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WHAT YOU NEED TO KNOW ABO U T W O RKI N G IN A. LAB- GENE R AL L A B OR G ANIZ A TION. D r . Fer n T sien Depart m ent o f Genetics LSU H SC. Wha t typ e o f laborator y do yo u wor k in?. In gener a l, labs ca n be d e scribed as:

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What you need to know abo u t w o rki n g in a

WHATYOUNEEDTOKNOW

ABOUT WORKING INA

LAB-

GENERALLABORGANIZATION

Dr.FernTsien DepartmentofGenetics LSUHSC


What you need to know abo u t w o rki n g in a

Whattypeoflaboratorydo

youworkin?

  • Ingeneral, labscanbedescribedas:

    • BasicScience–the researchersdo sciencefor the sakeof knowledge.Forexample,howdoesa specific transcription proteinwork?

    • Clinicalresearch–humanpatients or their samplesare usedto investigatea diseaseor syndrome.


Wher e i s you r la b located

Whereisyourlablocated?

  • Eachlabispartofa DepartmentorCenter.

  • Large piecesofequipmentareoftensharedbythe department membersand arehoused inaseparatelabor possiblyyourlab.

  • Thesecan includeultra-cenrifuges, liquid

  • nitrogentank,autoclaves,darkrooms, etc.

  • Alsothedepartmentmay haveaconference room andakitchen.

  • Makesure youcheckif you can usethesefacilitiesbefore goingahead.


What you need to know abo u t w o rki n g in a

KNOWYOURLABCOWORKERS


Principa l investigato r pi

PrincipalInvestigator(PI)

  • Peopleoftenask:Who is yourPI?

  • Thisisthe headof the lab, the boss,your

  • advisor.Mostprobablyyourmentor.

  • Thispersonoftenspendstimeintheir officewriting grantsor researchreports andmayspendlesstimeinthe lab.

  • Iftheyare clinicians, theymayalso see

  • patientsinadditionto reportwriting.

  • However,this personisthe intellectual guidebehindmostof theprojectsinthe lab

  • PIsare responsibleforfundingthelab research.


What you need to know abo u t w o rki n g in a

•I

I


What you need to know abo u t w o rki n g in a

Postdocs

  • LabPost-doctoral fellow

  • Thispersonhas recentlyreceivedtheirdoctorate(usuallyPhD)andisdoingatrainingperiodbefore becoming aPI

  • Theyusuallywork ontheirownproject, butwillcollaboratewith otherlab membersonvariousparts oftheproject.


What you need to know abo u t w o rki n g in a

Technicianorresearch

assistant

  • Canbeacollegestudentwho wantsto gainmoreexperienceinalabbefore enteringmedicalorgraduateschool

  • Canbe aprofessionalwithamasters degreeandappropriatepayandtitle, basedonyearsofexperience

  • Theydoavarietyoftasksincluding: orderinglabsupplies,preparingmedia,caringfor thelab’scelllines,assistingthelabwithexperiments,and theycancarry theirownexperiments.


La b supervisor

Labsupervisor

  • The day-to-dayoperationofthelabis

  • sometimesoverseenby a labsupervisor.

  • Thispersonusually hasamastersdegreeand extensivelabexperience.


Graduat e students

Graduatestudents

  • Theyaredoinglabworktoget

  • theirMastersdegreeor PhD.

  • Theyusuallyworklong hours andspenda lotof timeinvestedin theirspecificproject.

  • Theybecome increasingly independent duringtheir4-5 yearsinthelab.


Resident s an d fellows

Residentsandfellows

  • They havean MDandarespendingsometime(weeks,months,sometimesyears) traininginan areaof his/herfieldbeforethey becomeindependent


Workin g hours

Workinghours

  • Becauseexperimentsdonotfitintoaslotof9-5,lab workershaveunpredictableandsometimeseccentric hours.Thisdoesnotapplytoyou!

  • Clinicalresearchalsodependsontheclinicschedule.

  • Checkwithyourmentorwhatworkinghoursare

  • expectedofyou andtryto conformtothis.

  • Pleasecommunicatewithyourmentorandlab personnel whatyourvacation plansare.

