ASTM D7427-2014 7963 Standard Test Method for Immunological Measurement of Four Principal Allergenic Proteins &40 Hev b 1 3 5 and 6 02&41 in Natural Rubber and Its Products Derived.pdf

1、Designation: D7427 14Standard Test Method forImmunological Measurement of Four Principal AllergenicProteins (Hev b 1, 3, 5 and 6.02) in Natural Rubber and ItsProducts Derived from Latex1This standard is issued under the fixed designation D7427; the number immediately following the designation indica

2、tes the year oforiginal adoption or, in the case of revision, 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 met

3、hodknown as an immunoenzymetric assay to quantify the amountof 4 principal Hevea brasiliensis Hev b allergenic proteinsHev b 1, Hev b 3, Hev b 5 and Hev b 6.02 in natural rubberand its products2derived from latex using monoclonal anti-bodies specific for epitopes on these proteins. Since theseassays

4、 quantify the levels of only 4 of the known 14 officiallyacknowledged allergens potentially present in natural rubberlatex containing products, the sum of the four allergen levelsshall be viewed as an indicator of the allergen burden and notas a measure of the total allergen content that can be rele

5、asedfrom the product.1.2 For the purpose of this test method, the range ofallergenic protein will be measured in terms of nanogram tomicrogram quantities per gram or unit surface area of a naturalrubber containing product.1.3 The test method is not designed to evaluate the potentialof natural rubber

6、 containing materials to induce or elicit TypeI (IgE-mediated) hypersensitivity reactions.1.4 This test method should be used under controlledlaboratory conditions to detect and quantify the level of 4allergenic proteins found in natural rubber containing products.It should not be used to describe,

7、appraise or assess the hazardor risk of these natural rubber containing materials or productsunder actual in use conditions.1.5 The values stated in SI units are to be regarded asstandard. No other units of measurement are included in thisstandard.1.6 This standard does not purport to address all of

8、 thesafety concerns, if any, associated with its use. It is theresponsibility of the user of this standard to establish appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to use.2. Referenced Documents2.1 ASTM Standards:3D1193 Specification for

9、Reagent WaterD4483 Practice for Evaluating Precision for Test MethodStandards in the Rubber and Carbon Black ManufacturingIndustriesD4678 Practice for RubberPreparation, Testing,Acceptance, Documentation, and Use of Reference Mate-rialsD5712 Test Method for Analysis of Aqueous ExtractableProtein in

10、Natural Rubber and Its Products Using theModified Lowry MethodD6499 Test Method for The Immunological Measurement ofAntigenic Protein in Natural Rubber and its ProductsE691 Practice for Conducting an Interlaboratory Study toDetermine the Precision of a Test Method3. Terminology3.1 Definitions:3.1.1

11、accepted reference value (ARV)value that serves asan agreed upon reference for comparison and which is derivedas (1) a theoretical or established value, based on scientificprinciples, (2) an assigned or certified value, based on experi-mental work of some national or international organization, or(3

12、) a consensus or certified value, based on collaborativeexperimental work under the auspices of a scientific orengineering group.3.1.1.1 DiscussionARV is an average industrial referencematerial (IRM) property or parameter value established by wayof a specified test program. In this standard, theARVa

13、s definedin the IRMs for the reference antigens and capture anddetection antibodies is determined by analyzing a high and low1This test method is under the jurisdiction of ASTM Committee D11 on Rubberand is the direct responsibility of Subcommittee D11.40 on Consumer RubberProducts.Current edition a

14、pproved Sept. 1, 2014. Published September 2014. Originallyapproved in 2008. Last previous edition approved in 2008 as D7427 082. DOI:10.1520/D7427-14.2This procedure has not been validated for condoms, particularly lubricatedcondoms, which could contain surfactants or other ingredients that could i

15、nterferewith the assay.3For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM 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

16、 Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States1control in an inter-laboratory study and using the assignedvalues of these high and low controls to verify that the assay isin control and that the reagents are performing properly.3.1.2 accuracythe closeness of agreement be

17、tween a testresult and an accepted reference value.3.1.3 allergensprotein antigens which induce allergic im-mune reactions typically mediated through IgE antibodies.3.1.4 analyteany element, ion, compound, substance,factor, infectious agent, cell, organelle, activity (enzymatic,hormonal, or immunolo

