ASTM D6499-2016 3769 Standard Test Method for The Immunological Measurement of Antigenic Protein in Natural Rubber and its Products《天然橡胶及其制品中抗原性蛋白质的免疫测量的标准试验方法》.pdf
《ASTM D6499-2016 3769 Standard Test Method for The Immunological Measurement of Antigenic Protein in Natural Rubber and its Products《天然橡胶及其制品中抗原性蛋白质的免疫测量的标准试验方法》.pdf》由会员分享,可在线阅读,更多相关《ASTM D6499-2016 3769 Standard Test Method for The Immunological Measurement of Antigenic Protein in Natural Rubber and its Products《天然橡胶及其制品中抗原性蛋白质的免疫测量的标准试验方法》.pdf(9页珍藏版)》请在麦多课文档分享上搜索。
1、Designation: D6499 16Standard Test Method forThe Immunological Measurement of Antigenic Protein inNatural Rubber and its Products1This standard is issued under the fixed designation D6499; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revi
2、sion, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () indicates an editorial change since the last revision or reapproval.1. Scope1.1 This test method covers an immunological method todetermine the amount of antigenic protein in natur
3、al rubber andits products using rabbit antisera specific for natural rubberlatex (NRL) proteins. This immunoassay procedure quantita-tively measures the level of antigenic latex proteins in solutionusing an inhibition format. The samples may include glove orother rubber product extracts which have b
4、een collected inorder to measure the latex protein levels.Although this methoddetects antigenic proteins, it should not be considered as ameasure of allergenic proteins. Correlation of protein/antigenlevels with the level of allergenic proteins has not been fullyestablished.1.2 For the purpose of th
5、is test method, the range of proteinwill be measured in terms of microgram to milligram quanti-ties.1.3 This standard does not purport to address all of thesafety concerns, if any, associated with its use. It is theresponsibility of the user of this standard to establish appro-priate safety and heal
6、th practices and determine the applica-bility of regulatory limitations prior to use.2. Referenced Documents2.1 ASTM Standards:2D4483 Practice for Evaluating Precision for Test MethodStandards in the Rubber and Carbon Black ManufacturingIndustriesD5712 Test Method for Analysis of Aqueous Extractable
7、Protein in Latex, Natural Rubber, and Elastomeric Prod-ucts Using the Modified Lowry MethodE177 Practice for Use of the Terms Precision and Bias inASTM Test MethodsE691 Practice for Conducting an Interlaboratory Study toDetermine the Precision of a Test Method3. Terminology3.1 Definitions:3.1.1 alle
8、rgens, nprotein antigens which induce allergicimmune reactions typically mediated through IgE antibodies.3.1.2 antibody, nan immunoglobulin, a protein that isproduced as a part of the immune response which is capable ofspecifically combining with the antigen.3.1.3 antigen, nany substance that provok
9、es an immuneresponse when introduced into the body.3.1.4 background absorbance, nthe absorbance reading inthe solution resulting from the presence of chemicals, ions etc.other than the substrate being determined.3.1.5 blocking solution, na non-reactive protein solutionused to prevent nonspecific ant
10、ibody adsorption.3.1.6 calibration, nthe standardization of an instrumentsetting or an assay configuration.3.1.7 concentration range, nthe recommended analyteconcentration range in g/mL that produces an absorbancereading of 0.1 to 2.0 units.3.1.8 enzyme linked immunosorbent assay (ELISA), nanimmunol
11、ogical test method to quantify antigen or antibodylevels using an enzyme as the detection mechanism.3.1.9 primary antibody, nthe antibody used first in asequence that is specific for the antigen.3.1.10 reference solution, nthe solution to which the testsample is being compared against.3.1.11 repeata
12、bility, nthe variability or test error betweenindependent test results obtained within a single laboratory.3.1.12 reproducibility, nthe variability or error betweentest results obtained in different laboratories.3.1.13 secondary antibody, nthe enzyme conjugated anti-body used second in the sequence
