ASTM_D_3862_-_13.pdf

Designation:D386213Standard Test Method forRetention Characteristics of 0.2-m Membrane Filters Usedin Routine Filtration Procedures for the Evaluation ofMicrobiological Water Quality1This standard is issued under the fixed designation D3862;the number immediately following the designation indicates 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 a procedure to test membranefilters for their ability to retain bacteria whose diameter is equalto or slightly larger than the 0.2-m pore size of the membranefilter.1.2 The procedures described are for the use of userlaboratories as differentiated from manufacturers laboratories.1.3 The values stated in SI units are to be regarded asstandard.No other units of measurement are included in thisstandard.1.4 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 and health practices and determine the applica-bility of regulatory limitations prior to use.2.Referenced Documents2.1 ASTM Standards:2D1129 Terminology Relating to WaterD1193 Specification for Reagent Water3.Terminology3.1 Definitions:3.1.1 For definitions of terms used in this test method,referto Terminology D1129.3.2 Definitions of Terms Specific to This Standard:3.2.1 Grams stain,na routine bacterial stain that dividesbacteria into two categories,depending on whether they can bedecolorized with acetone,alcohol,or aniline oil after stainingwith one of the rosaniline dyes such as crystal violet,methylviolet,or gentian violet and treating with iodine.Those thatresist decolorization remain blue or violet and are designatedGram-positive;those that are decolorized and take up the redcounterstain,such as neutral red,safranin,or dilute carbolfuchsin are termed Gram-negative.3.2.2 vacuum,nfor the procedure used,source of suctionthat can produce a reading of 500 to 600 mm Hg on a vacuumgage.4.Summary of Test Method4.1 This test method is based on the cultivation of organ-isms whose diameters are equal to or slightly larger than poresof the membrane filter to be tested and then filtering a specificaliquot containing organisms through the membrane followedby an examination of the filtrate after incubation for sterility.Asterile filtrate indicates complete retention of the organism andvalidates the ability of the membrane to retain bacteria equal toor slightly larger than the stated pore size.5.Significance and Use5.1 Microbiological water testing procedures using mem-brane filtration are based on the premise that all bacteria withina specific size range will be retained by the membrane filterused.If the membrane filter does not retain these bacteria,falsenegative results or lowered density estimates may occur thatcould have serious repercussions due to the presence ofunrecognized potential health hazards in the water being tested,especially in drinking water.5.1.1 This procedure as devised will enable the user to testeach membrane filter lot number for its ability to retain allbacterial equal to,or larger than,the stated membrane poresize.5.2 Since this membrane is often used to sterilize nonauto-clavable liquids,it is essential that the retention characteristicsof this membrane are stable.6.Apparatus6.1 Membrane Filtration Units,six.6.2 Vacuum Source,with trap vessel.1This test method is under the jurisdiction of ASTM Committee D19 on Waterand is the direct responsibility of Subcommittee D19.08 on Membranes and IonExchange Materials.Current edition approved Feb.15,2013.Published July 2013.Originallyapproved in 1990.Last previous edition approved in 2011 as D3862 80(2001),which was withdrawn July 2010 and reinstated in February 2013.DOI:10.1520/D3862-13.2For 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 Harbor Drive,PO Box C700,West Conshohocken,PA 19428-2959.United States1 6.3 Filtering flasks,1-L,with vacuum tubing into which aglass tube and aY-tube have been incorporated as in Fig.1.Thefree end of the Y-tube is connected by tubing to a sterilebacterial air vent.The tubing to air vent is clamped shut duringfiltration and released after filtration.6.4 Forceps,blunt-nosed,and small beaker of 95%ethanol.6.5 Incubator,37C.6.6 Pinch-Cock Clamps.6.7 Autoclave or Other Sterilizing Equipment.6.8 Appropriate Equipment for producing reagent gradewaters.6.9 Appropriate Laboratory Glassware.6.10 Sterile Rubber Stoppers,to fit 1-L filtering flask.6.11 Expendables:6.11.1 Double-Strength Broth,140-mL aliquots.6.11.2 Sterile Pipets,1 and 10-mL.6.11.3 Sterile 0.1%Peptone,in