GB 18281.1-2000 Biological indicators for sterilization of health care products Part 1: General
Some standard content:
GB 18281. 1—2000
All technical contents of this standard are mandatory. This standard is equivalent to the international standard 1S011138-1: 1994\Medical and health products-Biological indicators-Part 1: General
Appendix A, Appendix B, Appendix C, Appendix D, Appendix E and Appendix F of this standard are all appendices of the standard. This standard is proposed by the State Food and Drug Administration. The water standard is under the jurisdiction of the National Technical Committee for Standardization of Disinfection Technology and Equipment. The drafting units of this standard are: Guangzhou Medical Device Quality Supervision and Inspection Center of the State Drug Administration, Jiangsu Provincial Health and Epidemic Prevention Station, Disinfection Department of Jilin Provincial Health and Epidemic Prevention Station, and Sichuan East Xinhua Medical Device Co., Ltd. The main drafters of this standard are: Chen Jiahua, Gu Jian, Huang Xinyu, and Yang Zhaoxu. 605
GB 18281. 1—2000
ISO Foreword
ISO (International Organization for Standardization) is a worldwide federation composed of national standardization bodies (ISO member bodies). The work of formulating international standards is usually completed by ISO's technical committees. Each member body has the right to participate in the work of a committee if it is interested in the standards proposed by a technical committee. International organizations (official or unofficial) that maintain contact with ISO () can also participate in the relevant work. In the field of electrotechnical standardization, ISO maintains a close cooperative relationship with the International Electrotechnical Commission (IEC). The draft international standards formally adopted by the technical committee are submitted to the member bodies for voting. The international standards must be approved by at least 75% of the member bodies participating in the voting before they can be officially released. International Standard ISO11138-1 was formulated by ISO/TC198 Technical Committee on Sterilization of Health Products.
ISO11138 includes the following parts under the general title "Sterilization of medical products - Biological indicators": Part 1: General
- Part 2: Biological indicators for cyclopentane sterilization - Part 3: Biological indicators for mixed thermophilic bacteria Annexes A, B, C, D, E and F are integral parts of this part of ISO11138. 605
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GB18281.1—2000
This standard specifies the general requirements for the production, labeling and performance of biological indicators intended for monitoring sterilization cycles (see Chapter 1). The specified steps and methods should be implemented by trained and appropriate personnel. Biological indicators shall not be used in any process not specified by the manufacturer on the label. Improper use of biological indicators may produce misleading results. Biological indicators should be used in conjunction with physical and/or chemical monitoring methods to demonstrate the efficacy of the sterilization process. Regardless of the results obtained with the biological indicator, or if the sterilization process base/chemical variables are outside the specified range, the sterilization process should be considered to have failed to achieve the expected results.
The performance of biological indicators can be affected by the storage environment before use, the method of use, or the technology used after exposure to the sterilization process. Therefore, storage and use should be carried out in accordance with the manufacturer's recommendations, and after sterilization, the biological indicator should be transferred to the specified recovery conditions as soon as possible. Biological indicators should not be used if they have exceeded the expiration date specified by the manufacturer. Biological indicators are used to monitor the effectiveness of the sterilization process and sterilization equipment. Such studies should be conducted by trained and appropriate personnel. 67
National Standard of the People's Republic of China
Sterilization of health care products
Biological indicators
Part 1: General
1 Scope
GB 18281. 1 : 2000
idtISo 11138-1:1994
This standard specifies the general requirements for the production, labelling and handling of biological indicators and test bacterial suspensions intended for confirmation and monitoring of sterilization cycles.
Note 1: Other parts of GB 18281-2000 (idl1SO11138) specify the specific requirements for biological indicators used for each specified sterilization process. This standard does not include requirements for products that have been directly inoculated with test bacteria or the recovery procedures of such inoculated products. This standard also does not specify the requirements for biological indicators using more than one strain or species on a carrier. 2 Referenced Standards
The provisions contained in the following standards constitute the provisions of this standard through reference in this standard. When this standard was published, the versions shown were valid. All standards will be revised, and parties using this standard should explore the possibility of using the latest versions of the following standards. GB/T 7408—1994 Data elements and exchange formats - Date and time representation for information interchange (eg VISO 8601: 1988) GB 18281.2.2000 Biological indicators for sterilization of health care products - Part 2: Biological indicators for ethylene oxide sterilization (idt IS0 11138-2, 1994)
GB 18281.3—2000
Biological indicators for sterilization of health care products - Part 3: Biological indicators for moist heat sterilization (idt IS0 11138.3: 1995)
GB/T 19002—1994 Quality system - Quality assurance model for production, installation and service (idt ISO 9002.1994) 3 Definitions
The following definitions are used in all parts of GB 18281--ISO 11138. 3.1 Biological indicator biological Indicatur (BI) is a contaminated carrier that has a certain resistance to specific sterilization treatment and is packed in the inner packaging and is available for use. Note 2: "Indicator" refers to various forms of indicator devices including indicator cards, sheets, strips, tapes, groups, etc. 3.2 Carrier rarricr
Support material coated with test bacteria.
3.3 Primary pack
A system that protects the inoculated carrier from damage and contamination without hindering the penetration of sterilizing agents. 3.4 Secondary pack
A packaging system containing packaged biological indicators for transportation and storage. 3.5 Inoculated carrier A carrier with a specified number of test bacteria on the shelf, approved by the State Quality and Technical Supervision on December 13, 2000 608
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3.6 Test organism test.organism
Microorganism used to prepare the inoculated carrier.
