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October2009ClinicalLaboratoryNews:NewParadigmsinFetalLung MaturityTesting

October2009:Volume35,Number10 NewParadigmsinFetalLungMaturityTesting WhatAretheOptionsforLabs? ByGennaRollins Sincetheearly1970s,fetallungmaturity(FLM)testinghasplayedanimportantroleinidentifyingneonatesatrisk ofrespiratorydistresssyndrome.Thevariousassays,developedinthe1970sand1980s,maintainedtheir popularityintothe1990s.However,FLMtestinghasbeenonthewaneinrecentyearsasphysicianshavefound theresultslessinformativetotheirdecisionmakingaboutwhentotimedelivery.Evenso,informationaboutFLM stillhasaplaceincertainsituations,andthepotentiallossofoneoftheassayswillrequirelaboratoriansand clinicianstocollaborateaboutavailableoptionsanddevelopcontinuityplans. Labscanreallyplayanimportantroleinthistransitiontime,saidDavidGrenache,PhD,DABCC,assistant professorofpathologyatUniversityofUtahSchoolofMedicineandmedicaldirectorofspecialchemistryatARUP LaboratoriesinSaltLakeCity.Itsagoodopportunityforthemtoworkintandemwithdoctors,notonlytoadapt tothechangingenvironment,butalsotohavediscussionsaboutFLMtestutilizationandhowandwhyitsbeing used.

ALookatTestUtilization
AbbottisoneoftheprimarymanufacturersofFLMtestsanditsTDxFLMIIassayisoneofthemorepopulartesting options.However,thepotentialdiscontinuationofthisplatformhaspromptedresearcherstoreviewtheutilization ofandindicationsforFLMtesting.AlthoughthecompanyhasnotmadeaformalannouncementaboutitsTDxFLM IItestforsurfactant/albuminratio(S/Aratio)byfluorescentpolarization,anAbbottspokespersonindicatedthatit isdecommissioningcertainlegacysystems.Abbottisretiringmanyoftheassaysthatrunonthecompanysolder diagnosticinstrumentplatformsIMx,TDxandTDxFLx.Theseplatformsaremorethan20yearsoldanduse olderequipmentandtechnologythatisdifficulttosupportandserviceaspartsandrawmaterialsarescarce.The decisiontoretiretheseolderproductswillenableAbbotttoinvestinstateofthearttechnologies,saidAnnFahey Widman,directorofglobaldiagnosticpublicaffairs.WehavenotmadeanannouncementregardingtheAbbott FLMassayonTDxaswe'recurrentlyevaluatingthisandotherproductsinternally,[but]whatIcantellyouisthat wearecommittedtocommunicatinganyadditionalassayretirementstoourcustomersatleast12monthsin advanceandwillcontinuetohelpcustomersthroughanytransitions. ShouldtheS/Aratioplatformbecomeunavailable,itappearsthatmanylabswillbeaffected,accordingtoasurvey ofAACCmembersconductedthisspringbyGrenacheandAnnGronowski,PhD,DABCC,FACB,associateprofessor ofpathology,immunology,andobstetricsandgynecologyatWashingtonUniversitySchoolofMedicineinSt.Louis. TheirresultsindicatedthattheS/Aratiowasbyfarthemostfrequentlyofferedtest,followedby phosphatidylglycerol(PG)byagglutination,lecithin/sphingomyelinratio,lamellarbodycount,PGbythinlayer chromatography,and(rarely)foamstability.CAPsurveydatafor20072008indicatedroughlythesameutilization pattern,exceptitdidnothavedataonlamellarbodycountandfoamstabilitytests.

