ASTM F2450-2018 Standard Guide for Assessing Microstructure of Polymeric Scaffolds for Use in Tissue-Engineered Medical Products.pdf

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1、Designation: F2450 18Standard Guide forAssessing Microstructure of Polymeric Scaffolds for Use inTissue-Engineered Medical Products1This standard is issued under the fixed designation F2450; 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 guide covers an overview of test methods that maybe used to obtain information relating to th

3、e dimensions ofpores, the pore size distribution, the degree of porosity,interconnectivity, and measures of permeability for porousmaterials used as polymeric scaffolds in the development andmanufacture of tissue-engineered medical products (TEMPs).This information is key to optimizing the structure

4、 for aparticular application, developing robust manufacturing routes,and providing reliable quality control data.1.2 The values stated in SI units are to be regarded asstandard. No other units of measurement are included in thisstandard.1.3 This standard does not purport to address all of thesafety

5、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 applicability of regulatory limitations prior to use.1.4 This international standard was developed in accor-dance w

6、ith internationally recognized principles on standard-ization established in the Decision on Principles for theDevelopment of International Standards, Guides and Recom-mendations issued by the World Trade Organization TechnicalBarriers to Trade (TBT) Committee.2. Referenced Documents2.1 ASTM Standar

7、ds:2D2873 Test Method for Interior Porosity of Poly(VinylChloride) (PVC) Resins by Mercury Intrusion Porosim-etry (Withdrawn 2003)3D4404 Test Method for Determination of Pore Volume andPore Volume Distribution of Soil and Rock by MercuryIntrusion PorosimetryE128 Test Method for Maximum Pore Diameter

8、 and Perme-ability of Rigid Porous Filters for Laboratory UseE1294 Test Method for Pore Size Characteristics of Mem-brane Filters Using Automated Liquid Porosimeter (With-drawn 2008)3E1441 Guide for Computed Tomography (CT) ImagingF316 Test Methods for Pore Size Characteristics of Mem-brane Filters

9、by Bubble Point and Mean Flow Pore TestF2150 Guide for Characterization and Testing of Biomate-rial Scaffolds Used in Tissue-Engineered Medical Prod-uctsF2603 Guide for Interpreting Images of Polymeric TissueScaffolds3. Terminology3.1 Definitions:3.1.1 bioactive agent, nany molecular component in, o

10、n,or within the interstices of a device that is intended to elicit adesired tissue or cell response.3.1.1.1 DiscussionGrowth factors and antibiotics are typi-cal examples of bioactive agents. Device structural compo-nents or degradation byproducts that evoke limited localizedbioactivity are not bioa

11、ctive agents.3.1.2 blind (end)-pore, na pore that is in contact with anexposed internal or external surface through a single orificesmaller than the pores depth.3.1.3 closed cell, na void isolated within a solid, lackingany connectivity with an external surface. Synonym: closedpore3.1.4 hydrogel, na

12、 water-based open network of polymerchains that are cross-linked either chemically or throughcrystalline junctions or by specific ionic interactions.3.1.5 macropore/macroporosity (life sciences) ,na struc-ture (including void spaces) sized to allow substantially unre-stricted passage of chemicals, b

13、iomolecules, viruses, bacteria,and mammalian cells. In implants with interconnecting pores,macroporosity provides dimensions that allow for ready tissue1This guide is under the jurisdiction of ASTM Committee F04 on Medical andSurgical Materials and Devices and is the direct responsibility of Subcomm

14、itteeF04.42 on Biomaterials and Biomolecules for TEMPs.Current edition approved Nov. 15, 2018. Published December 2018. Originallyapproved in 2004. Last previous edition approved in 2010 as F2450 10. DOI:10.1520/F2450-18.2For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact

15、 ASTM Customer Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.3The last approved version of this historical standard is referenced onwww.astm.org.Copyright ASTM International, 100 Barr Harbor Drive, PO

16、Box C700, West Conshohocken, PA 19428-2959. United StatesThis international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for theDevelopment of International Standards, Guides and Recommendations issued by

17、 the World Trade Organization Technical Barriers to Trade (TBT) Committee.1penetration and microvascularization after implantation. In-cludes materials that contain voids with the potential to beobservable to the naked eye (100 m).3.1.6 micropore/microporosity (life sciences) ,na struc-ture (includi