GB 18281. 1—2000
3.7 Viable test organism count The number of viable test organisms in a unit volume of bacterial suspension or on a carrier contaminated with bacteria is obtained by counting the number of individual colonies grown under the required culture conditions.
3.8 Inactivation
The loss of the ability of the test organism to germinate, grow and/or proliferate under the specified culture conditions. 3.9 Culture conditions The conditions that promote the germination, growth and/or proliferation of the test organism as specified by the producer, including the growth medium, culture time and temperature. 3.10 Recognized culture collection refers to an internationally recognized culture collection organization that preserves microorganisms in accordance with the Budapest Convention, patent laws and regulations. 3.11) Value I) value; Derimal reduction value The time or radiation absorbed dose required to inactivate 90% of the test organism under the specified conditions. 3.12 Survivor curve survivor Curve represents the curve graph of the relationship between the resistance or survival of the test bacteria and the exposure enhancement under specified conditions. 3.13 Process monitoring equipment process challenge device is an object that simulates the most unfavorable sterilization state of the sterilized items by the sterilization factor. Note 3: The composition of the process monitoring device can be to place the biological indicator in the most difficult part for the sterilization factor to reach. Note 4: The design of the process monitoring equipment should be based on the type of items to be sterilized and the sterilization steps. Biological indicators should not interfere with the performance of the items being tested. Note 5: The process monitoring equipment can be a dye carrier instead of a biological indicator. 3.14 Colony-forming unit (CFU) is a living microbial community visible to the naked eye that is produced by one or more cells. 3.15 Self-contained hialagicel indicator A biological indicator whose inner packaging contains the culture medium required for bacterial recovery and growth. 3. 16 Survival-kill winduw The exposure degree at which the biological indicator transitions from complete bacterial growth (survival exposure) to complete non-bacterial growth (killing exposure) when subjected to fire sterilization treatment under specified conditions.
3.17 Nominal population (dust producer) Nominal number of microorganisms
Note 6: The actual number of microorganisms may be different from the calibrated total number of microorganisms due to differences in the accuracy of the infection and resuscitation methods. 3.18 Resistometer
Special equipment for measuring resistance in order to produce a specified combination of physical and chemical changes in the sterilization process under defined conditions: 4 Production, operation and labeling requirements
4.1 Production control and quality system
4.1.1 All work required for this standard must be controlled in accordance with the quality control system that meets the requirements of GB/T 19002. 4.1.2 Records of the source of manufactured components must be kept. The manufactured components should include all materials and components that are mixed into or directly contact the test bacterial suspension, infection carrier or biological indicator.
4.1.3 The final product (bacterial suspension, infection carrier or biological indicator) provided by the manufacturer must be free of foreign bacteria. A large amount of foreign bacteria may impair the efficacy of the product. This must be confirmed, controlled, monitored and recorded during the production process. 4.2 Test bacteria
4.2-1 The test bacteria must be a strain that is easy to handle without special containers and equipment. 69
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4.2.2 The test bacteria must be a specified strain stored in a recognized strain repository and must be carefully identified by referring to the strain storage number. 4.2.3 When it is necessary to use a test bacteria that is not stored in a recognized strain repository, the producer must be responsible for depositing the special strain in a recognized strain repository.
4.2.4 The initial inoculum of each batch of test bacterial suspension must: a) be traceable to a standard strain stored in a recognized strain repository, and b) be verified for its species and purity
The method for storing the specified test bacteria strain should ensure that the culture is not contaminated and will not cause changes in its inherent properties. Each test bacteria has a specific identification experiment, and the manufacturer should provide relevant information and be verified. 4.3 Test bacterial suspension
4.3.1 The manufacturer must specify the culture medium and culture conditions for preparing the test suspension. These conditions can consistently produce test bacterial suspensions that meet the operational requirements of (J318281.1-1ISO11138-), and meet the special operational requirements of GB18281.2--IS011138-2 and GB 18281.3IS011138-3 respectively according to the situation. 4.3.2 The collection (culture) and subsequent treatment methods must ensure that the inoculation solution used for the carrier does not contain culture medium residues that may have an adverse effect on the efficacy of the inoculated carrier or biological indicator. If the manufacturer has demonstrated that culture medium residues will not have an adverse effect on the efficacy of the inoculated carrier or biological indicator, this requirement may be waived.
4.3.3 In order to be able to trace the strains used in the production of biological indicators and test bacterial suspensions to the strain collection, the manufacturer of the test bacterial suspension and (or) biological indicator must keep records that meet the requirements. 4.3.4 If the test bacterial suspension is used to prepare a contaminated carrier or contaminated product, each container or packaging box (box) containing the test bacterial suspension must be accompanied by the following instructions:
a) Name of the test suspension:
b) Provide the name or abbreviation of the strain repository of the test bacteria and the strain code of the strain; c) The nominal volume of the bacterial suspension is in milliliters (if it is not a suspension, it is in grams); d) A special code for tracing the production history: c) The number of viable cells of the test bacteria per liter of bacterial suspension; 1) Storage conditions of the sample:
g) Indicate the expiration date or validity period in the manner specified in GB/T 7408 (i.e.: X year × month × month); h) The name, trademark, address or other identification method of the producer; i) Disposal method
4.3.5 If required by the purchaser, the producer must provide detailed information on the resistance and performance characteristics of the test bacterial suspension, and these data must be measured by methods recognized by both the purchaser and the producer. 4.3.6 The producer must specify the storage conditions and expiration date of the test bacteria. These conditions must be monitored during storage. These conditions must ensure that the test bacterial suspension can always meet the operational requirements of (GB1318281.1-ISO11138-1), and meet the special operational requirements of GB18281.21SO11138-2 and GB1828].3-ISO11138-3 as appropriate. 4.3.7 The bacterial suspension must be counted for viable bacteria of the test bacteria. When the user requires the growth index of the test bacteria, the viable count should be expressed as a percentage of the total number of bacteria obtained by microscopic examination. 4.3.8 When the producer transfers the test bacterial suspension to a third party, It must be ensured that the storage is carried out under controlled conditions similar to the specified storage conditions (see 4-3.6)
4.4 Carrier, inner packaging and design
4.4.1 The carrier and inner packaging must not contain any contaminants (physical, chemical or microbiological) that may adversely affect the use of the biological indicator
4.4.2 The carrier and inner packaging must not be damaged during the sterilization process, otherwise the performance of the contaminated carrier will be adversely affected.