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ChangingVolumes,PracticePatterns
ThepotentialretirementoftheTDxplatformmayposeadilemmaforsomelabs,butperhapsnotasmuchasit mighthaveinthepast.GrenachesandGronowskissurveyshoweda10yeartrendofdecreasingFLMtest utilization,andinaseparateanalysis,UniversityofMississippiMedicalCenterresearchersreportedanoverall declineinFLMtestvolumeintheirlabfrom1,090in1994to389in2004(JPerinatol200828:2023).Theauthors alsosurveyedothermaternalfetalmedicinepracticesandfoundthattherewasanetdecreaseintestutilization forthreeoffourFLMtests,rangingfrom33%forfoamstabilityto56%forPG.TheexceptionwasAbbottsTDx FLMII,forwhichrespondentsreportedanetincreaseof29%.Thisspring,GrenacheandGronowskialso surveyedfellowsoftheSocietyforMaternalFetalMedicine.Althoughtheresponseratewasquitemodest(<5%), theresultsindicatedthatonequarterofparticipantswereorderingthetestslessoften. ThedeclineinFLMtestingstemsfromseveralfactors,accordingtoThomasMcElrath,MD,PhD,assistant professorofobstetricsandgynecologyatHarvardMedicalSchoolandBrighamandWomensHospitalinBoston. Multipletrendshaveeachcontributedtotheformertestingparadigmthathadbeendominant,heobserved.We werehopingatonepointthatfetallungmaturitywasamarkerforoverallmaturity.Wenowrealizethatitsless thatandmoreanissuespecifictothelungs.Fetallungmaturityisstillanimportantissue,butwewereprobably creditingitwithprovidingustoomuchinformationatanearliertimeandtodayhaveputitinamoreproper context. TrendsMcElrathcitedascontributingtothedeclineincludedstudiesindicatingthatthewindowofutilityforFLM testingismorenarrowthanmostofusexpecteditwouldbe,andmedicalpracticesthathaveenabledsafer deliveryofpretermbabies.NICUtechnologyhasimproveddramaticallyandaneonatologistcannowsupporta babyat34weeksmuchmoresafelythanheorshecouldhavedonetwodecadesago,hesaid.Inaddition, researchhasilluminatedtheoveralldysmaturityofneartermprematurebabies.Werealizethattheresmoreto maturitythanjustlungs.Theuseofsurfactanthasincreasedsodramatically,anditsbeenarevolutionintreating thesekids.Buttherearesomanyotherproblemsthatarentbeingtestedforlikenecrotizingenterocolitis, intravascularhemorrhages,andbehaviorproblems,thatFLMisstillinformativebutitsperhapslessusefulthanwe hadthoughtitwas. FLMtestingnotbeingneededforpatientcarewasbyfarthemostcommonreasoncitedforthedeclineintest utilizationreportedintheJournalofPerinatologystudy.Cliniciansaregoingmoreonclinicaljudgment,andtheyre notdoingamniocentesisasoftenisthebottomline,observedseniorauthorJohnMorrison,MD,professorof obstetricsandgynecologyandpediatricsatUniversityofMississippiMedicalCenterinJackson.

UpdatedGuidelinesClarifyIndications
UpdatedAmericanCollegeofObstetriciansandGynecologistspracticeguidelinesnotethatwhileFLMstatuscan assistcliniciansindeterminingwhendeliveryshouldoccur,FLMtestingshouldnotbeperformed,andis contraindicated,whendeliveryismandatedforfetalormaternalindications(ACOGPracticeBulletin,97, September2008).ThomasPeng,MD,amaternalfetalmedicinespecialistatBonSecoursRichmondHealthSystem inRichmond,Va.agrees.ThewayIputitis,ifyourstrategyformanagementisgoingtobetodeliverregardless ofthetestresults,thentheresnopointindoingthetest.Youusethetestwhenyouhavetheoptiontowaitto deliver.Forexample,amomwithdiabeteswithamacrosomicbabywhereyoureconcernedaboutfetalloss,youd doanamniocentesisat37weeks,andifthetestindicatedlowriskofneonatalrespiratorydistress,youcould proceedanddeliverthebaby,hesaid.PengservedasconsultanttoACOGinupdatingthepracticebulletin. OneofthemostcommoncircumstancesinwhichMcElrathusesFLMtestingiswhenthemotherhasruptured membranesandherpregnancyisatleast32weeksgestation.Itsbeendemonstratedinthemedicalliterature thatifababyhasmaturelungs,after32weekstheoverallriskofmorbiditytothebabyislowertogoaheadand deliverthantocontinuetomanageitinutero,heexplained.ThatscenariowasasmallpercentageoftheFLM

