1、Designation: D7391 09D7391 17Standard Test Method forCategorization and Quantification of Airborne FungalStructures in an Inertial Impaction Sample by OpticalMicroscopy1This standard is issued under the fixed designation D7391; the number immediately following the designation indicates the year ofor
2、iginal 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 is a procedure that uses direct microscopy t
3、o analyze the deposit on an inertial impaction sample.1.2 This test method describes procedures for categorizing and enumerating fungal structures by morphological type. Typically,categories may be as small as genus (for example, Cladosporium) or as large as phylum (for example, basidiospores).1.3 T
4、his test method contains two procedures for enumerating fungal structures: one for slit impaction samples and one forcircular impaction samples. This test method is applicable for impaction air samples, for which a known volume of air (at a rateas recommended by the manufacturer) has been drawn, and
5、 is also applicable for blank impaction samples.1.4 Enumeration results are presented in fungal structures/sample (fs/sample) and fungal structures/m3 (fs/m3).1.5 The range of enumeration results that can be determined with this test method depends on the size of the spores on thesample trace, the a
6、mount of particulate matter on the sample trace, the percentage of the sample trace counted, and the volumeof air sampled.1.6 This test method addresses only the analysis of samples. The sampling process and interpretation of results is outside thescope of this test method.1.7 The values stated in S
7、I 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 responsibilityof the user of this standard to establish appropriate safety and heal
8、th practices and determine the applicability of regulatorylimitations prior to use.1.9 This international standard was developed in accordance with internationally recognized principles on standardizationestablished in the Decision on Principles for the Development of International Standards, Guides
9、 and Recommendations issuedby the World Trade Organization Technical Barriers to Trade (TBT) Committee.2. Referenced Documents2.1 ASTM Standards:2D1193 Specification for Reagent WaterE691 Practice for Conducting an Interlaboratory Study to Determine the Precision of a Test Method3. Terminology3.1 AS
10、TM Definitions (see the ASTM Terminology Dictionary)Online Dictionary of Engineering Science and Technology3):3.1.1 numerical aperturenumerical aperture.3.2 Definitions of Terms Specific to This Standard:1 This test method is under the jurisdiction of ASTM Committee D22 on Air Quality and is the dir
11、ect responsibility of Subcommittee D22.08 on Sampling and Analysisof Mold.Current edition approved May 1, 2009March 15, 2017. Published June 2009April 2017. Originally approved in 2009. Last previous edition approved in 2009 as D7391 09. DOI: 10.1520/D7391-09.10.1520/D7391-17.2 For referencedASTM st
12、andards, visit theASTM website, www.astm.org, or contactASTM Customer Service at serviceastm.org. For Annual Book of ASTM Standardsvolume information, refer to the standards Document Summary page on the ASTM website.3 ASTM Online Dictionary of Engineering Science and Technology (Stock #: DEFONLINE)
13、is available on theASTM website, www.astm.org, or contactASTM CustomerService at serviceastm.org.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. Becauseit may not be technically p
14、ossible to adequately depict all changes accurately, ASTM recommends that users consult prior editions as appropriate. In all cases only the current versionof the standard as published by ASTM is to be considered the official document.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700,
15、 West Conshohocken, PA 19428-2959. United States13.2.1 circular impaction sample, na sample of airborne particulate matter collected via by means of a device that draws airthrough a round aperture at a specified rate, impacting the particles suspended in the air onto an adhesive medium, resulting in
16、 acircular area of deposition. A circular impaction sample may be collected by means of a cassette manufactured for that purpose,or by means of a sampling device that requires slides to be pre-coated with impaction medium.3.2.2 debris rating, na distinct value assigned to an impactor sample based on
17、 the percentage of the sample area potentiallyobscured by particulate matter, and ranging from 0 to 5.3.2.3 field blank, na sample slide or cassette carried to the sampling site, exposed to sampling conditions (for example, sealsopened), returned to the laboratory, treated as a sample, and carried t
18、hrough all steps of the analysis.3.2.4 fungal structure (sing.), na collective term for fragments or groups of fragments from fungi, including but not limitedto conidia, conidiophores, and hyphae and spores.3.2.5 fungus (s), fungi, (pl.), neukaryotic, heterotrophic, absorptive organisms that usually