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The carrier in the inner and outer packaging should withstand transportation and be delivered to the user without damage. The design of the carrier and (or) inner packaging should meet the following requirements: a) During storage, transportation and handling, the loss of the initially infected test bacteria can be minimized; b) It is suitable for use as a component of the sterilization process test device. 4.4.3 Compliance with the requirements of 4.4.2 must be verified by observing the extent to which the carrier and inner packaging are exposed to extreme changes during the sterilization process and the degree of chemical and physical changes.
Go to 7: These ranges are found in other relevant parts of this standard. 4.4.4 During the sterilization process and after sterilization, the carrier and inner packaging must not retain or release any substances into the culture medium, otherwise it will inhibit the growth of a small number of surviving test bacteria under culture conditions. The test should be carried out in accordance with Appendix F to check whether it meets this requirement. 4.4.5 If the purchaser requires, the manufacturer must provide the maximum and minimum values of each carrier size. 4.5 Infected carrier
4.5.1 When preparing a batch of infected carriers, only the same strain of test bacteria can be used. 4.5.2 The preparation of the infected carrier must be to stain the test bacteria suspension on the carrier and then dry it under controlled conditions. 4.5.3 The carrier infection conditions must be specified, confirmed and controlled to ensure that the infected carrier is not contaminated by other bacteria other than the test bacteria, otherwise it will have an adverse effect on the use of the product specified in GL18281.2--ISO11138-2 and GB18281.3--ISO11138-3. 4.5.4 In the same batch of biological indicators, the total number of test bacteria marked on each infected carrier must be the same. 4.5.5 The dust manufacturer must specify the storage conditions and expiration date of the infected carrier. These conditions must be monitored during storage and must consistently comply with the operating requirements of GB18281.1-ISO11138-1 and, depending on the situation, the special operating requirements of GB18281.2-ISO11138-2 and CB18281.3-1SO11138-3 for contaminated carriers. 4.5.6 When contaminated carriers are packaged as biological indicators, the packaging method must not affect the total number of test bacteria and its performance of the single carrier.
4.5.7 Each batch of contaminated carriers must be accompanied by the following instructions: a) marked "contaminated carrier"
b) the name of the test bacteria;
c) instructions for use, especially data on the culture medium and culture conditions for the test of live bacteria after sterilization; d) the name of the strain storage bank of the test bacteria and the number of the strain; e) the number of test bacteria for each contaminated carrier;
b) the batch number or special code for traceability of production history; g) data on the resistance characteristics of the contaminated carrier to the applicable sterilization process, including test conditions and methods for determining the characteristics; h) the number of contaminated carriers in the outer packaging; i) recommended storage conditions; j) the expiration date of the contaminated carrier indicated in the manner specified in GB/T 7408 (i.e.: × year × month ×); k) the name, trademark, address or other identification method of the manufacturer; 1) the sterilization process applicable to the contaminated carrier:
i) the disposal sequence.
4.6 Biological indicators
4.6.1. Individual bacterial carriers must be placed in the inner packaging as biological indicators. 4.6.2 The design, construction and confirmation of the inner packaging must ensure that the biological indicators in the inner packaging meet the requirements of GB18281.2-ISO111382 and GB18281.3-ISO11138-3 respectively. 4.6.3 The design, construction and confirmation of the inner packaging must ensure that the biological indicator and the carrier can avoid contamination and loss of the infected test bacteria on the carrier during storage and transportation according to the manufacturer's method. 4.6.4 The environmental conditions of the inner packaging must be specified, confirmed and controlled to ensure that the carrier is not contaminated by other bacteria other than the test bacteria. Otherwise, it will have an adverse effect on the use of the product specified in GB 18281.2-ISO11138-2 and GB 18281.3ISO 11138-3.
4.6.5 The inner packaging to be used must be confirmed. It should meet the requirements of 4.4 of this standard.
4.6.6 The inner packaging of each biological indicator must be marked with the following information: a) the name of the test bacteria;
b) the batch number of the biological indicator;
c) the expiration date of the biological indicator in the manner specified in GB/T7408 (i.e.: × year × month × day);
d) the bacterial count of the biological indicator tablet;
e) the name, trademark, address and other identification methods of the manufacturer. 4.6.7 Biological indicators must be placed in outer packaging for transportation and storage. 4.6.8 The outer packaging must indicate the following: a) the name of the "biological indicator"; b) the inner packaging specified in 4.6.6: c) the name of the strain repository providing the test bacteria and the strain number; d) the number of test bacteria for each biological indicator is the same as the number measured for the batch of infected carriers; e) the number of biological indicators in the outer packaging; f) recommended storage conditions; g) the resistance of the test bacteria on the infected carriers in the inner packaging, including the test conditions and the method for determining the characteristics; h) instructions for use, especially data on the culture medium and culture conditions used to resuscitate the test activities after sterilization; and i) disposal methods.