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testingweusedtodobuthasnowgrowntoapproximately50percentofalloftheFLMtestsweorder. TheACOGguidelinescallforFLMtestinginscheduleddeliveriesatlessthan39weeksunlessfetalmaturitycanbe deducedfromothertestsperformedearlierinthepregnancy,includingultrasound,Dopplerultrasonographyto detectfetalhearttones,andpositiveserumorurineHCGpregnancytestresults.Likewise,NACBsLaboratory ManagementPracticeGuidelineonMaternalFetalRiskAssessmentandReferenceValuesinPregnancystatesthat FLMtestingisnotindicatedinnormalpregnanciesifthegestationalageisaccuratelyknowntobeatleast36 weeks. AnothertrendinFLMtestingisthatrepeattestingislesscommonnow.TheupdatedACOGpracticebulletinnotes alackofconsensusaboutthebenefitsofrepeattestingwhenthefirstresultsareeitherimmatureor indeterminate,achangefromACOGspriorrecommendationin1996.Thatchangewasmadefortwopractical reasons.First,mostpractitionersingeneraldontdosequentialtesting,Pengexplained.Also,onestudy evaluatedtherateofincreaseinTDxFLMIIresultsandfoundittobeaconstantofapproximately15milligrams pergramperweek.Sodependingontheinitialvalue,onecouldextrapolatetodetermineifbywaitinganother weekonaveragetheFLMtestwillbemature.

FLMTestOptions
ForlabsthatoffertheS/Aratiotest,thekeyistobeginreviewingFLMtestingoptionsnow,accordingto Gronowski.Withoutadoubt,labsneedtostartthinkingaboutthisandworkingtogethernow.Mostofusdont haveahugevolumeofthistesting,andovertimeithasdeclined,sothoseofuswhoneedtovalidateatestwill needtohelpeachotherandsharesamples,shesaid.Includingcliniciansearlyonisamust,advisedPeng.Ifyou springanewtestonthematthelastminute,theywillhavenofaithintheparticulartestthatyoureoffering becausetheywillneverhaveuseditbefore.Inordertofostertheirtrust,youhavetogetthemtogetherand developaplan,henoted. AccuracyandtimelinessarethetwomostimportantfactorsinphysicianpreferenceforFLMtests,accordingto Peng.Thetesthastobereproducibleandreasonablyaccurateintermsofpredictingrespiratorydistress syndrome,hesaid.Werealsolookingforashortturnaroundtime.MostrespondentstoGrenacheand GronowskissurveyofSocietyofMaternalFetalMedicinefellowsindicatedtheywouldlikeFLMresultswithin12 hours.NACBLMPGrecommendsthatFLMtestingbeavailableinhospitalsthatdeliverinfants,withresultsavailable routinelyonceperdayandinurgentsettingswithinanhourofcollectingthespecimen. AccordingtoNACBLMPG,allFLMtestshaveahighatleast95%sensitivity,butmediocrespecificity.Takingthe prevalenceofrespiratorydistresssyndromeintoaccount,thetestshaveahighnegativepredictivevalue,meaning theprobabilityoftheneonatehavingrespiratorydistresssyndromeislowwhenresultsindicatethelungsare mature.Thepositivepredictivevalue,orlikelihoodofrespiratorydistresssyndromewhentheFLMtestindicates thelungsareimmature,rangesfrom13%to61%,accordingtoACOG.Veryfewbabieswithmatureresultsgo ontodeveloprespiratorydistresssyndrome,Grenacheexplained.Butthetestsimmature,orpositive,predictive valueisnotgoodatall.Therearelotsofbabiesbornwhohadanimmatureresultandneverdeveloprespiratory distresssyndrome.Meconiumandbloodcontaminationcanaffecttheperformanceoftheteststovaryingdegrees, dependingontheassay. IntheirsurveyofSocietyofFetalMaternalMedicinefellows,GrenacheandGronowskiaskedrespondentstogive theirimpressionoftheclinicalaccuracyofthevariousFLMtests.RespondentsratedS/AratioandPGbyTLC equallyhigh,with>90%reportingtheteststobeexcellentorgood.Nearly90%alsorankedtheL/Sratioaseither excellentorgood,whereasabouttwothirdsratedPGbyagglutinationandlamellarbodycountaseithergoodor excellent.Foamstabilitywasadistantsixth.GrenacheandGronowskihavealsopublishedareviewarticleabout FLMtesting(ClinBiochem200639:110)seetable,below,forasummaryofthevarioustests.