18、ng void spaces) sized to allow substantially unre-stricted passage of chemicals, biomolecules, and viruses whilesized to control or moderate the passage of bacteria, mamma-lian cells, and/or tissue. Includes materials with typical poresizes greater than 0.1 m (100 nm) and less than about 100 m(100 0

19、00 nm), with a common microporous context encom-passing the range of 20 m or less for the filtration of cellsranging from bacteria to common mammalian cells and above30 m for the ingrowth of tissue. Objects in this size rangetypically can be observed by conventional light microscopy.3.1.7 nanopore/n

20、anoporosity (life sciences),na structureinclusive of void spaces sized to control or moderate thepassage of chemicals, biomolecules, and viruses while sized tosubstantially exclude most bacteria and all mammalian cells.Includes materials with typical pore sizes of less than 100 nm(0.1 m), with commo

21、n nanoporous context in the range of 20nm or less for the filtration of viruses.3.1.8 permeability, na measure of fluid, particle, or gasflow through an open pore structure.3.1.9 polymer, na long chain molecule composed ofmonomers including both natural and synthetic materials.Examples include colla

22、gen and polycaprolactone.3.1.10 pore, na fluid (liquid or gas) filled externallyconnecting channel, void, or open space within an otherwisesolid or gelatinous material (for example, textile meshescomposed of many or single fibers (textile based scaffolds),open cell foams, hydrogels). Synonyms: open-

23、pore, through-pore.3.1.11 porogen, na material used to create pores within aninherently solid material.3.1.11.1 DiscussionFor example, a polymer dissolved inan organic solvent is poured over a water-soluble powder.Afterevaporation of the solvent, the porogen is leached out, usuallyby water, to leave

24、 a porous structure. The percentage ofporogen needs to be high enough to ensure that all the pores areinterconnected.3.1.12 porometry, nthe determination of the distributionof open pore diameters relative to the direction of fluid flow bythe displacement of a non-volatile wetting fluid as a function

25、 ofpressure.3.1.13 porosimetry, nthe determination of the pore vol-ume and pore size distribution through the use of a non-wettingliquid (typically mercury) intrusion into a porous material as afunction of pressure.3.1.14 porosity, nproperty of a solid which contains aninherent or induced network of

26、 channels and open spaces.Porosity can be determined by measuring the ratio of pore(void) volume to the apparent (total) volume of a porousmaterial and is commonly expressed as a percentage.3.1.15 scaffold, na support, delivery vehicle, or matrix forfacilitating the migration, binding, or transport

27、of cells orbioactive molecules used to replace, repair, or regeneratetissues.3.1.16 through-pores, nan inherent or induced network ofvoids or channels that permit flow of fluid (liquid or gas) fromone side of the structure to the other.3.1.17 tortuosity, na measure of the mean free path lengthof thr

28、ough-pores relative to the sample thickness. Alternativedefinition: The squared ratio of the mean free path to theminimum possible path length.4. Summary of Guide4.1 The microstructure, surface chemistry, and surface mor-phology of polymer-based tissue scaffolds plays a key role inencouraging cell a

29、dhesion, migration, growth, and prolifera-tion. The intention of this guide is to provide a compendium oftechniques for characterizing this microstructure. The breadthof the techniques described reflects the practical difficulties ofquantifying pore sizes and pore size distributions over lengthscale

30、s ranging from nanometres to sub-millimetres and theporosity of materials that differ widely in terms of theirmechanical properties.4.2 These microstructural data when used in conjunctionwith other characterization methods, for example, chemicalanalysis of the polymer (to determine parameters such a

31、s themolecular mass (molecular weight) and its distribution), willaid in the optimization of scaffolds for tissue-engineeredmedical products (TEMPs). Adequate characterization is alsocritical to ensure the batch-to-batch consistency of scaffolds;either to assess base materials supplied by different

32、suppliers orto develop robust manufacturing procedures for commercialproduction.4.3 Application of the techniques described in this guidewill not guarantee that the scaffold will perform the functionsfor which it is being developed but they may help to identifythe reasons for success or failure.4.4

33、This guide does not suggest that all listed tests beconducted. The choice of technique will depend on theinformation that is required and on the scaffolds physicalproperties; for example, mercury porosimetry will not yieldmeaningful data if used to characterize soft materials thatdeform during the t

