培养基与试剂相关标准解读
发布时间：
2022-12-24
作者：
即用型液体培养基公司
培养基与试剂相关标准解读
微生物学检验培养基质量控制的意义
4789系列)都是以传统微生物学培养为基础的经典检验方法。培养基及相关配套试剂是微生物学常规检验方法必须采用的生化试剂，培养基及试剂质量会直接影响到检验结果的准确性，因此，必须做好培养基及试剂的质量控制。涉及食品微生物学检验的培养基种类繁多。近年来，随着市场经济的发展，商品化培养基的应用越来越普及。基于种种原因，它们的配方、用途、使用方法、保存方式各不相同，为实验室质量控制带来新问题。
目前，作为我国现行食品安全标准体系配套的强制性标准《食品安全国家标准食品微生物学检验》(GB1538.2—2016《培养基制备指南第2部分：培养基性能测试实用指南》进行[2，3]，但是，这些相关标准或规定受限于应用领域，作为微生物学检验培养基和试剂的质量控制依据也不完备，多数商品化培养基生产企业也未据此作为质量控制标准，而检验机构对自行配制的培养基进行质量控制的情况更少，导致培养基和试剂质量参差不齐。据以往相关调查，国产培养基的质量均有严重问题[4]。中国国家认证认可监督管理委员会网站通报的CNAS实验室现场评审在《服务和供应品的采购》这个环节的常见不符合项最多，主要是：未建立培养基和试剂质量控制程序、不能提供关键培养基技术性验收评估记录和不能提供实验室用水符合性评价记录，说明多数检验机构对于培养基和试剂的相关标准和规定的理解、执行都存在偏差。第1部分：实验室培养基制备质量保证通则》和SN/T1538.1—2016《培养基制备指南1133制定的SN/T11133—2014《食品、动物饲料和水的微生物学—营养培养基的制备、生产、存储和性能测试》或修改采用ISO1 现行的相关微生物学检验用培养基质量控制的标准或规定，除了2013版标准之外，CNAS—CL09：2013《检测和校准实验室能力认可准则在微生物检测领域的应用说明》规定实验室对于培养基和试剂的验收可参考ISO。
培养基和试剂质量控制的要求。
在中国国家认证认可监督管理委员会发布的《检验检测机构资质认定评审准则》4.5.6条款中，规定“检验检测机构应建立和保持选择和购买对检验检测质量有影响的服务和供应品的程序。明确服务、供应品、试剂、消耗材料的购买、验收、存储的要求”，就是要求“在中华人民共和国境内，向社会出具具有证明作用的数据、结果的检验检测机构”为保证采购物品和相关服务的质量，应当按制定的程序对试剂进行有效的控制和管理，以保证检验检测结果的质量[5]。虽然文件有规定，但是，具体如何执行，需要检验机构结合自身实际情况制定程序性文件。2013版标准只是食品安全国家标准，适用范围仅限于食品微生物学检验领域，而目前其他微生物学检验领域并没有适用的培养基和试剂质量控制标准，所以，建议所有涉及微生物学检验领域的检验机构都执行或参照执行2013版标准，制定适合本实验室的培养基和试剂质量控制程序性文件。
2013版标准的具体要求和存在问题
术语和定义
11133分为6类有所不同，例如，将选择性计数培养基分为选择性固体计数培养基和选择性液体计数培养基，明显就是画蛇添足。 本章节主要缺陷是定义描述不到位和前后术语不一致。同时，遗漏了诊断血清、测试纸片、生化鉴定试剂盒等常用微生物检验试剂。另外，标准将培养基和试剂分为11类，分类方法与ISO
缺少“设备和仪器”
作为一个既有检验方法又有判定依据的标准，缺少对于设备和仪器的规定，是不可想象的。
培养基及试剂质量保证
这个章节分为证明文件、贮存、培养基的实验室制备、培养基的使用、培养基的弃置五个部分。建议检验机构按照这个框架规定培养基和试剂的质量控制程序。这个章节存在的问题，主要是对培养基保质期规定太松、配制用水电阻率错误、pH测定温度不适用、培养基的弃置可操作性不强，而目前现场评审时，实验室被核查到的常见错误，除了培养基配制原始记录信息不全这个通病外，几乎所有实验室都没有做到“倾注琼脂量达到3mm以上厚度”，属于未认真学习标准规定。
质控菌株的保藏及使用
19489—2008等其他实验室质量控制管理规定要求不一致[2，6，7]，容易理解错误。总则》、GB食品微生物学检验4789.1—2016《食品安全国家标准 这个章节，除了使用术语与第二章规定不一致外，最重要的问题是描述和规定不够详尽，也未考虑生物安全相关标准要求，因此，与CNAS—CL09：2013、GB
培养基和试剂的质量要求
1.基本要求
本条规定供应商和制备者应进行培养基和试剂的20℃～25℃的pH和缓冲能力特性评估，前者与标准全文都在25℃测定pH的规定不符，后者则是全文都没有出现如何检验、如何判定的描述。
2.微生物学要求
概论规定实验室应“制定验收程序，如需进行验证，可按6.2执行”。对此条款，目前，多数检验机构并未理解，除了没有制定明确的培养基和试剂验收程序外，也不知道哪些培养基和试剂需要进行验证。其实，就是有第三方检验机构检验报告判定“达到附录D质量控制标准的要求”的，以及对检验结果没有直接影响的培养基和试剂，不需要验证。在“5.2.3.2测试菌株”条，使用了与第四章“质控菌株”不一样的术语。这个条款中，规定了根据培养基和试剂类型的不同，采用的测试菌株数量。在现场评审时，常见评审员要求实验室必须使用附录E里面的全部菌株进行测试或实验室仅用一株菌株进行选择性培养基测试等情况，都属于对标准理解错误。
培养基和试剂性能测试方法
总体来说，2016版标准考虑到我国中小型实验室进行全部培养基和试剂验证的工作量，并未一味照搬国际标准的要求，人性化规定了实验室可以使用半定量和定性测试方法验证培养基和试剂，有利于实验室执行并开展。不足之处是，本章节文字、语法及描述性错误太多，影响了标准的实施效果。
测试结果的记录
7.1条要求“培养基制造商或供应商应按客户的要求提供培养基常规信息和相关测试菌株生长特性信息。”与3.1.1条“生产企业提供的文件”属于重复规定。
7.2条要求“按照质量体系的要求，对所有培养基性能测试的数据归档并在有效期内进行适当的保存。”实际上，任何质量控制体系都没有规定必须对所有培养基进行性能测试，用于辅助检验用的培养基，如，菌种保存、实验室内部环境监控的那些培养基并不强制进行性能测试，只要对关键性培养基和试剂进行验证即可。
Standard interpretation of medium and reagent
Significance of microbiological examination of medium quality control
