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.
Table of Contents
- 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|