  • If youaresick,pleaseletthemknow.

  • Even thoughyoudonotgetpaidbythehour,youneed toletthemknowoutofcourtesy.


La b meetings

Labmeetings

  • Largelabsusuallyhavemeetingsto discusstheresearchof

  • eachmemberofthe team.

  • Inthesemeetings,sometimesonly one or two peopleare assignedtotalkeachweek, orallmembersare expected to speakbriefly.

  • Somemeetingsare casual, and someareformalwitha slide

  • projector.

  • Youmaybe askedto participate.Ask yourmentorahead of timeif he/she expectsyouto participate so youcan prepare afew statementsorquestionsyou mayhave.

  • Check out theatmosphereandaskquestionsduring appropriatetimes.

  • DO:take notesandlookuptermsyou maynot

  • understand

  • DONOT:fallasleep,text messageduringthemeeting


Th e firs t week

Thefirstweek

  • Bynow youshouldhavebeenassignedadesk

  • orlab bench.

  • Your mentor shouldhave discussedyour projectwithyou.

  • Ifanother labmember offers to teach you, go

  • aheadandlearnfromthem.

  • Youmayget alabkey.Becareful withit!

  • Yourresearchmaybe slowrightnow while youwaitforreagents tobeordered,cellsto grow,etc.


Wha t t o d o o n the firs t fe w weeks

Whattodoonthe

firstfewweeks

  • Setupandorganizeyourlab benchor desk

  • Introduceyourselfto everyone in thelab if the mentorhas notdoneso.

  • TAKENOTESON EVERYTHING!Youdo not wantyour instructorsto keeprepeatingthemselvesortheywillget annoyed.

  • Familiarizeyourselfwith where thingsare kept.

  • ASK.Ofcourse,youdo not want tobotheranyone unnecessarily,but itis muchbetter to ask about a procedure,a reagent,equipment,etc. than towastedtime andmoney.

  • Ifyoumakea mistake,it’sOK.Justmake sure youlearnand

  • askcoworkersforhelp.

  • Askyourmentorforrelevantliterature abouttheproject.Thiswillimpressthem!


Wha t no t t o d o o n your firs t fe w weeks

WhatNOTtodoonyour

firstfewweeks

  • Don’tsay“we did not do it thisway inmyprevious job”. Eachlab does experimentstheir ownway (example:cellculture)

  • Donot propupyour feetandread,playcomputergames, takeanap,or lookat videos onYoutube.Itistruethatat thebeginning,there willbe somedead(non-experiment)time untilthe experimenttakes off. However, playingwhileothersare workinghardwillnot makea goodimpression. Insteadtryto readabout yourresearch topic.

  • Donotaskand/orcomplainto otherstudents or co-workersaboutmoney

  • orsalaries.Itcanirritate mentors andotherco-workers.

  • Don’tsayyouare workingina labfor anyother reasonthanfor interestin the field. Ifyousay youare workingtherejustfor the money, you willnot betakenseriouslyanditcancostyouyourjob.


What you need to know abo u t w o rki n g in a

Basicsurvivalrules:

Attitude

  • Ask,donot command.Othersare takingthe trouble tohelpyouanddo not HAVEto doit.

  • Becourteoustoeveryone,not just your mentor.

  • Donot assumethatsomeoneis goingto dropwhat

  • theyare doingto helpyou.Bepatient.

  • Writedowneverythingwhensomeoneisgiving you instructions.You arenot expectedto remember everything,andthat person is not expectedto repeateverythingoverand overagain.

  • Makeappointmentswithbusy people.Some mentorsarealso directorsof adepartmentor center, andare hardto knowwhenthey are available for themto focus on whatyouhaveto say.


What you need to know abo u t w o rki n g in a

Basicsurvival

rules:Courtesyat

ScienceFriction

thelabbench

  • Donot use reagentsinsomeoneelse’slab bench without asking.

  • Ifeveryonesharesa reagent andyoufinishit,let someoneknow.Donot placethe emptybottleback on theshelf.

  • Ifsomethingbreaks,tellsomeoneimmediately.You will notgetintroubleunlessyou tryto hideit.

