Membrane filters have a known uniform porosity of predetermined size (generally 0.45 µm ) sufficiently small to trap microorganisms. Using the membrane filter technique, sample is passed through the membrane using a filter funnel and vacuum system. Any organisms in the sample are concentrated on the surface of the membrane.
The membrane, with its trapped bacteria, is then placed in a special plate containing a pad saturated with the appropriate medium. The passage of nutrients through the filter during incubation facilitates the growth of organisms in the form of colonies, on the upper surface of the membrane. Discrete colonies thus formed can be easily transferred to confirmation media.
Membrane filter technique is an effective, accepted technique for testing fluid samples for microbiological contamination. It involves less preparation than many traditional methods and is one of a few methods that will allow the isolation and enumeration of microorganisms. Membrane filters are used extensively in the laboratory and in the industry to sterilize fluid materials.
- Collect the sample and make any necessary dilutions.
- Select the appropriate nutrient or culture medium. Dispense the broth into a sterile Petri dish, evenly saturating the absorbent pad.
- Flame the forceps, and remove the membrane from the sterile package.
- Place the membrane filter into the funnel assembly.
- Flame the pouring lip of the sample container and pour the sample into the funnel.
- Turn on the vacuum and allow the sample to draw completely through the filter.
- Rinse funnel with sterile buffered water. Turn on the vacuum and allow the liquid to draw completely through the filter.
- Flame the forceps and remove the membrane filter from the funnel.
- Place the membrane filter into the prepared Petri dish.
- Incubate at the proper temperature and for the appropriate time period.
- Count and confirm the colonies and report the results.
Advantage of Membrane Filter Technique
- Permits testing of large sample volumes. Theoretically, almost any volume of non-turbid water could be filtered through the disk, the organisms from any given volume being deposited in the disk.
- The membrane can be transferred from one medium to another for purposes of selection or differentiation of organisms thus allowing isolation and enumeration of discrete colonies of bacteria.
- Results can be obtained more rapidly than by the conventional MPN standard methods. It provides presence or absence information within 24 hours.
Uses of Membrane Filters
- Membrane filters are used extensively in the laboratory and in the industry to sterilize materials likely to be damaged by heat sterilization. These materials include nutritional supplements of culture media, and pharmaceutical products such as drugs, hormones, sera, and vitamins.
- Effective and acceptable technique to monitor drinking water, air quality, etc.
- Useful for bacterial monitoring in the pharmaceutical, cosmetics, electronics, and food and beverage industries.
- Allows for removal of bacteriostatic or bactericidal agents that would not be removed in pour plate, spread plate, or MPN techniques.
- Can be used to allow selective passage of the organism of interest by selecting a membrane filter of appropriate porosity. Such practices are done in the pharmaceutical industry to allow the passage of a particular virus strain while preparing vaccines.
Disadvantages of Membrane Filter Technique
- Many membrane filters allow viruses and some mycoplasmas to pass through.
- They may absorb relatively large amounts of the filtrate and may introduce metallic ions into the filtrate.
Comparison of Most Probable Number (MPN) and Membrane Filtration Technique for analysis of coliform bacteria
|Most Probable Number (MPN) technique||Membrane filter technique|
|Slower: requires 48 hours for a positive||More rapid: quantitative results in or presumptive positive about 18 hours|
|More labor-intensive||Less labor-intensive|
|Requires more culture medium||Requires less culture medium|
|Requires more glassware||Requires less glassware|
|More sensitive||Less sensitive|
|Result obtained indirectly by statistical approximation (low precision)||Results obtained directly by colony count (high precision)|
|Not readily adaptable for use in the field||Readily adapted for use in the field|
|Applicable to all types of water||Not applicable to turbid waters|
|Consumables readily available in most countries||Cost of consumables is high in many countries|
|May give better recovery of stressed or damaged organisms in some circumstances||No such difference observed|
14 thoughts on “Membrane Filter Technique”
can u write bacterial endotoxin method briefly
I would be happy if you give information about various pore sizes for filtration of virus, fungi, bacteria.
please i need to know the detailed microbial principle that is involved in the vacuum filtration for testing for Escherichia coli
I was taught that after filtering, say E. Coli or K. Pneumoniae through a MF unit using a 0.45um membrane filter that it was safe to dispose of the filtered water down the drain. Is this true?
Filtration using Membrane Filter is not the recommended method for the waste water treatment. “I did not understand what do you mean by “safe to dispose”? There are other parameters (chemical pollution) that are equally important as microbial contamination.
How many time we can perform membrane filtrating method for research purpose
For samples with high coliforms counts eg raw and primary treated water from sewage treatment plant which method is preferable- membrane filtration of diluted sample or direct spreading/ pour plating of undiluted/ less diluted sample?
The above description given on membrane filtration technique is very useful for water analysis laboratories.
We have recently introduced ISO 9308-1 : 2014 Microbiology test methods for water. We filter 100 ml of undiluted water and inoculate the membrane on chromogenic coliform agar (CCA) and count the colonies and report as the number of colonies / 100 ml . In this standard under section 9 has mentioned that for the expression of results it is necessary to refer ISO 8199. Please let me know whether our way of expression of result agree with ISO 8199.
Hi, thank you very much for the detailed description of the MF method. Now I have two questions
1. Can I substitute the sterile buffered water rinse for just sterile water?
2. After you use the filter, howt is the appropiate way to clean or wash the filter?
Thank you in advance for your advice.
You can use sterile water to rinse but I did not understand the 2nd question. Do you want to reuse the filter? As the organisms are trapped in the filter paper, you won’t be able to reuse the filter paper after washing or cleaning it.
would you kindly inform me any source to obtain bacterial reference cultures required for membrane filtration technique and also any organization who conduct PT programe for Membrane filtration technique.
my questions are,(1) how do I express results when using MF technique? is it number of colonies/100ml?
(2) what is the standard for malt or flavoured drinks using MF technique?
in case if have any growth on area other than the filter paper, what we can do?
Why is buffer peptone water used for rinsing and pre wetting in case of sterile testing?