Bacterial spores are used to monitor the efficiency of sterilization processes. Effective sterilization requires complete destruction of microorganisms achieved through a complex interaction of physical and/or chemical conditions. Biological indicators can be used to assure the required Sterility Assurance Level (SAL) ≤ 10-6 gke biological indicators are suitable for the validation and monitoring of sterilization processes. For special application it is possible to prepare specific indicators. Standard indicators are used for the sterilization processes mentioned in the table below.
gke produces biological indicator spore strips, suspensions, ampoules and self-contained biological indicators (SCBI). For SCBIs see separate data sheet. All biological indicators are referenced to the numbers of American Type Culture Collection (ATCC™) and are specified in a certificate in each pack.
1. Spore Strips
The gke Steri-Record® biological indicators are used for validation and monitoring of dry heat, ethylene oxide, steam, formaldehyde and hydrogen peroxide (plasma) sterilization processes. The biological indicator strips can be used in packs, containers or in special process challenge devices (PCD) for validation and/or routine monitoring without losing sensitivity. If spore strips are put in PCDs, they have to be opened aseptically in a microbiological laboratory and incubated in the corresponding gke growth medium afterwards.
The gke Steri-Record®stearo ampoules can be used for sterilization monitoring of liquids or waste. All thermal processes such as steam sterilization, liquid sterilization, microwaves or mixtures of the same, except dry heat sterilization, can be monitored from 110 to 134°C. The FBio-value of the ampoule should be adapted to the F0-value of the sterilization process.
The ampoule should be placed in an area considered to be the most difficult to sterilize: directly in fluids, in the centre of packs or inside waste bags. If waste in PP plastic bags is sterilized, the bag should be filled with about one litre of water. The ampoule, connected with a string, should be positioned inside the water.
Suspensions are used if other indicators cannot be positioned in narrow lumens or valves or if special FBio-values are required which are not available in forms of other indicators. An example is the validation of hollow instruments. The suspension is placed at the most difficult location of the sterilizing agent. After sterilization the instrument can be put into a culture media to be incubated.
4. Direct Inoculation Kit
This kit is used for direct inoculation to test sterility in critical areas inside of complex hollow instruments at the most difficult penetration location. The
inoculation and the sterility test procedure afterwards is the only way to check sterilization inside of complex instruments and sealed areas. The test can be used for steam, formaldehyde or hydrogen peroxide sterilization processes with G. Stearothermophilus suspension. All types of instruments may be tested. Addtionally sterility in ethylene oxide
processes can be tested if B. atrophaeus suspension is purchased. This procedure cannot be used for routine monitoring but only for validation. To check the efficacy of the sterilization procedure, the correct inoculation with a defined population at the worst-case location is necessary. It is important to validate quantitative recovery before sterilization. After sterilization the quantitative microbiological evaluation is necessary. This whole validation procedure shall be carried out from an experienced laboratory.
5. Growth Medium
gke offers specific growth media with a pHindicator. The growth media have been developed to incubate spores G. stearothermophilus and B. atrophaeus to easily monitor growth. During the growth of the bacteria the pH-value of the medium is changing the colour allowing a quicker evaluation of biological indicators. The test tubes are optimized with the medium volume and the filling height have been adapted to incubate spore strips.
1. Spore Strips:
The gke Steri-Record® biological spore strips consist of a contaminated carrier of 6 x 38 mm in a glassine packaging that protects the spore strip from re-contamination after sterilization. For steam, ethylene oxide and formaldehyde filter paper is used strips uses a carrier made of PET. The glassine packaging has to be opened in a micro-biological laboratory where the indicator strip is aseptically transferred into the gke Steri-Record®growth medium. Populations of 105and 106 spores per strip are offered. The strips are available in different package sizes. Other populations and D-values can be produced on request.
