The Membrane Filter (MF) Technique was developed in the late 1950s as an alternative method to the Most Probable Number (MPN) technique for conducting microbiological analysis of water samples.
This technique utilizes membrane filters—thin, porous sheets made from cellulose esters or other similar polymeric materials.
These filters function as two-dimensional sieves, effectively trapping all particles, whether biological or non-biological, that are larger than the filter’s pore size on their surface as fluids pass through.
Membrane filters with a pore size of 0.45 micrometers (μm) are commonly used, as they are capable of retaining bacteria and other microorganisms present in water samples.
Objective of Membrane Filter (MF) Technique
To assess the quality of water samples through the application of the membrane filter method.
Principle of Membrane Filter (MF) Technique
Membrane filters are designed with a consistent and uniform pore size, typically 0.45 µm, which is small enough to retain microorganisms.
A water sample is passed through a sterile membrane filter that is mounted in a specially designed filtration apparatus connected to a suction flask.
After filtration, the membrane filter, now holding the trapped microorganisms, is aseptically placed into a sterile Petri dish containing an absorbent pad saturated with the appropriate culture medium.
During incubation, nutrients from the medium pass through the membrane, supporting the growth of microorganisms as colonies on the upper surface of the filter.
The technique relies on the assumption that each bacterium, bacterial clump, or particle with attached bacteria will form a single visible colony, known as a colony forming unit (cfu).
The microbial results are expressed as colony forming units per unit volume of water analyzed.
The individual colonies that develop can be conveniently transferred to confirmation media for further testing.
After incubation, the number of colonies on the membrane filter is counted using a microscope.
Procedure of Membrane Filter (MF) Technique
Collect the water sample and perform any required dilutions.
Choose the suitable nutrient or culture medium and dispense it into a sterile Petri dish, ensuring the absorbent pad is evenly saturated.
Sterilize the forceps by flaming, then carefully remove the membrane filter from its sterile packaging.
Position the membrane filter within the funnel assembly.
Flame the pouring edge of the sample container and gently pour the sample into the funnel.
Activate the vacuum system to draw the entire sample through the membrane filter.
Rinse the funnel with sterile buffered water, then turn the vacuum back on to allow the rinse solution to pass completely through the filter.
Re-flame the forceps and remove the membrane filter from the funnel assembly.
Transfer the membrane filter into the prepared Petri dish.
Incubate the Petri dish at the appropriate temperature for the required duration.
After incubation, examine the filter and count the colonies using 10–15X magnification.
Confirm the identity of the colonies and record the results accordingly.
Uses of Membrane Filter (MF) Technique
The technique is employed to examine a range of water sample dilutions taken both upstream and downstream from the discharge point of a sewage treatment facility.
It is widely used following EPA-approved procedures, such as EPA Method 1103.1, to detect fecal contaminant organisms in water before the treated water is released into national water bodies.
The total number of coliform bacteria detected through this method helps determine the potability or safety of water sources.
Membrane filtration is also useful for sterilizing sensitive media components that cannot endure steam sterilization in an autoclave, such as serum, specific carbohydrate solutions, certain antibiotics, and other heat-sensitive substances.
In the pharmaceutical and cosmetics industries, membrane filter techniques are commonly used to monitor process water for the presence of Pseudomonas species.
Merits of Membrane Filter (MF) Technique
Provides results within a shorter time frame.
Capable of processing larger volumes of water samples efficiently.
Offers high accuracy, making the results easily reproducible.
Facilitates the isolation and counting of distinct bacterial colonies.
Enables the removal of bacteriostatic or bactericidal substances that are not eliminated by techniques like the Pour Plate, Spread Plate, or MPN methods.
Requires less preparation compared to many conventional techniques and is among the few methods that permit both isolation and enumeration of microorganisms.
Limitations of Membrane Filter (MF) Technique
When processing turbid samples containing high amounts of suspended solids, the particulate matter can block the membrane pores, preventing the proper passage of the intended water volume.
In cases where only small volumes of highly contaminated samples (such as sewage effluent or heavily polluted surface water) are available, it becomes necessary to dilute the sample with a sterile diluent to ensure enough volume is present for even distribution across the membrane surface.
References
Pall Corporation. Laboratory Membrane Filter Technique for Microbiology Quality Control. Available at: https://www.pall.com/en/laboratory/microbiology-qc/laboratory-membrane-filter-technique.html
Membrane Solutions. Overview of Membrane Filter Techniques in Microbiology. Accessed from: https://www.membrane-solutions.com/News_80.htm
ScienceDirect. Application of Membrane Filters in Microbiological Analysis. Available at: https://www.sciencedirect.com/science/article/pii/S0580951708701387
World Health Organization (WHO). Water Quality Monitoring – Chapter 10: Microbiological Techniques. Available at: http://www.who.int/water_sanitation_health/resourcesquality/wqmchap10.pdf
The Lab Depot Inc. Standard Operating Procedures: Membrane Filtration Method (Method 9222 A). Available at: http://www.labdepotinc.com/admin/uploads/9222_a.pdf
Europe PMC. Use of Membrane Filters in Water Microbiology – Research Article. Available at: http://europepmc.org/backend/ptpmcrender.fcgi?accid=PMC547049&blobtype=pdf
Applied and Environmental Microbiology (ASM). Quantitative Analysis Using Membrane Filtration. Available at: https://aem.asm.org/content/aem/34/1/42.full.pdf
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