Series 4789) are all classical test methods based on traditional microbial culture. Medium and related auxiliary reagent is routine microbiological testing method must be used biochemical reagents, culture medium and reagent quality will directly affect the accuracy of test results, therefore, culture medium and the quality control of the reagents must be done. There are many kinds of media involved in food microbiological testing. In recent years, with the development of market economy, the application of commercial media is more and more popular. For various reasons, their formulations, USES, use methods and preservation methods are different, which bring new problems to laboratory quality control.
Currently, as a compulsory standard in China's current food safety standard system, food safety national standard food microbiology test (gb1538.2-2016) Medium performance test and practical guide to [2, 3], however, the relevant standards or provisions are limited by application field, as the quality control of microbiology test reagent and medium basis is not complete, most of the commercialization of media production enterprises is not as a quality control standard, accordingly the inspection institution to preparation of culture medium for quality control of less, leading to medium and reagents are of variable quality. According to previous relevant surveys, the quality of domestic media has serious problems [4]. The supervision and administration of the China national accreditation committee website report CNAS lab site assessment in the procurement of services and supplies at this stage the most common nonconformities, mainly: medium and reagent quality control program has not been established, is unable to provide the key medium technical acceptance evaluation records and cannot provide laboratory water conformity assessment records, shows that most of the inspection institution for the understanding of culture medium and the related standards and regulations of the reagents, execution, there is a deviation. Part 1: general principles for quality assurance laboratory culture medium preparation and SN/T1538.1-2016 "medium preparation guidelines set by the 1133 SN/T11133-2014" food, animal feed and water microbiology, nutrient medium preparation, production, storage, and the performance test, or modify the ISO1 current microbiology related media quality control standards or regulations, in addition to the 2013 version of the standard, CNAS - CL09: In 2013, the application description of the laboratory accreditation criteria for testing and calibration in the field of microbiological detection stipulated that the laboratory's acceptance of media and reagents could refer to ISO.
Quality control requirements for media and reagents.