4.6.9 Each outer package must contain a copy of the certificate of conformity for the batch of indicators, the contents of which must include: a) the contents specified in 4.6.8; b) the characteristic data of the resistance of the biological indicator to the sterilization process applicable to the monitoring of contaminated carriers; and c) reference to this standard GB18281.1ISO 11138-1 and other relevant international standards: 4.6.10 Each outer package must contain instructions for the user to handle and resuscitate the biological indicator, the contents of which must specify:) the biological indicator must be stored under the conditions specified by the manufacturer; b) a batch of biological indicators must not be used beyond the expiration date specified by the manufacturer; c) after the dust indicator has undergone the sterilization procedure being tested, the test bacteria recovery count must be tested within the time specified by the manufacturer; d) when testing the test bacteria recovery count of the biological indicator, the method and conditions specified by the manufacturer must be used. If other methods are used, they must be confirmed.
4.7 Self-contained biological indicators
4.7.1 Self-contained biological indicators must meet all the requirements of this standard. 4.7.2 Self-contained biological indicator systems should be tightly packed and placed in outer packaging to withstand transportation and handling, and not damaged when used. The design of self-contained biological indicator systems should: a) minimize the loss of initially infected test bacteria during transportation and handling; b) be suitable as components for monitoring devices for sterilization processes. 4.7.3 During and after sterilization treatment, the materials of the self-contained biological indicator system must not retain or release any substances that can inhibit the growth of a small number of surviving test bacteria under culture conditions (see Appendix F). 5 Resistance determination
5.1 Resistance test requirements
5.1.1 Each batch of biological indicators must be tested to demonstrate whether its resistance meets the use requirements specified in GB18281.2ISO11138-2 and 612
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GI3 18281. 3—-ISO 11138-3. 5.1.2 Resistance test (see 5.4 and 5.5) must include the determination of viable counts in the recovery test and the determination of resistance by two or more of the following methods:
a) Determination of D value by establishing a survival curve; b) Determination of D value by partial negative analysis; c) Calculation of survival-kill interval using the measured D value and verification of the survival-kill reaction characteristics. 5.1.3 The values obtained from these tests must be within the ranges specified in GB18281.2-IS011138-2 and GR18281.3-IS011138-3 respectively. At least two of the above data shall be marked on the label of the outer packaging (see 4.6.8) and the certificate of conformity of each batch of contaminated carriers (see 4.5.7).
Note 8: Some critical parts of this standard may require additional test items, such as GB18281.3IS011138-3 biological indicators for moist heat and fire bacteria, with additional test Z value.
5.2 Calculation of survival-kill interval
The survival-kill interval must be calculated using one of the D values measured in Appendix D and the following formula: survival time or survival dose ≥ D value × (IgN.-2) Killing time or killing dose ≤ D value × (IgV.·4) Note: N is the number of test viable bacteria labeled on each carrier. 5.3 Determination of viable count in recovery test
The viable count in the recovery test shall be determined in accordance with Appendix A. 5.4 Determination of D value
5.4.1 The data required for calculating the D value of the biological indicator must be determined in accordance with Appendix R (i.e., establishing a survival curve by direct counting) and (or) Appendix C (partial negative analysis method or most likely number [MPN] method). 5.4.2 The D value must be calculated in accordance with Appendix I3 and (or) Appendix T). 5.4.3 Other methods for analyzing partial negative data may be used and must be proven to be equivalent to the standard method. 5.5 Survival-Killing Reaction Determination
The survival-killing reaction characteristics must be determined and verified according to Appendix E. 613
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Appendix A
(Subject Appendix)
Determination of the number of viable test bacteria
A1 The viable test bacteria revived on the infected carrier must be checked according to A2~~A4 or other methods. If other methods are used, the relationship with the standard method must be understood.
A2 At least four infective carriers shall be used for each batch or exposure. Each infective carrier shall be placed in a certain volume of culture medium and the test bacteria shall be eluted from the infective carrier by a specific method (such as shaking with glass beads, grinding in a homogenizer, using ultrasound or other appropriate methods).
A3 The test bacterial suspension shall be diluted with an appropriate bacterial diluent so that the bacteria inoculated on each plate medium F produce between 30 and 300 CFU. Then appropriate aliquots of the suspension shall be mixed with melted liquid agar medium or spread on plates with solid culture medium. The manufacturer of the biological indicator shall identify or provide a suitable recovery medium and/or provide all data for the preparation of the medium.
A4 The sample plates shall be incubated at the temperature and time specified by the manufacturer. Note 1: - For thermophiles, the temperature shall be 5℃~60℃ for not less than 48 hours, and for thermophiles, the temperature shall be 30℃-37℃ for not less than 48 hours. 11: If the culture temperature is too high, the culture medium will dry out and affect the growth of bacteria. A5 After the appropriate period of culture, the colony forming units on the plate must be counted, and then the average number of test live bacteria on each contaminated carrier must be calculated.
Appendix B
(Appendix of the standard)
Survival curve method
Note 12: The ideal survival curve is linear in the entire inactivation range, but in reality it often deviates. However, its linearity must be maintained within an acceptable range. When the number of surviving bacteria is greater than 5×10, the survival curve constructed by the direct counting method can be used to determine its resistance. When the number of surviving bacteria is less than 5×10, the maximum energy number (MPN) method should be used to determine its resistance. If the D values measured by the two methods are well correlated, the established inactivation survival curve is particularly serious.
