1、Designation:D649918Standard Test Method forImmunological Measurement of Antigenic Protein in HeveaNatural Rubber(HNR)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 re
2、vision,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 Hevea Nat
3、uralRubber and its products using rabbit antisera specific forHNRL proteins.This immunoassay procedure quantitativelymeasures the level of antigenic latex proteins in solution usingan inhibition format.The samples may include glove or otherproduct extracts which have been collected in order to measu
4、rethe HNR levels.Although this method detects antigenicproteins,it should not be considered as a measure of allergenicproteins.Correlation of protein/antigen levels with the level ofallergenic proteins has not been fully established.1.2 For the purpose of this test method,the range of proteinwill be
5、 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,health,and environmental practices and deter-mine the
6、applicability of regulatory limitations prior to use.1.4 This international standard was developed in accor-dance with internationally recognized principles on standard-ization established in the Decision on Principles for theDevelopment of International Standards,Guides and Recom-mendations issued
7、by the World Trade Organization TechnicalBarriers to Trade(TBT)Committee.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 ExtractableProtein in La
8、tex,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 allergens,nprotein a
9、ntigens 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 provokes an immuneresponse
10、 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 antibody adsorption.3.1.6
11、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),nanimmunological test method to quant
12、ify 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 repeatability,nthe variability or te
13、st error betweenindependent test results obtained within a single laboratory.3.1.12 reproducibility,nthe variability or error betweentest results obtained in different laboratories.1This test method is under the jurisdiction of ASTM Committee D11 on Rubberand Rubber-like Materials,and is the direct
14、responsibility of Subcommittee D11.40on Consumer Rubber Products.Current edition approved Aug.1,2018.Published August 2018.Originallyapproved in 2000.Last previous edition approved in 2016 as D6499 16.DOI:10.1520/D6499-18.2For referenced ASTM standards,visit the ASTM website,www.astm.org,orcontact A
15、STM 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 StatesThis international standard was deve
16、loped in accordance with internationally recognized principles on standardization established in the Decision on Principles for theDevelopment of International Standards,Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade(TBT)Committee.1 3.1.13 secondary antibody,nthe enzyme conjugated anti-body used second in the sequence that is specific for the heavychain of the primary antibody.3.1.14 standard solution,nthe preparation of standardanalyte used as a re