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Table1 FetalLungMaturityTests Mature Predictive Value (%) 95 Immature Predictive Value (%) 15

Test Foam Stability Index

Platform Labdeveloped

Pros Rapid

Cons Bloodand meconium affectresults Qualitative Technically challenging

Lamellar Body Count

Labdeveloped

Rapid,simple Instrumentation widelyavailable

Bloodand meconium affectresults QC/PTnot available until2010

9798

2935

L/S Ratio

TLC(lab developedor commercially available)

Quantitative QC/PTavailable

Bloodand meconium affectresults Technically challenging Time consuming

86100

1983

PG

AmnioStat FLM (Agglutination)

Rapid,simple QC/PTavailable Bloodand meconiumdo notaffect results

PGlast surfactantto rise Subjective test interpretation Singlevendor

97100

950

PG

TLC(lab developedor commercially available)

QC/PTavailable Quantitative Bloodand meconiumdo notaffect results

PGlast surfactantto rise Technically challenging

95100

2353

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S/A Ratio

TDx

Rapid,precise Quantitative Standardized QC/PTavailable

Bloodand meconium affectresults Singlevendor

99100

1436

Legend:S/ARatio=surfactant/albuminratioPG=phosphatidylglycerolL/SRatio= lecithin/sphingomyelinratio AllbutthreeFLMtestsarelabdeveloped,soanotherconsiderationforlabsinchoosingatestishowlabor intensiveandtechniquedependentthemethodsare.L/SratiobyTLCcanbeperformedaloneorincombination withPGbyTLC.Boththetestsareoldstandbysbuttheyalsorequireanexperiencedpersontoknowhowto prepareandreadthegels,Pengobserved.Alabheworkedwithstoppedofferingthetestbecauseitlosttheone personwhohadexpertiseindoingthegels,andwhenhewasgone,thatwasit.Ingeneral,thetestistime consumingandmanylabsdonothavevolumesrecommendedtomaintainproficiencywiththeassay,accordingto Gronowski.AcommercialversionoftheL/Sratio,FetalTek200,isavailablefromHelenaLaboratories. Anotherlabdevelopedtestthatafewlabsofferisthefoamstabilityindextest.Ithastheadvantageofrapid results,butisonlysemiquantitativeandcanposetechnicalchallengesowingtotheuseofhygroscopicethanol. Theassayalsoishighlyoperatordependent.PhysiciansatamajorteachinghospitalinformedPengthatthey preferredtheassaybutthattherewasonlyonepersoninthelabtheytrustedtodothetest.Relyingonatest thatissotechnologistdependentisarecipefordisaster,accordingtoGronowski.Withourautomatedtestswe trynottohaveasingleinstrumentthatdoesatest,sowhywouldalabhaveasingleemployeethatcandoatest? sheasked.Bythesametoken,labscanbeequallyvulnerableiftheyofferonlyoneofthethreecommercialFLM tests. IrvineScientificsAmnioStatFLMPGbyagglutinationtestgivesrapidresultsandiseasytoperform.HoweverPGis thelastsurfactanttoincrease,atabout36weeks,andmakesuparelativelymodestproportionofoverall pulmonarysurfactant,soamatureresultiscloselytiedtotimingofthetest.Inaddition,themanufacturertwice hasexperiencedproductionchallenges,onceduringmuchof2004,andforaperiodofaboutthreemonthsthis year.Theproblemswereunrelatedandhavebeenfullyresolved,accordingtoJanieKim,seniorproductmanager.

