We evaluated the performance of our MicroFunnel™ filter funnels with ME25 and Supor™ PES membranes for microbial recovery and water flow rate. Testing showed ≥ 90% recovery of five key microorganisms and consistent flow rates across membrane types. Supor™ PES 0.45 µm delivered the fastest flow rate, while all tested membranes supported reliable microbial analysis. These results confirm that our MicroFunnel™ filter funnels are optimal for membrane filtration workflows in regulated environments.
Introduction
Membrane filtration is a widely accepted and effective method for testing fluid samples for microbiological contamination. Compared to traditional methods, it offers several advantages such as a reduction in preparation time and lower use of culture media.
This technique requires a porous, hydrophilic filtration membrane, composed of different polymers like polyethersulfone (PES) or mixed ester cellulose (ME). With pore sizes typically ranging from 0.1 µm to 1.2 µm , these membranes retain microorganisms and allow detection and enumeration across industries such as pharmaceutical, food and beverage, and environmental testing.
In this study, we focused on five key organisms: Escherichia coli (E. coli), Bacillus subtilis (B. subtilis), Staphylococcus aureus (S.aureus), Pseudomonas aeruginosa (P. aeruginosa), and Candida albicans (C. albicans).
Our MicroFunnel™ filter funnels simplify your microbial testing. Designed to eliminate the cleaning and sterilization steps required with reusable filter funnels, we offer our preassembled MicroFunnel™ filter funnels in either 100 mL or 300 mL volumes. Containing a variety of filter membrane options such as ME25 (mixed ester cellulose membrane) and Supor™ PES membrane, with pore sizes ranging from 0.2 to 0.8 μm, these single-use units are also gamma irradiated and supplied in multiple packing options to meet your needs.
In this application note, we evaluate the performance of MicroFunnel™ filter funnels with ME25 and Supor™ PES membranes, focusing on water flow rate and microbial recovery of the five test organisms.
Our results indicate that the water flow rate is influenced by both pore size and membrane material, with the fastest flow rate being observed in funnels containing 0.45 µm Supor™ PES membrane. We also achieved ≥ 90% recovery of all test organisms using membrane filtration when compared to the traditional plate count method, regardless of the membrane type used.
These findings support the use of the MicroFunnel™ filter funnels with ME25 0.45 µm, Supor™ PES 0.45 µm, and Supor™ PES 0.2µm as reliable tools for microbial analysis.
Material and methods
All experiments were performed in the Cytiva Harbourgate site (Portsmouth, UK).
Water flow rate assessment using MicroFunnel™ filter funnels with ME25 and Supor™ PES membranes (100 mL)
To assess water flow rate, we tested three types of MicroFunnel™ filter funnels. Their characteristics are summarised in Table 1.
Table 1. MicroFunnel™ filter funnel specifications
| Membrane type | ME25 | Supor™ | Supor™ |
| Membrane diameter | 47 mm | 47 mm | 47 mm |
| Color | White gridded | White gridded | White gridded |
| Sterile | Yes | Yes | Yes |
| Membrane pore size | 0.45 µm | 0.45 µm | 0.2 µm |
| Funnel volume | 100 mL | 100 mL | 100 mL |
We tested 12 units of each MicroFunnel™ filter funnel variants. Each MicroFunnel™ filter funnel was placed in a 3-position vacuum manifold, and 100 mL of water was filtered at 0.69 bar (10 psi, 0.069 MPa) at room temperature. The time taken to filter the water was recorded and used to calculate the flow rate.
Microbial recovery assessment using ME25 and Supor™ PES membranes
We evaluated microbial recovery of E. coli, B. subtilis, S. aureus, P. aeruginosa and C. albicans using MicroFunnel™ filter funnels containing either 0.45 µm ME25, 0.2 µm or 0.45 µm Supor™ PES membranes. Testing was performed according to guidance from USP <1227>, USP <61> and USP<62>, and the culture media used for the specific microorganisms tested in this study are summarized in Table 2.
Table 2. Microorganisms and corresponding culture media
| Organism tested | ID strain | Culture media | Temperature (°C) |
| E. coli | ATCC 8739 | Chromogenic coliform agar (CCA), Nutrient broth supplemented with agar | 34 |
| B. subtilis | ATCC 6633 | Brain heart infusion broth (BHI) supplemented with agar | 34 |
| S. aureus | ATCC 6538 | Tryptone soy broth (TSB) supplemented with agar | 34 |
| P. aeruginosa | ATCC 9027 | Pseudomonas agar (PA) base with C-N supplement | 34 |
| C. albicans | ATCC 10231 | Sabouraud dextrose broth (SDB) supplemented with agar | 24 |
Test cultures were harvested after 24 h growth and adjusted to 50 to 100 CFU/mL. We then added to 1× phosphate buffered saline (PBS) to the diluted culture, and filtered 100 mL of the solution through the MicroFunnel™ filter funnels (containing either 0.45 µm ME25, 0.2 µm or 0.45 µm Supor™ PES membranes) using a vacuum manifold at 0.69 bar (10 psi, 0.069 MPa). We used twelve of each funnel variant per microorganism tested.