13、that is specific for the heavychain of the primary antibody.3.1.14 standard solution, nthe preparation of standardanalyte used as a reference to which the unknown sample beingmeasured is compared.1This test method is under the jurisdiction ofASTM Committee D11 on Rubber,and is the direct responsibil
14、ity of Subcommittee D11.40 on Consumer RubberProducts.Current edition approved July 1, 2016. Published August 2016. Originallyapproved in 2000. Last previous edition approved in 2012 as D6499 12. DOI:10.1520/D6499-16.2For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact AST
15、M Customer Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States13.1.15 substrate, nthe materia
16、l or substance upon whichan enzyme reacts.3.1.16 titer, nthe strength of the antibody solution (forexample, concentration and affinity of antibody).4. Summary of Test Method4.1 The latex device is extracted for2hinanaqueousbuffer. The extract is recovered and the antigen levels aredetermined using i
17、nhibition Enzyme Linked ImmunoSorbentAssay (ELISA) technology (1).3The ELISA assay is based onpolyclonal antiserum which can detect NRL proteins. ELISAtechnology takes advantage of the specificity and sensitivity ofthe antibody-antigen reaction. A variation of the ELISAmethod (an inhibition ELISA) h
18、as been developed for thedetection and quantification of latex protein antigens. In theinhibition ELISA, the latex antigen is immobilized by absorp-tion to the wells of a 96-well test plate. The sample extract ismixed with antibody specific for NRL protein in a dilutionplate. Following a brief incub
19、ation to allow for antibodyrecognition of the relevant NRL antigens, the mixture is addedto the immobilized antigen in the assay plate. Anti-NRLantibody which is not bound to the soluble NRL protein in thesample will bind to the immobilized antigen. The plate iswashed to remove the soluble antigen a
20、ntibody complexes anda secondary antibody (enzyme-labeled anti-immunoglobulin)is added which attaches to the immobilized antigen-boundspecific antibody. Next, the enzyme substrate is added and thereaction of the enzyme on the substrate results in a colorchange.Areduction in the amount of color in co
21、mparison to anuninhibited control is an indicator of the amount of antigenpresent in the sample. Comparison to a standard curve gener-ated using known amounts of NRL protein permits quantifica-tion. The assay is highly sensitive and can quantitate NRLproteins in the nanogram per millilitre range.5.
22、Significance and Use5.1 Type 1 latex allergy most commonly manifests aslocalized urticaria after contact of skin with natural rubber butcan also include symptoms of allergic rhinoconjunctivitis,asthma and rarely anaphylaxis. This immediate (Type I) allergyis caused by natural proteins inherent to th
23、e rubber tree, whichremain on the finished natural rubber products. The quantifi-cation of protein levels in NRL products using the standardcolorimetric protein assays may give spurious results due tochemical additives in the latex formulations that interfere withthe assay (2,3). Furthermore, the am
24、ount of protein found inNRL products are often below the detection limits of thestandard colorimetric protein assay (4,5).5.2 This test method describes an immunological methodfor quantitation of natural rubber latex proteins using rabbitanti-NRL serum. Rabbits immunized with NRL proteins reactto th
- 1.请仔细阅读文档,确保文档完整性,对于不预览、不比对内容而直接下载带来的问题本站不予受理。
- 2.下载的文档,不会出现我们的网址水印。
- 3、该文档所得收入(下载+内容+预览)归上传者、原创作者;如果您是本文档原作者,请点此认领!既往收益都归您。
下载文档到电脑,查找使用更方便
5000 积分 0人已下载
下载 | 加入VIP,交流精品资源 |
- 配套讲稿:
如PPT文件的首页显示word图标,表示该PPT已包含配套word讲稿。双击word图标可打开word文档。
- 特殊限制:
部分文档作品中含有的国旗、国徽等图片,仅作为作品整体效果示例展示,禁止商用。设计者仅对作品中独创性部分享有著作权。
- 关 键 词:
- ASTMD649920163769STANDARDTESTMETHODFORTHEIMMUNOLOGICALMEASUREMENTOFANTIGENICPROTEININNATURALRUBBERANDITSPRODUCTS
链接地址:http://www.mydoc123.com/p-522687.html