1 The sample must be exposed to the specified exposure conditions in stages, and the exposure range should be indicated. The time or dose of each exposure stage should be different from the previous exposure time or dose. The difference is equal. Note 13: For detailed information on the requirements for the equipment used for exposure, see GR18281.2IS11138-2 and GB18281.3--I50 11138-3,
B2 There must be at least 5 exposures, including:
a) In one exposure, the sample is not treated with sterilizing factors (sterilizing factors may not be used or non-lethal gases may be used instead); b) At least one exposure reduces the total number of viable bacteria to no more than 0.01% of the initial inoculum. B3 In each exposure, the number of infective carriers used in each measurement must not be less than 4, and each exposure must be from the same number of samples.
B4 Within 2 hours after each exposure, the sample must be treated to allow the test bacteria to fall off the carrier, and the viable bacteria must be counted using the culture conditions and methods specified by the manufacturer.
B5 Use the common logarithm of the total number of surviving bacteria obtained to plot time (min) or dose, and use the least squares method for regression analysis to determine the best linear curve. The survival data points within the 0.5log range of the original bacterial count should not be included in the regression analysis. The negative reciprocal value of the slope of the calculated straight line is equal to the D value expressed in minutes or absorbed radiation dose. B6 The linear correlation coefficient of the tear line of the viable bacteria obtained must not be less than 0.8. 614
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Appendix'
(Appendix with difficulty)
Partial negative analysis method or the maximum probable number (MPV) method of continuous determination of D values by the limited Spearman-Karber method Note 14: Partial negative data are usually used to calculate the characteristics of the test bacteria's response to the sterilization process by the maximum probable number (MPN) method. C1 The test specimens should be exposed to the specified exposure conditions in stages. Except for time or dose, other process variables should be carefully determined. Each exposure must use no less than 20 identical specimens. The time or dose of each exposure level should be different from the previous exposure time or dose, but the difference is equal. Each exposure must use the same specimens that have been the same for several months. Note 15: For details on the equipment operation requirements required for this clause, refer to GB 18251.2ISO 11138-2 and GB 18281.3IS( 11138-3.
2 Within 2 hours after each exposure, each infected carrier should be aseptically transferred to a test tube containing an appropriate amount of sterile culture medium. The same volume of culture medium must be used for each sample. If the producer uses the culture medium as part of the biological indicator, the producer's culture requirements must be followed. The producer of the biological indicator should identify or provide a suitable recovery medium and/or provide all data for the preparation of the culture medium.
C3 The infected carrier is incubated at the temperature recommended by the producer. After incubation for the incubation time recommended by the producer, check the growth of the test bacteria. Depending on the characteristics of the test bacteria, if the broth medium becomes turbid, or there is bacterial growth on the surface of the broth, or there is precipitation at the bottom of the test tube, it indicates bacterial growth.
C4 The result is recorded as the ratio of the number of infected carriers that can recover the test bacteria to the number of infected carriers exposed to a sublethal dose each time. Appendix D
(Recommended Appendix)
Calculation of D value D1 using the limited Spearman-Karher method Calculation of D value
Note 16: The limited Spearial1-Karber method requires that the intervals between successive exposures be fixed, and that the number of months (n) used in the same test column be the same for each exposure.
D7.1 The selected exposure interval or level (1.UU) must cover the entire partial negative area, and the initial exposure (1) must show no samples to be sterile or r = 0. The final exposure (U) must be sterile for all samples or r = n. The test is valid only if no samples are negative in the exposure before (r0), and all samples are negative in the exposure after U (r-n), and there are at least two partial reaction intervals between U and U (f ≥ 0 and n). The average exposure to sterility (U) or the limited Spearman-Karber method is used as follows: x
Wu:
Ua—average exposure to sterility (limited Spearinan-Karber method estimation), Uk
The exposure that initially shows sterility of all samples, n; i—exposure time or dose;
—time or dose interval between exposures (see Note 16),
Number of bio-indicator samples at each exposure, number of samples reaching sterility at each exposure t
The longest exposure without a negative bio-indicator, 0; U.-——the previous exposure,
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Art r: — the sum of the number of sterile biological indicators (r) in all exposures between Ui and Uh-:. D1.2 Calculate the value of
by the following formula:
Uk — the average exposure to sterility;
IgN, + 0.250 7
the average number of viable spores on each biological indicator measured by the total viable count, Nu
D2 Calculate the variance, standard deviation and confidence interval of U by the finite Spearman-Karbher method. Note 17: The finite Spearman-Karbher method makes it possible to calculate the variance of U, and then derive its standard deviation and confidence interval. D2.1 Calculate the variance of U according to the following formula: (U)Tisk
Where:
Time or dose interval between exposures:
Number of days for each exposure sample;
r-Number of each exposure sample;
2n(nr)-The sum of the values of each exposure between U: and Uc1 (n-r.). D2.2 Calculate the standard deviation (STD) of Uk according to the following formula: SDek = VVe
D2.3 Use the upper and lower credible intervals of Uk (the credible interval of Ua is Uk=2SD, iP=0.95) to calculate the upper and lower credible intervals of D value. Formula:
U lower limit: value
upper limit; force value -
1gN.0. 250 7
IgN,-0. 250 ?
Appendix E
(Standard Appendix)
Survival-killing reaction characteristics
Note 18: Monitoring the survival-killing reaction characteristics of some biological monitors can provide another way to ensure the consistent performance of all indicators in the batch. E1 The D value calculated by establishing a survival curve or partial negative analysis (see Appendix B and Appendix D) can be used to calculate the survival and killing time or dose.
