ASTM F2148-18
(Practice)Standard Practice for Evaluation of Delayed Contact Hypersensitivity Using the Murine Local Lymph Node Assay (LLNA)
Standard Practice for Evaluation of Delayed Contact Hypersensitivity Using the Murine Local Lymph Node Assay (LLNA)
SIGNIFICANCE AND USE
5.1 The propensity of a material to stimulate delayed contact hypersensitivity must be assessed before clinical application of devices containing this material. Delayed hypersensitivity may occur anywhere in the body. Systemic delayed hypersensitivity may have a complex set of reactions and consequences depending on the actual tissue/organ site of reaction. Although the reactions are seldom life-threatening, severe tissue and organ damage my result over time. Skin is the usual test site to determine the propensity of a material to cause delayed hypersensitivity.
5.2 The standard historical test methods have involved the use of guinea pigs with a cutaneous application and observation of the reaction site. The use of the murine local lymph node assay results in a numerical quantitation of stimulation, rather than subjective evaluation and could be used to determine dose responses.
5.3 This practice may not be predictive of events occurring during all types of implant applications. The user is cautioned to consider the appropriateness of the method in view of the materials being tested, their potential applications, and the recommendations contained in Practice F748.
SCOPE
1.1 This practice provides a methodology to use a combination of in vivio and in situ procedures for the evaluation of delayed contact hypersensitivity reactions.
1.2 This practice is intended to provide an alternative to the use of guinea pigs for evaluation of the ability of a device material to stimulate delayed contact hypersensitivity reactions. This alternative is particularly applicable for materials used in devices that contact only intact skin. However, the guinea pig maximization test is still the recommended method when assessing the delayed hypersensitivity response to metals or when testing substances that do not penetrate the skin but are used in devices that contact deep tissues or breached surfaces. This practice may be used for testing metals, with the exception of nickel-containing metals, unless the unique physicochemical properties of the materials may interfere with the ability of LLNA to detect sensitizing substances.
1.3 This practice consists of a protocol for assessing an increase in lymphocyte proliferation in the lymph nodes draining the site of test article administration on the ears of mice.
1.4 The LLNA has been validated only for low-molecular-weight chemicals that can penetrate the skin. The absorbed chemical or metabolite must bind to macromolecules, such as proteins, to form immunogenic conjugates.
1.5 This practice is one of several developed for the assessment of the biocompatibility of materials. Practice F748 may provide guidance for the selection of appropriate methods for testing materials for a specific application.
1.6 Identification of a supplier of materials or reagents is for the convenience of the user and does not imply a single source. Appropriate materials and reagents may be obtained from many commercial supply houses.
1.7 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.
1.8 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.
1.9 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
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This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the
Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
Designation: F2148 − 18
Standard Practice for
Evaluation of Delayed Contact Hypersensitivity Using the
1
Murine Local Lymph Node Assay (LLNA)
This standard is issued under the fixed designation F2148; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
1. Scope 1.7 The values stated in SI units are to be regarded as
standard. No other units of measurement are included in this
1.1 This practice provides a methodology to use a combi-
standard.
nation of in vivio and in situ procedures for the evaluation of
1.8 This standard does not purport to address all of the
delayed contact hypersensitivity reactions.
safety concerns, if any, associated with its use. It is the
1.2 This practice is intended to provide an alternative to the
responsibility of the user of this standard to establish appro-
use of guinea pigs for evaluation of the ability of a device
priate safety, health, and environmental practices and deter-
material to stimulate delayed contact hypersensitivity reac-
mine the applicability of regulatory limitations prior to use.
tions. This alternative is particularly applicable for materials
1.9 This international standard was developed in accor-
used in devices that contact only intact skin. However, the
dance with internationally recognized principles on standard-
guinea pig maximization test is still the recommended method
ization established in the Decision on Principles for the
when assessing the delayed hypersensitivity response to metals
Development of International Standards, Guides and Recom-
orwhentestingsubstancesthatdonotpenetratetheskinbutare
mendations issued by the World Trade Organization Technical
used in devices that contact deep tissues or breached surfaces.
Barriers to Trade (TBT) Committee.
This practice may be used for testing metals, with the excep-
tion of nickel-containing metals, unless the unique physico-
2. Referenced Documents
chemical properties of the materials may interfere with the
2
2.1 ASTM Standards:
ability of LLNA to detect sensitizing substances.
F619 Practice for Extraction of Medical Plastics
1.3 This practice consists of a protocol for assessing an
F720 PracticeforTestingGuineaPigsforContactAllergens:
increase in lymphocyte proliferation in the lymph nodes
Guinea Pig Maximization Test
draining the site of test article administration on the ears of
F748 PracticeforSelectingGenericBiologicalTestMethods
mice.
for Materials and Devices
F750 Practice for Evaluating Material Extracts by Systemic
1.4 The LLNA has been validated only for low-molecular-
Injection in the Mouse
weight chemicals that can penetrate the skin. The absorbed
3
chemical or metabolite must bind to macromolecules, such as
2.2 Other Documents:
proteins, to form immunogenic conjugates.