34、est and cannot be used for hydratedscaffolds.4.5 Table 1 provides guidance for users of this guide byproviding a brief overview of the applicability of a range ofdifferent measurement techniques that can be used to physi-cally characterize tissue scaffolds. This list of techniques is notdefinitive.5

35、. Significance and Use5.1 The ability to culture functional tissue to repair damagedor diseased tissues within the body offers a viable alternative toxenografts or heterografts. Using the patients own cells toproduce the new tissue offers significant benefits by limitingrejection by the immune syste

36、m. Typically, cells harvestedfrom the intended recipient are cultured in vitro using aF2450 182temporary housing or scaffold. The microstructure of thescaffold can be defined by the existence, type, size distribution,interconnectivity, and directionality of pores all of which arecritical for cell mi

37、gration, growth, and proliferation (AppendixX1). Optimizing the design of tissue scaffolds is a complextask, given the range of available materials, different manufac-turing routes, and processing conditions. All of these factorscan, and will, affect the surface texture, surface chemistry, andmicros

38、tructure of the resultant scaffolds. Surface texture, sur-face chemistry, and microstructure of the scaffolds may or maynot be significant variables depending on the characteristics ofa given cell type at any given time (that is, changes in cellbehavior due to the number of passages, mechanicalstimu

39、lation, and culture conditions).5.2 Tissue scaffolds are typically assessed using an overallvalue for scaffold porosity and a range of pore sizes, though thedistribution of sizes is rarely quantified. Published mean poresizes and distributions are usually obtained from electronmicroscopy images and

40、quoted in the micrometer range. Tissuescaffolds are generally complex structures that are not easilyinterpreted in terms of pore shape and size, especially in threedimensions. Therefore, it is difficult to quantifiably assess thebatch-to-batch variance in microstructure or to make a system-atic inve

41、stigation of the role that the mean pore size and poresize distribution has on influencing cell behavior based solelyon electron micrographs (Tomlins et al, (1).45.2.1 Fig. 1 gives an indication of potential techniques thatcan be used to characterize the structure of porous tissuescaffolds and the l

42、ength scale that they can measure. Clearly arange of techniques must be used if the scaffold is to becharacterized in detail.5.2.2 The classification and terminology of pore sizes, suchas those given in Table 2, has yet to be standardized, withdefinitions of terms varying widely (as much as three or

43、ders ofmagnitude) between differing applications and industries. BothTable 2 and the supporting detailed discussion included withinAppendix X2 describe differences that exist between IUPAC(International Union of Pure and Applied Chemistry) defini-tions and the common terminology currently used withi

44、n mostlife science applications, which include both implant and tissueengineering applications.5.2.2.1 Since the literature contains many other terms fordefining pores (Perret et al (3), it is recommended that theterms used by authors to describe pores be defined in order toavoid potential confusion

45、. Additionally, since any of thedefinitions in Table 2 can shift, depending on the pore sizedetermination method (see Table 1 and Fig. 1), an accompa-nying statement describing the assessment technique used isessential.5.2.3 All the techniques listed in Table 1 have limitations forassessing complex

46、porous structures. Fig. 2a and Fig. 2b showa through- and a blind-end pore respectively. Porometrymeasurements (see 7.4) are only sensitive to the narrowestpoint along a variable diameter through-pore and therefore cangive a lower measure of the pore diameter than other investi-gative techniques, su

47、ch as scanning electron microscope(SEM), which may sample at a different point along the pore.The physical basis of porometry depends on the passage of gasthrough the material. Therefore, the technique is not sensitiveto blind-end or closed pores. Estimates of porosity based onporometry data will th

48、erefore be different from those obtainedfrom, for example, porosimetry (see 7.3), which is sensitive toboth through- and blind-pores or density determinations thatcan also account for through-, blind-end, and closed pores. Thesignificance of these differences will depend on factors such asthe percen

49、tage of the different pore types and their dimensions.Further research will enable improved guidance to be devel-oped.5.2.4 Polymer scaffolds range from mechanically rigidstructures to soft hydrogels. The methods currently used tomanufacture these structures include, but are not limited to:5.2.4.1 Casting a polymer, dissolved in an organic solvent,over a water-soluble particulate porogen, followed by leaching.5.2.4.2 Melt mixing of immiscible polymers followed byleaching of the water-soluble component.5.2.4.3 Dissolution of supercritical carbon dioxi