E2 At least 50 identical samples must be used to confirm the survival time or survival dose and the ignition time or ignition dose. E3 At least 50 identical samples should be used in the determination of survival time or survival dose and killing time or killing dose. The exposure time or dose for survival of the test bacteria on each sample indicated indicates the survival characteristic. The exposure time or dose for killing all bacteria on each sample indicated indicates the killing characteristic. E4 survival-killing reaction characteristics must be measured using a biological indicator resistance tester. The survival and killing time or dose are expressed as: survival time or survival dose (1gN.2) × D value killing time or killing dose ≤ (lgN.4) × 1) value Note 19: The number of samples used in each exposure should be based on the volume and operating characteristics of the dust indicator resistance tester used. When measuring the survival and killing time or dose, several exposures may be required to test whether the total number of samples meets the requirements. 616
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Appendix'
(Difficult Appendix)
Partial Negative Analysis Method or Maximum Probable Number (MPV) Method for Continuous Determination of D Values by Limited Spearman-Karber Method Note 14: Partial negative data are usually used to evaluate the characteristics of the test bacteria's response to the sterilization process by the maximum probability number (MPN) method. C1 The test specimens shall be exposed to the specified exposure conditions in stages. Except for time or dose, other process variables shall be carefully determined. Each exposure shall use no less than 20 identical test specimens. The time or dose of each exposure stage shall be different from the previous exposure time or dose, but the interval shall be equal. Each exposure shall use the same test specimens that have been the same for several months. Note 15: For details on the equipment operation requirements for exposure, refer to GB 18251.2ISO 11138-2 and GB 18281.3IS( 11138-3.
2 Within 2 hours after each exposure, each infected carrier should be aseptically transferred to a test tube containing an appropriate amount of sterile culture medium. The same volume of culture medium must be used for each sample. If the producer uses the culture medium as part of the biological indicator, the producer's culture requirements must be followed. The producer of the biological indicator should identify or provide a suitable recovery medium and/or provide all data for the preparation of the culture medium.
C3 The infected carrier is incubated at the temperature recommended by the producer. After incubation for the incubation time recommended by the producer, check the growth of the test bacteria. Depending on the characteristics of the test bacteria, if the broth medium becomes turbid, or there is bacterial growth on the surface of the broth, or there is precipitation at the bottom of the test tube, it indicates bacterial growth.
C4 The result is recorded as the ratio of the number of infected carriers that can recover the test bacteria to the number of infected carriers exposed to a sublethal dose each time. Appendix D
(Recommended Appendix)
Calculation of D value D1 using the limited Spearman-Karher method Calculation of D value
Note 16: The limited Spearial1-Karber method requires that the intervals between successive exposures be fixed, and that the number of months (n) used in the same test column be the same for each exposure.
D7.1 The selected exposure interval or level (1.UU) must cover the entire partial negative area, and the initial exposure (1) must show no samples to be sterile or r = 0. The final exposure (U) must be sterile for all samples or r = n. The test is valid only if no samples are negative in the exposure before (r0), and all samples are negative in the exposure after U (r-n), and there are at least two partial reaction intervals between U and U (f ≥ 0 and n). The average exposure to sterility (U) or the limited Spearman-Karber method is used as follows: x
Wu:
Ua—average exposure to sterility (limited Spearinan-Karber method estimation), Uk
The exposure that initially shows sterility of all samples, n; i—exposure time or dose;
—time or dose interval between exposures (see Note 16),
Number of bio-indicator samples at each exposure, number of samples reaching sterility at each exposure t
The longest exposure without a negative bio-indicator, 0; U.-——the previous exposure,
GB 18281.1 :2000
Art r: — the sum of the number of sterile biological indicators (r) in all exposures between Ui and Uh-:. D1.2 Calculate the value of
by the following formula:
Uk — the average exposure to sterility;
IgN, + 0.250 7
the average number of viable spores on each biological indicator measured by the total viable count, Nu
D2 Calculate the variance, standard deviation and confidence interval of U by the finite Spearman-Karbher method. Note 17: The finite Spearman-Karbher method makes it possible to calculate the variance of U, and then derive its standard deviation and confidence interval. D2.1 Calculate the variance of U according to the following formula: (U)Tisk
Where:
Time or dose interval between exposures:
Number of days for each exposure sample;
r-Number of each exposure sample;
2n(nr)-The sum of the values of each exposure between U: and Uc1 (n-r.). D2.2 Calculate the standard deviation (STD) of Uk according to the following formula: SDek = VVe
D2.3 Use the upper and lower credible intervals of Uk (the credible interval of Ua is Uk=2SD, iP=0.95) to calculate the upper and lower credible intervals of D value. Formula:
U lower limit: value
upper limit; force value -
1gN.0. 250 7
IgN,-0. 250 ?
Appendix E
(Standard Appendix)
Survival-killing reaction characteristics
Note 18: Monitoring the survival-killing reaction characteristics of some biological monitors can provide another way to ensure the consistent performance of all indicators in the batch. E1 The D value calculated by establishing a survival curve or partial negative analysis (see Appendix B and Appendix D) can be used to calculate the survival and killing time or dose.