The ampoule contains a 1.5 ml media with a pH-indicator and G. stearothermophilus spores with a population of 105or 106 CFU/ampoule which are used for steam sterilization monitoring. At the end of the sterilization process the glass ampoules are incubated. If not all spores were inactivated during the sterilization process, the germs replicate and produce acids. The resulting pH changes the colour of the liquid from purple to yellow-green. 3. Suspensions:The 10 ml suspension in 40% ethanol/water is packaged in glass bottles with rubber septum. They are available with G. stearothermophilus and B. atrophaeus spores with different populations.
4. Direct Inoculation Kit:
One test kit contains 2x 1.5 ml of G. stearothermophilus suspension in 40% Ethanol, a precision syringe and a needle of 20 cm length. The syringe is used to inject 100 µl of suspension.
5. Growth Medium:
The glass ampoule, length 10 cm, and an inner diameter of 1 cm containing 5 ml sterile CASO bouillon medium optimized for a quick growth. The size has been optimized so that spore strips of 6 x 38 mm can be transferred in the glass ampoule. The medium contains a pH-indicator that shows bacteria growth quicker than conventional media.A special growth medium for low temperature steam formaldehyde (LTSF) sterilization processes contains an extra formaldehyde neutralizer so that a pre-treatment with Na2SO3 is not required as described in EN ISO 11138-5.
gke offers incubators for ampoules, growth media and self-contained biological indicators in whichbiological indicators can be incubated. The colour change can be monitored without having to take out the biological indicators. Details see separate data sheet „incubator“.
All biological indicators comply with the standard EN ISO 11138 series and meet the performance characteristics published in the current United States Pharmacopeia (USP) and the European Pharmacopeia (EP). Population and resistances are checked for each lot and are guaranteed in a certificate. All products have a lot number. Thus, in combination with the certificate, population, D-value and expiry date can be evaluated. The above-mentioned criteria ensure continuous product quality. The continuous quality is guaranteed by the gke quality management system according to EN ISO 13485 and the inspection of various pharmaceutical companies.
The resistance characteristic is expressed by the D-value and is measured for each lot using the EN ISO 11138-1, USP and the European Pharmacopoeia (EP). The resistance against steam, dry heat, ethylene oxide and formaldehyde is measured using highly sophisticated resistometers according to EN ISO 18472. The resistance of hydrogen peroxide is tested in a resistometer designed by gke because up until now no resistometer for H2O2 is described in EN ISO 18472.The total resistance of a biological indicator is expressed as the FBio-Value:
FBio = lg Pop x D
Usually, the resistances are measured with spores immobilized on a standard carrier. The resistance value of the spores will vary with the carrier or inside different aqueous liquids. For measuring the resistance characteristic of a germ on a specific carrier or inside a liquid, please contact our application laboratory.PurityThe gke biological indicator suspensions are monitored in all stages of production assuring a purity according to EN ISO 111381 5.3.3 at all population levels.Appropriate growth characteristics and typical biochemical reactions are verified for each parent crop. Technical information about culturing, microscopic data and biochemical analysis are available for each organism type.
Spore populations are calibrated and carefully controlled at all stages of production. The spore population of suspensions are routinely determined for each production lot by means of a standard plate count determined and documented in a certificate. A test procedure is available.
- All specifications of each lot are available in a certificate available in each pack.
- Guaranteed stability of 2 years after production date.
- With the z-value mentioned in the certificate the kill time that is available for D-value at 121°C, can also be calculated for other temperatures.
- The pH-indicator allows an easier and quicker growth detection than the conventional turbidity method.
- The composition of the media and the pHindicator for G. Stearothermophilus and B. atrophaeus is different for quick recognition and growth optimization.
- The media glass ampoules are optimized in volume and diameter to allow spore strip transfer. Due to the low volume the growth is visible faster.
- The media for formaldehyde also contains a neutralizer to remove absorbed formaldehyde in order to not restrain the growth. Remaining formaldehyde can restrain the growth.
- Rapid incubation by combination of culture media with a pH-indicator.