18、gical), or property the presence orabsence, concentration, activity, intensity, or other characteris-tics of which are to be determined.3.1.5 antibodyan immunoglobulin, a protein that is pro-duced as a part of the humoral immune response which iscapable of specifically combining with antigen.3.1.5.1

19、 DiscussionAny of numerous Y-shaped proteinmolecules produced by B lymphocytes as a primary immuneresponse, each molecule and its clones having a unique bindingsite that can combine with the complementary site of anantigen, as on a virus or bacterium, thereby signaling otherimmune responses. (See mo

20、noclonal antibody.)3.1.6 antigenany substance that can stimulate the produc-tion of antibodies within an organism and combine specificallywith them.3.1.7 background absorbancethe absorbance reading inthe solution resulting from non-specific interactions caused bythe presence of chemicals, ions, etc.

21、, other than the analytebeing measured.3.1.8 binding capacitywithin the context of this document,refers to the number of Hev b allergen molecules that a primarycapture antibody can bind reproducibly under standardizedassay conditions (pH, ionic strength, protein matrix, time,temperature).3.1.9 block

22、ing solutiona non-reactive protein solutionused to prevent nonspecific antibody adsorption and to reducebackground absorbance.3.1.10 calibrationthe standardization of an instrumentsetting or an assay configuration.3.1.11 calibration material/calibratora material (forexample, solution) of known quant

23、itative/qualitative character-istics (for example, concentration, activity, intensity, reactivity)used to calibrate, graduate, or adjust a measurement procedureor to compare the response obtained with the response of a testspecimen/sample.3.1.12 concentration rangethe recommended analyte con-centrat

24、ion range in nanograms per mL to micrograms per mLthat produces an absorbance reading from 0.1 to 2.03.0 units(depending on the instrument).3.1.13 data reduction algorithma mathematical processthat converts assay-response data (for example, absorbanceunits) into interpolated dose results.3.1.13.1 Di

25、scussionThe doseresponse relationship in theassay is defined by the standard, reference, or calibration curve.3.1.14 detection limit/limit of detectionthe smallest quan-tity of an analyte that can be reproducibly and a statisticallysignificant manner distinguished from the variance of thebackground,

26、 or a zero calibrator in a given assay system.3.1.14.1 DiscussionIt is usually defined at the 95 % con-fidence interval and has also been called the lower detectionlimit or positive threshold of the assay; this term is notsynonymous with analytical sensitivity.3.1.15 enzyme linked immunosorbent assa

27、y (ELISA)animmunological test method to quantify antigen or antibodylevels using an enzyme as the detection mechanism.3.1.16 epitope/determinant(1) the minimum molecularstructure of the antigenic site that will react with an antibody;(2) any site on an antigen molecule at which an antibody canbind;

28、the chemical structure of the site determining the specificcombining antibody.3.1.17 IgEhuman IgE is an immunoglobulin of the ap-proximate molecular weight of 190 000, which exists normallyin monomeric form and constitutes approximately 0.0005 % ofthe total serum immunoglobulins.3.1.17.1 DiscussionI

29、t (IgE) binds with high affinity toFcR1 receptors on mast cells and basophils and FcRIIreceptors on a number of cells. IgE mediates the release ofvasoactive mediators following the binding of allergen.3.1.18 immunoenzymetric assay (IEMA)a two-site non-isotopic immunological test method that employs

30、twoantibodies, a primary antibody to capture and a secondaryenzyme conjugated antibody to detect the analyte of interest.3.1.19 immunoglobulina glycoprotein composed of twoheavy and two light chains that functions as an antibody.Human immunoglobulins have been subdivided into differentisotypes (IgM,

31、 IgG, IgA, IgD, IgE), each of which possess aunique set of antigenic markers, physiochemical properties,and each of which produce a different pattern of effectorfunctions (receptor binding, complement activation, opsoniza-tion).3.1.19.1 DiscussionAll antibodies are immunoglobulins,but it is not cert

32、ain that all immunoglobulins possess antibodyfunction.3.1.20 industry reference materials (IRM)materials thathave been prepared according to a specified production processto generate a uniform lot; the parameters that define the qualityof the lot are evaluated by a specified measurement program.3.1.