E2 At least 50 identical samples must be used to confirm the survival time or survival dose and the ignition time or ignition dose. E3 At least 50 identical samples should be used in the determination of survival time or survival dose and killing time or killing dose. The exposure time or dose for survival of the test bacteria on each sample indicated indicates the survival characteristic. The exposure time or dose for killing all bacteria on each sample indicated indicates the killing characteristic. E4 survival-killing reaction characteristics must be measured using a biological indicator resistance tester. The survival and killing time or dose are expressed as: survival time or survival dose (1gN.2) × D value killing time or killing dose ≤ (lgN.4) × 1) value Note 19: The number of samples used in each exposure should be based on the volume and operating characteristics of the dust indicator resistance tester used. When measuring the survival and killing time or dose, several exposures may be required to test whether the total number of samples meets the requirements. 616
TKAoNiKAca-1--2000
Appendix'
(Difficult Appendix)
Partial Negative Analysis Method or Maximum Probable Number (MPV) Method for Continuous Determination of D Values by Limited Spearman-Karber Method Note 14: Partial negative data are usually used to evaluate the characteristics of the test bacteria's response to the sterilization process by the maximum probability number (MPN) method. C1 The test specimens shall be exposed to the specified exposure conditions in stages. Except for time or dose, other process variables shall be carefully determined. Each exposure shall use no less than 20 identical test specimens. The time or dose of each exposure stage shall be different from the previous exposure time or dose, but the interval shall be equal. Each exposure shall use the same test specimens that have been the same for several months. Note 15: For details on the equipment operation requirements for exposure, refer to GB 18251.2ISO 11138-2 and GB 18281.3IS( 11138-3.
2 Within 2 hours after each exposure, each infected carrier should be aseptically transferred to a test tube containing an appropriate amount of sterile culture medium. The same volume of culture medium must be used for each sample. If the producer uses the culture medium as part of the biological indicator, the producer's culture requirements must be followed. The producer of the biological indicator should identify or provide a suitable recovery medium and/or provide all data for the preparation of the culture medium.
C3 The infected carrier is incubated at the temperature recommended by the producer. After incubation for the incubation time recommended by the producer, check the growth of the test bacteria. Depending on the characteristics of the test bacteria, if the broth medium becomes turbid, or there is bacterial growth on the surface of the broth, or there is precipitation at the bottom of the test tube, it indicates bacterial growth.
C4 The result is recorded as the ratio of the number of infected carriers that can recover the test bacteria to the number of infected carriers exposed to a sublethal dose each time. Appendix D
(Recommended Appendix)
Calculation of D value D1 using the limited Spearman-Karher method Calculation of D value
Note 16: The limited Spearial1-Karber method requires that the intervals between successive exposures be fixed, and that the number of months (n) used in the same test column be the same for each exposure.
D7.1 The selected exposure interval or level (1.UU) must cover the entire partial negative area, and the initial exposure (1) must show no samples to be sterile or r = 0. The final exposure (U) must be sterile for all samples or r = n. The test is valid only if no samples are negative in the exposure before (r0), and all samples are negative in the exposure after U (r-n), and there are at least two partial reaction intervals between U and U (f ≥ 0 and n). The average exposure to sterility (U) or the limited Spearman-Karber method is used as follows: x
Wu:
Ua—average exposure to sterility (limited Spearinan-Karber method estimation), Uk
The exposure that initially shows sterility of all samples, n; i—exposure time or dose;
—time or dose interval between exposures (see Note 16),
Number of bio-indicator samples at each exposure, number of samples reaching sterility at each exposure t
The longest exposure without a negative bio-indicator, 0; U.-——the previous exposure,
GB 18281.1 :2000
Art r: — the sum of the number of sterile biological indicators (r) in all exposures between Ui and Uh-:. D1.2 Calculate the value of
by the following formula:
Uk — the average exposure to sterility;
IgN, + 0.250 7
the average number of viable spores on each biological indicator measured by the total viable count, Nu
D2 Calculate the variance, standard deviation and confidence interval of U by the finite Spearman-Karbher method. Note 17: The finite Spearman-Karbher method makes it possible to calculate the variance of U, and then derive its standard deviation and confidence interval. D2.1 Calculate the variance of U according to the following formula: (U)Tisk
Where:
Time or dose interval between exposures:
Number of days for each exposure sample;
r-Number of each exposure sample;
2n(nr)-The sum of the values of each exposure between U: and Uc1 (n-r.). D2.2 Calculate the standard deviation (STD) of Uk according to the following formula: SDek = VVe
D2.3 Use the upper and lower credible intervals of Uk (the credible interval of Ua is Uk=2SD, iP=0.95) to calculate the upper and lower credible intervals of D value. Formula:
U lower limit: value
upper limit; force value -
1gN.0. 250 7
IgN,-0. 250 ?
Appendix E
(Standard Appendix)
Survival-killing reaction characteristics
Note 18: Monitoring the survival-killing reaction characteristics of some biological monitors can provide another way to ensure the consistent performance of all indicators in the batch. E1 The D value calculated by establishing a survival curve or partial negative analysis (see Appendix B and Appendix D) can be used to calculate the survival and killing time or dose.