19、 develop a rather diffuse, branched,tubular body (for example, network of hyphae) and usually reproduce by means of spores. The terms mold and mildew arefrequently used by laypersons when referring to various fungal colonization.3.2.6 hyaline, adjcolorless.3.2.7 impaction medium, na substance applie
20、d to a microscope slide used to collect (or capture) particulate matter duringsampling.3.2.8 impaction sample, na sample taken using impaction, for example, slit impaction sample, circular impaction sample.3.2.9 inertial impactor, na device for collecting particles separated from an air stream by in
21、ertia to force an impact onto anadhesive surface. Inertial impactors are available in many designs, including those having a slit jet, yielding a rectangular sampletrace, and a circular jet, yielding a circular sample trace.3.2.10 magnification/resolution combination 1, n150-400 150400 total magnifi
22、cation and a point to point resolution of0.7 m or better, as checked by a resolution check slide.3.2.11 magnification/resolution combination 2, n 400 or greater total magnification and a point to point resolution of 0.5m or better, as checked by a resolution check slide.3.2.12 minimum reporting limi
23、t (fs/sample); minimum reporting limit (fs/m3), nthe lowest result to be reported for total sporesor any spore category. Since both fs/sample and fs/m3 are reported, there are two minimum reporting limits.3.2.13 morphology, nthe form and structure of an organism or any of its parts; for fungi, the s
24、hape, form, and/orornamentation. ornamentation, or combination thereof.3.2.14 mounting medium, na liquid, for example, lactic acid or prepared stain, used to immerse the sample particulate matterand to attach a cover slip to an impaction sample.3.2.15 sample trace, nthe area of particle deposition,
25、that is, the deposit on a slit impaction sample resembling a narrowrectangle, or the circular deposit on a circular impaction sample.3.2.16 septum (pl.: septa), na cell wall or partition.3.2.17 slide adherent, nan adhesive or liquid used to affix an impaction sample substrate to a microscope slide.3
26、.2.18 slit impaction sample, na sample of airborne particulate matter collected via by means of a device that draws airthrough a linear aperture at a specified rate, impacting the particles suspended in the air onto an adhesive medium, resulting in arectangular area of deposition. A slit impaction s
27、ample may be collected via by means of a cassette manufactured for that purpose, or via purpose, or by means of a sampling device that requires slides to be pre-coated with impaction medium.3.2.19 spore category, na grouping used for identification and quantifation of fungal structures.Aspore catego
28、ry may containa specific genus (for example, Stachybotrys), or it may represent a combination of genera (for example, Aspergillus/Penicillium-like).3.2.20 traverse, na portion of analysis of an impactor sample consisting of one scan under the microscope from a sample-lessportion of the impaction med
29、ium across the deposit to a corresponding sample-less portion of the impaction medium on the otherside.3.3 Symbols:3.3.1 fsfungal structure3.3.2 fs/m3fungal structures per cubic metre3.3.3 m3cubic metre3.3.4 mmmillimetre3.3.5 mmicrometreD7391 1724. Summary of Test Method4.1 Samples have been previou
30、sly collected utilizing an impaction device operating at the device manufacturers recommendedsample flow rate. Each sample consists of an optically clear substrate coated with an adhesive and optically transparent mediumonto which particles have been deposited through inertial impaction.4.2 A sample
31、 is mounted to a microscope slide and examined by bright field microscopy using at least two magnification/resolution combinations.4.3 Spores are differentiated from each other, other fungal structures, and from non-fungal material by color, size, shape,presence of a septum or septa, attachment scar
32、s, surface texture, etc., by means of a taxonomic comparison with standard referencetexts and/oror known standard samples (see samples, or both (see Section A1.1 for suggested references). The number of sporesthat match each spore category are then calculated in units of fungal structures per sample
33、 (fs/sample) and also fungal structuresper cubic meter of air (fs/m3).5. Significance and Use5.1 This test method is used to estimate and categorize the number and type of fungal structures present on an inertial impactorsample.5.2 Fungal structures are identified and quantified regardless of whethe
34、r they would or would not grow in culture.5.3 It must be emphasized that the detector in this test method is the analyst, and therefore results are subjective, depending onthe experience, training, qualification, and mental and optical fatigue of the analyst.6. Interferences6.1 Differentiation of Fu