33、20.1 DiscussionIRMs are divided into two types ac-cording to the production process for generating the material.3.1.21 linearitythe ability (within a given range) of anassay to provide results that are directly proportional to theconcentration amount of the analyte in the test sample.3.1.22 monoclon

34、al antibodyantibody produced by cellscreated through the fusion of an antibody producing cell(B-lymphocyte) with immortal cancer cells.3.1.22.1 DiscussionThis fusion process produces a hybrid(hybridoma) that expresses properties of both cells. The cellsare all identical since they derive from a sing

35、le cell and arecalled “monoclonal.”3.1.23 parallelismextent to which the doseresponse re-lationship between two materials (that is, calibrator versusunknown specimens) is constant for the examined range ofconcentrations.D7427 1423.1.23.1 DiscussionParallelism is a property (and a re-quirement) of qu

36、antitative immunoassays in which the calibra-tor and test sera produce parallel doseresponse curves.3.1.24 precisionthe closeness of agreement between inde-pendent test results obtained under prescribed conditions;agreement between replicate measurements.3.1.24.1 DiscussionPrecision has no numerical

37、 value butis expressed in terms of imprecisionthe standard deviation(SD) or the coefficient of variation (CV: SD/mean) of theresults in a set of replicate measurements.3.1.25 precision profilethe precision of an assay across theanalyte concentration range of interest.3.1.25.1 DiscussionA precision p

38、rofile is constructed bydetermining the standard deviation (or coefficient of variation)of replicate measurements (within assays, between assays, orbetween specimen dilutions within an assay) spanning theentire analyte concentration range, albeit without the exactknowledge of the true analyte concen

39、tration that is contained inthe serum specimens. When the CVdose(Y-axis) is graphedagainst the dose (X-axis), a precision profile plot is generated.The precision profile is also referred to as the “imprecisionprofile” by some investigators.3.1.26 primary antibodythe antibody used first in an assayse

40、quence that is specific for the antigen and is sometimesreferred to as the capture antibody that binds the analyte ofinterest from a biological specimen.3.1.27 proficiency testing (PT)an independent (non-manufacturer sponsored) program in which challenge speci-mens are sent to participating laborato

41、ries to be evaluated inassays that measure a spectrum of analytes.3.1.28 qualitative assayan assay system that produces anindication of the presence or absence of an analyte but does notprovide a precise estimate of the concentration of that analyte.3.1.28.1 DiscussionApositive test result implies o

42、nly thatthe assay signal exceeds the analytical threshold or positivecutoff point that has been set to obtain an arbitrary combinationof diagnostic sensitivity and specificity.3.1.29 quantitative assayan assay system that produces anaccurate and reproducible estimate of the concentration of ananalyt

43、e in the test specimen.3.1.29.1 DiscussionIts (quantitative assay) analysis in-volves interpolation from a calibration curve, which is refer-enced to a readily available standard reference preparation.3.1.30 quality control responselevel of analyte producedby an assay for a quality control specimen

44、that has a previouslydefined analyte concentration range as defined by the manu-facturer.3.1.30.1 DiscussionAssay performance was evaluated bydetermining the agreement in Hev b 1, 3, 5 or 6.02 levelsobtained for two quality control extracts containing a high orlow level of each Hev b allergen, follo

45、wing analysis in multiplelaboratories participating in the multi-center study.3.1.31 reference solutionthe solution against which thetest sample is being compared.3.1.32 relative standard deviation (RSD)the coefficient ofvariation which is the standard deviation divided by the mean.3.1.33 repeatabil

46、ityprecision under conditions where in-dependent test results are obtained with the same method onidentical test items in the same laboratory by the same operatorusing the same equipment within short intervals of time.3.1.34 repeatability limit (r)the value below which theabsolute difference between

47、 two individual test results obtainedunder repeatability conditions may be expected to occur with aprobability of approximately 0.95 (95 %).3.1.34.1 DiscussionThe repeatability limit is 2.8(1.96 square root of 2) times the repeatability standarddeviation. This multiplier is independent of the size o

48、f theinter-laboratory study.3.1.35 reproducibilityprecision obtained under conditionswhere test results are obtained with the same method onidentical test items in different laboratories with differentoperators using different equipment.3.1.36 reproducibility limit (R)the value below which theabsolu

49、te difference between two test results obtained underreproducibility conditions may be expected to occur with aprobability of approximately 0.95 (95 %).3.1.36.1 DiscussionThe reproducibility limit is 2.8(1.96 square root of 2) times the reproducibility standarddeviation. This multiplier is independent of the number oflaboratories participating.3.1.37 secondary antibodyin an IEMA, it is the enzymeconjugated antibody used second in a sequence that is specificfor the analyte or interest and that completes the sa