  • Ifyoudo somethingwrong,confess.Everyonemakes mistakes,but sneakingto covera mistakeis unacceptable.

  • Clean up aftertheexperiment.Donot leaveit for the technicianor graduatestudent todoit.


What you need to know abo u t w o rki n g in a

Checkyourlab’sownSafety

rules

  • No eating,drinking,orsmokingin thelabunless

  • approvedbyyourmentor.

  • Do notwearopentoedshoesin thelab.Yourfeet willbe vulnerable ifachemicalspillsonyou.

  • No shortsorbaremidriffsareallowedforsafety

  • reasons.

  • Do notmouthpipet,evenwater.

  • Askwhattodoincaseofemergency-wherethefirstaidkitiskept,howtocallforemergency


What you need to know abo u t w o rki n g in a

  • Ifyouhaveanyquestionsor

  • problems,pleasemakesuretolet

usknowearlyonintheinternship.

Dr. Paula GregoryDr.FernTsien

[email protected]@lsuhsc.edu


Today s experiment

TODAY’S EXPERIMENT


W h a t a r e h e l a c e ll s

WhatareHeLaCells?

  • HeLacellsarecervicalcancercellsthatwere

  • obtainedformawomannamedHenriettaLacks.

  • These cellswereobtainedfromabiopsyofthe cervixbyherdoctorwithouther knowledge/consent.

  • Thecellswereimmortalized.

Henrietta

Lacks


Immortalize d cell s an d hela

ImmortalizedcellsandHeLa

  • HeLacellsobtainedfromthebiopsyarestillalive and havebeenusefulin theunderstandingof canceranddisease,aswellasdiscovery of treatmentsand therapy.

  • HeLawasthefirstcelllineimmortalized

  • Was usedin thedevelopmentofpoliovaccine, andtofindlinkbetweenhumanpapillomavirus (HPV)andcervicalcancer


T h e imm o rta l l if e o f henr i ett a l a ck s

TheImmortalLifeofHenrietta

Lacks

  • byRebeccaSkloot

  • Littlewasknow about HenriettaLacksuntilrecently.

  • Recently,RSkloot wrote thebiography ofMs.Lacks:

    • PoorAfricanAmerican womanfromBaltimore

    • Had5children

    • Hadcervicalcancer

    • Had littleunderstandingofher diagnosisofcervical cancer,andnoknowledgeofwhatthedoctoratJohns Hopkins wasplanningto do withherbiopsy.

  • Died ofcancer in 1951lessthan a yearafterdiagnosis


Ethica l implications

Ethicalimplications

  • Most widelyused andpropagatedcelltypeusedfor

  • research.

  • Billionsofdollarsintreatmentandtherapydiscoveriesby researchersandpharmaceutical companies

  • Lastyear, theNationalInstitutesofHealth (NIH)madeanagreementwiththeLacksfamily

  • The Lacks family agreedtoallowresearchersto have limitedaccessto her DNA sequence

  • Lacksfamilyhas controloverher information


Ne w regulations

Newregulations

  • Currently,therearespecificregulationsfor ensuringpatientconfidentialityandtoallow patientstogive consenttousetheircellsfor research.

  • Cellsarenotallowedtobeusedforresearch

  • withoutthepatientconsent.


Hel a i n th e news

HeLainthenews

  • “The HeLaGenome:AnAgreementonPrivacy

  • andAccess”;StatementreleasedbytheNIH

  • HBOtofilmmoviebyOprahWinfrey


Today s experiments

Today’sexperiments:

  • Learn howto micropipette

  • HeLaand normalDNA

  • PCRanalysisforthedetectionof human papillomavirus(HPV)

  • Gelelectrophoresis


What you need to know abo u t w o rki n g in a

WhatisPCR?

  • Thepolymerasechainreaction(PCR)isa fasttechniqueused inmanylabsto "amplify“orcopysmallsegmentsofDNA.

  • Itisone of the mostimportantscientificadvancesinmolecular

  • biology.

  • Itscreator,KaryB.Mullis,wasawarded theNobel Prize for Chemistryin 1993.