35、ngal Genera/SpeciesBecause of the similar size and morphology of some fungal spores of differentgenera and the absence of growth structures and mycelia in airborne samples, differentiation by microscopic examination aloneis difficult and spores must be grouped into categories based strictly on morph
36、ology. In many cases, identification at the genus levelis presumptive. For example, differentiation between Aspergillus and Penicillium using this test method is not typical, so acombined Aspergillus/Penicillium-like category is used. When differentiation between such genera is desired, a different
37、testmethod must be used. Unequivocal identification of every spore in each category is not possible due to optical limitations, theatypical nature of some of the spores, and/or overlapping morphology among different spore types, or combination thereof, andtherefore, certain spores must be categorize
38、d as Miscellaneous/Unidentifiable.6.2 Look-alike Non-fungalLook-Alike Non-Fungal ParticlesCertain types of particles of non-fungal origin may resemblefungal spores. These particles and artifacts may include air or plant resin bubbles, starch, talc, cosmetic particles, or combustionproducts. Standard
39、s (mounted similarly to impactor samples) should be examined by laboratory analysts to know how to identifysuch particles. Examination of suspect particles using optical conditions other than bright field microscopy (for example, polarizedlight microscopy, phase contrast microscopy, differential int
40、erference contrast) may be helpful whenever significant concentrationsof look-alike particles are present. In some cases dust and debris can mimic the morphology of particles of interest. Whenlook-alike particles are present in high concentration, accurately counting spores with similar morphology i
41、s difficult. When theseconditions exist, they should be reported in the analysis notes section of the report.6.3 Particle OverloadingHigh levels of particulate matter on an impaction sample will bias the analysis in two ways:(1) Particle capture efficiency decreases, and(2) Debris obscures or covers
42、 spores.Both of these factors produce a negative bias.6.4 StainingStaining, while optional, may help the analyst differentiate spores from debris. Without staining, clear spores(especially small ones) may exhibit negative bias because the analyst has insufficient contrast to notice them while scanni
43、ng.Also,because spores of different fungal species absorb stains at different rates, under or over-staining makes identification difficult. Theproblem can be eliminated by careful control of stain concentrations.6.5 Impaction Medium Stability and ClarityChemicals present in some mounting media may a
44、ffect the physical stability orclarity of the impaction medium. For instance:(1) Samples collected on silicone grease medium should first be warmed on a hot plate at approximately 40C to “fix” thesample in place, when using lacto-phenol cotton blue stain, and(2) Slides and cassettes using methyl cel
45、lulose ester + solvent adhesive medium, which is stable in lacto-phenol cotton bluestain, will “fog” with Calberlas stain due to the water and alcohol mixture; warming fogged slides may temporarily clear them.The lab or analyst should develop through experimentation an impaction medium/mounting medi
46、um combination that will resultin acceptable stability, clarity, and spore visibility.6.6 Uneven Impaction Medium UniformityUneven thickness may be present in greased slides, pre-coated slides andmanufactured cassettes. The microscopist will compensate by adjusting the plane of focus. When grease is
47、 too thick, differentiatingD7391 173small spores from background artifacts (especially air bubbles) in the grease preparation becomes difficult. When grease is too thin,shrinkage and pooling may have occurred, causing particle loss during sampling.7. Apparatus7.1 Marking pen, for marking sample slid
48、es.7.2 Microscope or magnification system, having a precision x-y mechanical stage. The microscope or magnification system usedfor analysis shall be capable of at least two magnification/resolution combinations as follows: magnification/resolutioncombination 1 shall be 150-400150400 total magnificat
49、ion and a point to point resolution of 0.7 m or better;magnification/resolution combination 2 shall be 400 or greater total magnification and a point to point resolution of 0.5 m orbetter. It is recommended that at least one microscope or magnification system in the lab be capable of magnification of 1000total magnification and a point to point resolution of 0.3 m or better. That the resolution for combinations 1 and 2 is suitable isto be checked using a resolution check slide (see 13.2.3).7.3 Reference Slidesa series