Released in China national accreditation supervision and administration commission of the assessment criteria testing institutions aptitude 4.5.6 terms, provisions of "the examination institution shall establish and maintain affects the examination quality to select and purchase services and supplies. Specific services, supplies, reagents, consumable material purchase, acceptance, storage requirements ", is required to the society "within the territory of the People's Republic of China, has the effective evidence issued by the data, the results of the examination institutions" to ensure the quality of procurement of goods and related services, shall be formulated according to the procedure of reagents for effective control and management, to ensure the quality of the examination results [5]. Although there are provisions in the documents, it is necessary for the inspection authorities to formulate procedural documents according to their actual conditions. 2013 version of the standard is the national food safety standards, the applicable scope is limited to food microbiology test field, and the other field of microbiology test was not applicable medium and reagent quality control standards, therefore, recommended that all involved in the field of microbiology test inspection institutions are executed at or around the 2013 version of the standard, suitable for the laboratory reagent and medium quality control programmatic document.
Specific requirements and existing problems of the 2013 edition standard
Terms and definitions
For example, the selective count medium was divided into the selective solid count medium and the selective liquid count medium. The main drawback of this chapter is that the definition description is not in place and the terms are inconsistent. At the same time, the diagnostic serum, test paper, biochemical identification kit and other commonly used microbial test reagents were omitted. In addition, the standard divides media and reagents into 11 categories, classification methods and ISO
Lack of "equipment and equipment"
As a standard that has both inspection method and judgment basis, it is impossible to imagine the lack of provisions on equipment and instruments.
Medium and reagent quality assurance
This chapter is divided into five parts: proof document, storage, laboratory preparation of medium, use of medium and discard of medium. It is recommended that inspection bodies establish quality control procedures for media and reagents in accordance with this framework. Problems existing in this chapter, mainly for medium shelf life is too loose, mixed water resistivity error, shall not apply to the pH measurement temperature and medium disposed operability is not strong, and the site assessment, laboratory verification to common errors, in addition to the culture medium preparation records the information is not complete and the common fault, almost all the laboratory do not "pour into AGAR volume reached more than 3 mm thickness, is not seriously learning standards.
Storage and use of quality control strains
Other laboratory quality control regulations, such as 19489-2008, have inconsistent requirements [2,6,7] and are easy to understand and make mistakes. General rules ", GB food microbiology test 4789.1-2016 "the national food safety standards of this section, in addition to use the term inconsistent with the provisions of the second chapter, the most important problem is not enough detailed descriptions and regulation, also did not consider biological safety standard requirements, therefore, with CNAS - CL09:2013, GB
Quality requirements for medium and reagent
1. Basic requirements
Should be carried out the provisions of the supplier and the preparation of culture medium and reagents of 20 ℃ to 25 ℃ and buffering capacity of the pH value of the property evaluation, the former with the standard does not conform to that stipulated in full all at 25 ℃ was developed for the determination of pH, the latter is the full text is not how to test, how to determine the description.
2. Microbiological requirements
The introduction stipulates that the laboratory should "develop acceptance procedures, and if verification is required, it can be carried out in accordance with 6.2". At present, most inspection institutions do not understand this provision, and they do not know which media and reagents need to be verified except that there is no clear culture medium and reagent acceptance procedure. Actually, is the third party inspection agency inspection report to judge "to achieve the requirement of the appendix D quality control standard", and not directly influence the result of the test medium and reagents, don't need to test and verify. In the section "5.2.3.2 test strains", a different term is used than that used in chapter 4, "quality control strains". This provision specifies the number of test strains to be used depending on the type of medium and reagent. On site review, common judges require laboratory must use the inside of the appendix E all the strains tested or lab in only one strains tested selective medium, etc., all belong to understanding standard error.
Test method for performance of culture medium and reagent
2016 version of the standard, in general, considering our country small and medium-sized lab for all media and reagents to verify the amount of work, not blindly copy the requirements of international standard, humanized rules the lab can use half quantitative and qualitative test method validation of reagent and medium, is advantageous to the laboratory to execute and carry out. The disadvantage is that there are too many text, grammar and descriptive errors in this chapter, which affect the implementation effect of the standard.
Records of test results
7.1 requirements: "the medium manufacturer or supplier shall provide the conventional information of the medium and the relevant information on the growth characteristics of the tested strains as required by the customer." And 3.1.1 "documents provided by production enterprises" are duplicated.
7.2 "according to the requirements of the quality system, the data of all performance tests of the medium shall be filed and properly kept within the validity period." In fact, did not set any quality control system must be conducted for all media performance test, used for auxiliary test in culture medium, such as, species conservation, internal environmental monitoring laboratory performance testing of the medium is not mandatory, as long as the key media and reagents for validation.