ICCVAM NIH Publication No: 99-4494 The Murine Local
Lymph Node Assay, 1999
1.5 This practice is one of several developed for the
ICCVAM NIH Publication No: 10-7512 Test Method Evalu-
assessment of the biocompatibility of materials. Practice F748
ation Report on Using the Murine Local Lymph Node
may provide guidance for the selection of appropriate methods
Assay for Testing Pesticide Formulations, Metals, Sub-
for testing materials for a specific application.
stances in Aqueous Solutions, and Other Products, 2010
1.6 Identification of a supplier of materials or reagents is for
ICCVAM NIH Publication NO: 11-7709 Usefulness and
the convenience of the user and does not imply a single source.
Limitations of the Murine Local Lymph Node Assay for
Appropriate materials and reagents may be obtained from
Potency Categorization of Chemicals Causing Allergic
many commercial supply houses.
Contact Dermatitis in Humans
1 2
ThispracticeisunderthejurisdictionofASTMCommitteeF04onMedicaland For referenced ASTM standards, visit the ASTM website, www.astm.org, or
Surgical Materials and Devices and is the direct responsibility of Subcommittee contact ASTM Customer Service at service@astm.org. For Annual Book of ASTM
F04.16 on Biocompatibility Test Methods. Standards volume information, refer to the standard’s Document Summary page on
Current edition approved Dec. 1, 2018. Published February 2019. Originally the ASTM website.
3
approved in 2001. Last previous edition approved in 2013 as F2148 – 13. DOI: Available from NICEATM, NIEHS, 79 Alexander Dr., Mail Drop EC-17,
10.1520/F2148-18. Research Triangle Park, NC 27709.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700,
...
This document is not an ASTM standard and is intended only to provide the user of an ASTM standard an indication of what changes have been made to the previous version. Because
it may not be technically possible to adequately depict all changes accurately, ASTM recommends that users consult prior editions as appropriate. In all cases only the current version
of the standard as published by ASTM is to be considered the official document.
Designation: F2148 − 13 F2148 − 18
Standard Practice for
Evaluation of Delayed Contact Hypersensitivity Using the
1
Murine Local Lymph Node Assay (LLNA)
This standard is issued under the fixed designation F2148; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
1. Scope
1.1 This practice provides a methodology to use an in-situ procedure a combination of in vivio and in situ procedures for the
evaluation of delayed contact hypersensitivity reactions.
1.2 This practice is intended to provide an alternative to the use of guinea pigs for evaluation of the ability of a device material
to stimulate delayed contact hypersensitivity reactions. This alternative is particularly applicable for materials used in devices that
contact only intact skin. However, the guinea pig maximization test is still the recommended method when assessing the delayed
hypersensitivity response to metals or when testing substances that do not penetrate the skin but are used in devices that contact
deep tissues or breached surfaces. This practice may be used for testing metals, with the exception of nickel-containing metals,
unless the unique physicochemical properties of the materials may interfere with the ability of LLNA to detect sensitizing
substances.
1.3 This practice consists of a protocol for assessing an increase in lymphocyte proliferation withinin the lymph nodes draining
the site of test article administration on the ears of mice.
1.4 The LLNA has been validated only for low-molecular-weight chemicals that can penetrate the skin. The absorbed chemical
or metabolite must bind to macromolecules, such as proteins, to form immunogenic conjugates.
1.5 This practice is one of several developed for the assessment of the biocompatibility of materials. Practice F748 may provide
guidance for the selection of appropriate methods for testing materials for a specific application.
1.6 Identification of a supplier of materials or reagents is for the convenience of the user and does not imply a single source.
Appropriate materials and reagents may be obtained from many commercial supply houses.
1.7 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.
1.8 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility
of the user of this standard to establish appropriate safety safety, health, and healthenvironmental practices and determine the
applicability of regulatory limitations prior to use.
1.9 This international standard was developed in accordance with internationally recognized principles on standardization
established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued
by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
2. Referenced Documents
2
2.1 ASTM Standards:
F619 Practice for Extraction of Medical Plastics
F720 Practice for Testing Guinea Pigs for Contact Allergens: Guinea Pig Maximization Test
F748 Practice for Selecting Generic Biological Test Methods for Materials and Devices
F750 Practice for Evaluating Material Extracts by Systemic Injection in the Mouse
3
2.2 Other Documents:
ICCVAM NIH Publication No: 99-4494 The Murine Local Lymph Node Assay, 1999
1
This practice is under the jurisdiction of ASTM Committee F04 on Medical and Surgical Materials and Devices and is the direct responsibility of Subcommittee F04.16
on Biocompatibility Test Methods.
Current edition approved June 1, 2013Dec. 1, 2018. Published August 2013February 2019. Originally approved in 2001. Last previous edition approved in 20122013 as
F2148 – 07 (2012).F2148 – 13. DOI: 10.1520/F2148-13.10.1520/F2148-18.
2
For referenced ASTM standards, visit the ASTM website, www.astm.org, or contact ASTM Customer Service at service@astm.org. For Annual Book of ASTM Standards
volume information, refer to the standard’s Document Summary page on the ASTM website.
3
Available from NICEATM, NIEHS, 79 Alexander Dr., Mail Drop EC-17, Research Triangle Park, NC 27709.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
1
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F2148 − 18
ICCVAM NIH Publication No: 1
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