To assess recovery, membranes were transferred to a Petri dish containing the specific solid medium and incubated under the conditions indicated in Table 2 until colonies were visible.
Positive controls were also performed using traditional direct enumeration methods, in accordance with USP <1227>, USP <61> and USP<62> guidance. These results were considered 100% recovery for each tested microorganism. Microbial recovery from membrane filtration was expressed relative to the recovery observed by the direct enumeration method. We also performed negative controls from non-inoculated sterile PBS throughout the filtration process.
Results and discussion
Water flow rate
This study evaluated the water flow rate of MicroFunnel™ filter funnels containing 0.45 µm ME25, 0.2 and 0.45 µm Supor™ PES membranes. We observed the fastest flow rate (434.8 ± 22.4 mL/min) with funnels containing 0.45 µm Supor™ PES membrane. Funnels with 0.45 µm ME25 membranes showed the second fastest flow rate of 319 ± 12.1 mL/min, while the slowest flow rate (215.4 ± 10.3 mL/min) was recorded in filter funnels containing 0.2 µm Supor™ PES membrane.
As expected, tighter pore sizes resulted in slower flow rates due to increased resistance. However, differences between membrane types can also be attributed to factors such as:
- Pore structure
- Membrane asymmetry
- Polymerized structure
Fig 1. Water flow rate in MicroFunnel™ filter funnels. Each bar represents the mean flow rate of 12 devices tested with 100 mL water filtered at 10 psi (0.69 MPa).
Microbial recovery on ME25 and Supor™ PES 47 mm membranes
We found ≥ 90% recovery for all five test microorganisms (E. coli, B. subtilis, S. aureus, P. aeruginosa and C. albicans) on all membranes following the membrane filtration technique (Fig 2) in comparison to the positive controls from the traditional plate count method. As can be seen in Figure 2, in several cases, recoveries exceeded 100%, including:
- E. coli on 0.45 and 0.2 µm Supor™ PES membrane
- S. aureus on ME25 membrane
- C. albicans on ME25 and 0.45 µm Supor™ PES membrane
This higher recovery is likely due to an increased detection limit, as the full-sample filtration approach retains all microorganisms present—as opposed to traditional plate count methods that rely on aliquots. Additionally, the membrane filtration technique allows for better dispersal of the microorganisms on the membrane surface, rather than the manual spreading of aliquots on the agar surface.
Recovery of B. subtilis on all membrane types following the membrane filtration technique was shown to have a high level of variability. This was likely due to agglutination of colonies of the microorganism, and this impacted on our ability to discern individual colonies. The pattern of growth was also observed on our positive control plates which confirms that the membrane filtration technique was not the cause (Fig 2).
Fig 2. Microbial recovery of the five test organisms following membrane filtration technique relative to control plates.
Colony morphology and membrane grid lines
To assess whether membrane grid lines affected colony growth, we imaged colonies of E. coli grown on TSA and CCA, and C. albicans on Sabouraud media on ME25 0.45 µm, 0.2 µm and 0.45 µm Supor™ PES membranes through the membrane filtration technique.
Colonies appeared full grown, with round shape and characteristic colour when the microorganisms were grown on their selected media (Fig 3). Additionally, the grid lines did not show any signs of inhibiting colony growth of the test organisms (Figs 3, 4 and 5).
Fig 3. Pictures of microbial colonies grown on 0.45 µm ME25 membrane from a) E. coli ATCC 8739 on TSA, b) E. coli ATCC 8739 on CCA, and c) Candida albicans ATCC 1021 on Sabouraud Dextrose Agar after membrane filtration technique.
Fig 4. Pictures of microbial colonies grown on 0.45 µm Supor membrane from a) E. coli ATCC 8739 on TSA, b) E. coli ATCC 8739 on CCA, and c) Candida albicans ATCC 1021 on Sabouraud Dextrose Agar after membrane filtration technique
Fig 5. Colonies grown on 0.45 µm ME25 membrane from (A) E. coli on TSA; (B) E. coli on CCA; and (C) C. albicans on Sabouraud dextrose agar following membrane filtration.
Conclusion
Our evaluation of MicroFunnel™ filter funnels with ME25 and Supor™ PES membranes confirms their suitability for microbial analysis using membrane filtration. Our findings include:
- Fast and consistent flow rates: the 0.45 µm Supor™ PES membrane gave the highest flow rate at 434.8 ± 22.4 mL/min. ME25 0.45 µm and Supor™ PES 0.2 µm membranes also showed reliable performance, with expected variation based on pore size.
- High microbial recovery across all membrane types: recovery rates were ≥ 90% for all five test organisms. In some cases, recovery exceeded 100%, likely due to full-sample filtration and improved microorganism dispersal.
- No interference from membrane grid lines. Grid lines did not affect colony morphology or growth, supporting their use for accurate colony counting.
- Reliable performance across pore sizes and membrane materials. All tested configurations (ME25 0.45 µm membranes, Supor™ PES 0.2 µm membranes, and Supor™ PES 0.45 µm membranes) were well suited for microbial testing workflows.
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