TheCaseforLamellarBodyCount
AwareofthelimitationsofthevariousFLMtests,aswellasthepossibilitythatAbbottmightonedayretirethe TDXFLMIIplatform,GrenacheandGronowskibeganexploringotheroptionssometimeago.Theirresearchhas leadbothtoconcludethatthelamellarbodycountassayisthebestavailablealternative.Lamellarbodiesare hydrophobicstructuresmadeofsurfactantsandsynthesizedintypeIIpneumocytes. AlthoughGronowskiisbullishonthetestnow,thatwasntalwaysthecase.IhadanFDAapprovedtestthathad externalproficiencytesting,standardizedmethodsandqualitycontrol,sowhywouldIwanttosubstitutethatfor somethingIwasgoingtohavetovalidateinhouse,thatIdidnthaveexternalproficiencytestingforandthere wasntalotofdataon?sherecalled.However,overtime,bycompilingdataIveconvincedmyself,andhopefully others,thatclinicallyitperformsaswellastheS/Aratio. Thetest,whichinvolvescountinglamellarbodiesusingstandardhematologycounters,isquickandeasyto perform,andcanbedonewithsmallsamplevolumes.However,itisalabdevelopedtestthatwillrequireinternal validationwithhardtocomebyamnioticfluidsamples.Inaddition,bothlaboratoriansandcliniciansarenotas familiarwithitasotherFLMtests,accordingtoGrenacheandGronowskissurveys.

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Lamellarbodycountperformedonhematologycounters usesanamnioticfluidsample.CAPwillofferproficiencytesting forthisanalytenextyear. PhotocourtesyofAnnGronowski,PhD Thetwomainhurdlestoimplementingthetestareovercomingtheperceptionthatitisinsomewayinjuriousto cellcountersandaccumulatingenoughamnioticfluidsamplestovalidatethetest.Lamellarbodycountis quantitativeandeasytodo,andIdbewillingtobetthateverylabinthecountryhasacellcounter.Butpeopleare afraidofrunningsamplesthroughcellcountersthatarenotwholeblood,Grenacheobserved.However,amniotic fluidisnotgoingtoharmthecellcounter. Itmaytakeawhileforlabstoaccumulatesufficientamnioticfluidsamplesforvalidation,butthegoodnewsisthe fluidisverystable,itcanberefrigeratedandyoudonthavetorushitoffanywhereordoanyspecialhandling,so youcanjustholdituntilyouhaveenoughsamples,explainedGronowski.Sherecommendsthatlabsinterestedin implementingthetestpoolsamplestosupporttheirvalidationrequirements.CAPhasannouncedthatitwillbegin offeringproficiencytestingfortheassaynextyear. RegardlessofwhichFLMtest(s)labsofferinthefuture,itisimperativethattheystartthereviewprocessnow, accordingtoGronowski.LabshavetoopenadialoguewiththeirObGyns.WhethertheirdecisionistosendFLM testsoutordeveloptheminternally,honestly,itcouldtakeawhiletogeteverythingworkedoutandready. Drs.GrenacheandGronowskihavereceivedconsultantfeesfromAbbott.

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