can alter turbidity measurements and affect the accuracy of results.
References
McFarland, J. (1907). Nephelometer: An instrument for estimating the number of bacteria in suspensions used for calculating the opsonic index and for vaccine preparation. Journal of the American Medical Association, 14, 1176–1178.