E2 At least 50 identical samples must be used to confirm the survival time or survival dose and the ignition time or ignition dose. E3 At least 50 identical samples should be used in the determination of survival time or survival dose and killing time or killing dose. The exposure time or dose for survival of the test bacteria on each sample indicated indicates the survival characteristic. The exposure time or dose for killing all bacteria on each sample indicated indicates the killing characteristic. E4 survival-killing reaction characteristics must be measured using a biological indicator resistance tester. The survival and killing time or dose are expressed as: survival time or survival dose (1gN.2) × D value killing time or killing dose ≤ (lgN.4) × 1) value Note 19: The number of samples used in each exposure should be based on the volume and operating characteristics of the dust indicator resistance tester used. When measuring the survival and killing time or dose, several exposures may be required to test whether the total number of samples meets the requirements. 616
TKAoNiKAca-The test is considered valid. The average exposure (U) to sterility is calculated using the following formula or the limited Spearman-Karber method: x
Wu:
Ua—average exposure to sterility (limited Spearinan-Karber method estimation), Uk
the exposure that initially shows sterility of all samples, n; i—exposure time or dose;
—time or dose interval between exposures (see Note 16),
the number of bio-indicator samples at each exposure, the number of samples reaching sterility at each exposure, t
the longest exposure without a negative bio-indicator, 0; U.—the number of exposures before,
GB 18281.1:2000
techniquer:—the sum of the number of bio-indicators of sterility (r) in all exposures between Ui and Uh-:. D1.2 Calculate the value of (tt||) by the following formula:
Uk——the average exposure to sterility;
IgN, + 0.250 7
the average number of viable spores on each biological indicator measured by the total viable count, Nu
D2 Calculate the variance, standard deviation and confidence interval of U by the finite Spearman-Karbher method. Note 17: The finite Spearman-Karbher method makes it possible to calculate the variance of U, and then deduce its standard deviation and confidence interval. D2.1 Calculate the variance of U according to the following formula: (U)Tisk
Where:
Time or dose interval between exposures:
Number of days for each exposure sample;
r-Number of each exposure sample;
2n(nr)-The sum of the values of each exposure between U: and Uc1 (n-r.). D2.2 Calculate the standard deviation (STD) of Uk according to the following formula: SDek = VVe
D2.3 Use the upper and lower credible intervals of Uk (the credible interval of Ua is Uk=2SD, iP=0.95) to calculate the upper and lower credible intervals of D value. Formula:
U lower limit: value
upper limit; force value -
1gN.0. 250 7
IgN,-0. 250 ?
Appendix E
(Standard Appendix)
Survival-killing reaction characteristics
Note 18: Monitoring the survival-killing reaction characteristics of some biological monitors can provide another way to ensure the consistent performance of all indicators in the batch. E1 The D value calculated by establishing a survival curve or partial negative analysis (see Appendix B and Appendix D) can be used to calculate the survival and killing time or dose.
E2 At least 50 identical samples must be used to confirm the survival time or survival dose and the ignition time or ignition dose. E3 At least 50 identical samples should be used in the determination of survival time or survival dose and killing time or killing dose. The exposure time or dose for survival of the test bacteria on each sample indicated indicates the survival characteristic. The exposure time or dose for killing all bacteria on each sample indicated indicates the killing characteristic. E4 survival-killing reaction characteristics must be measured using a biological indicator resistance tester. The survival and killing time or dose are expressed as: survival time or survival dose (1gN.2) × D value killing time or killing dose ≤ (lgN.4) × 1) value Note 19: The number of samples used in each exposure should be based on the volume and operating characteristics of the dust indicator resistance tester used. When measuring the survival and killing time or dose, several exposures may be required to test whether the total number of samples meets the requirements. 616
TKAoNiKAca-The test is considered valid. The average exposure (U) to sterility is calculated using the following formula or the limited Spearman-Karber method: x
Wu:
Ua—average exposure to sterility (limited Spearinan-Karber method estimation), Uk
the exposure that initially shows sterility of all samples, n; i—exposure time or dose;
—time or dose interval between exposures (see Note 16),
the number of bio-indicator samples at each exposure, the number of samples reaching sterility at each exposure, t
the longest exposure without a negative bio-indicator, 0; U.—the number of exposures before, Www.bzxZ.net
GB 18281.1:2000
techniquer:—the sum of the number of bio-indicators of sterility (r) in all exposures between Ui and Uh-:. D1.2 Calculate the value of (tt||) by the following formula:
Uk——the average exposure to sterility;
IgN, + 0.250 7
the average number of viable spores on each biological indicator measured by the total viable count, Nu
D2 Calculate the variance, standard deviation and confidence interval of U by the finite Spearman-Karbher method. Note 17: The finite Spearman-Karbher method makes it possible to calculate the variance of U, and then deduce its standard deviation and confidence interval. D2.1 Calculate the variance of U according to the following formula: (U)Tisk
Where:
Time or dose interval between exposures:
Number of days for each exposure sample;
r-Number of each exposure sample;
2n(nr)-The sum of the values of each exposure between U: and Uc1 (n-r.). D2.2 Calculate the standard deviation (STD) of Uk according to the following formula: SDek = VVe
D2.3 Use the upper and lower credible intervals of Uk (the credible interval of Ua is Uk=2SD, iP=0.95) to calculate the upper and lower credible intervals of D value. Formula:
U lower limit: value
upper limit; force value -
1gN.0. 250 7
IgN,-0. 250 ?
Appendix E
(Standard Appendix)
Survival-killing reaction characteristics
Note 18: Monitoring the survival-killing reaction characteristics of some biological monitors can provide another way to ensure the consistent performance of all indicators in the batch. E1 The D value calculated by establishing a survival curve or partial negative analysis (see Appendix B and Appendix D) can be used to calculate the survival and killing time or dose.
E2 At least 50 identical samples must be used to confirm the survival time or survival dose and the ignition time or ignition dose. E3 At least 50 identical samples should be used in the determination of survival time or survival dose and killing time or killing dose. The exposure time or dose for survival of the test bacteria on each sample indicated indicates the survival characteristic. The exposure time or dose for killing all bacteria on each sample indicated indicates the killing characteristic. E4 survival-killing reaction characteristics must be measured using a biological indicator resistance tester. The survival and killing time or dose are expressed as: survival time or survival dose (1gN.2) × D value killing time or killing dose ≤ (lgN.4) × 1) value Note 19: The number of samples used in each exposure should be based on the volume and operating characteristics of the dust indicator resistance tester used. When measuring the survival and killing time or dose, several exposures may be required to test whether the total number of samples meets the requirements. 616
TKAoNiKAca-
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