W h a t i s pc r u s e d fo r

WhatisPCRusedfor?

  • Onceamplified,theDNAproducedby PCR(PCR product)canbeusedinmanydifferent laboratory procedures.

  • DNAfingerprinting inforensicsandpaternity testing

  • Detectionofbacteria or viruses(HIV/AIDS)

  • Diagnosis ofgeneticdisorders

  • Many types of research!


What you need to know abo u t w o rki n g in a

PCR:PolymeraseChainReaction

  • Usedto makemillionsofexactcopiesof

  • DNAfroma biologicalsample

  • Allowsverysmallsamplesto beanalyzed,

  • suchasasampleofa fewskincells

  • Mustbe verycarefulabout contaminationin

  • thisprocess


What you need to know abo u t w o rki n g in a

InsidethePCR reactiontube:

  • DNAthat youwant to

  • analyze

  • Primers(littlepiecesofDNAthat define thearea thatyouwant to analyze)

  • Nucleotides(A,T,C,G)

  • Taq polymerase(drivesthis

  • entire reaction)

  • Magnesium, salts,etc.(for solutionstability)

  • Sterile distilledwater


What you need to know abo u t w o rki n g in a

Millionsof

DNAcopies

45°C

72°C

95°C

ThePCR machineheatsandcoolstheDNA,primers,andotherreagentsin thetubeandallows theamplificationto takeplace.

This resultsinmillionsofcopies.


What you need to know abo u t w o rki n g in a

1.Restrjctionenzymescleave

DNAintosmaJmersegments

ofvarioussizes.

2.DNAsegmentsare

loadedintowellsina

porous gel.Thegelfloatsin

abuffersolutionwithinachamber

betweentwoelectrodes_

3.wnenanelectriccurrentispassedthroughthechamber.

EBDNAfragmentsmovetoward

tileposilively--cilargedcalhode.

111

I

III

II

4.SmallIerDNIAsegments movefasterandfa.riheir thanlargerDNAsegments_


What you need to know abo u t w o rki n g in a

VariationsofPCRandgel

electrophoresis

  • RT-PCR

  • Q-PCR

  • Western blots

  • Northern

  • blots

  • Etc….


What you need to know abo u t w o rki n g in a

THEMICROPIPETTE


Warning s abou t gel electrophoresis

warningsaboutgel

electrophoresis:

  • THEPOWERSUPPLYGENERATESANELECTRIC CURRENT.DONOTIMMERSEYOURHANDINTHE BUFFER OFTHEGELAPPARATUSWHILETHEPOWER SUPPLYISCONNECTED ANDTURNEDON.

  • UVLIGHTDAMAGESEYES.MAKESURETOKEEP THEUVTRANSILLUMINATORLIDDOWNTOLOOKAT GEL.MAKESURENOTTOOPENTHELIDWHILETHE UVLIGHTISTURNEDON.


Ge l electrophoresis

Gelelectrophoresis

  • Remove thegel combsanddams.

  • Arrangegelonitsgel tray intothe gel electrophoresis apparatus, making sure that thenegativeside(black)is onthesidewiththewells.

  • Addenoughelectrophoresis bufferto submerge thegel.


Ge l electrophoresis1

Gelelectrophoresis

  • Retrievethe PCR reactiontubesfrom the PCR machineandmicrocentrifuge5 seconds

  • Toeachtubeadd5 µl of loadingdyetoeachsample. Mix bypipettingup and down.

  • Loadgelby pipetting10 ulof thesolutioninto each

  • ofthe wells.

  • Writedownthe orderinwhichyouloadedthe

  • samples.


Ge l electrophoresis2

Gelelectrophoresis

  • Attachthecovertotheelectrophoresis apparatusmatchingthecolor codedcablesand electrodes(e.g.,redtoredandblacktoblack).

  • Connectthecolorcodedcablestothepowersupply.Turnthepower supply“ON”andsetthe voltageto“100V.”Press“RUN.”

  • Lookforbubblestomakesureitisworking

  • properly.

  • Letsamplesuntiltheloadingdyehasmoved about¾ofthewaydownthegel(abouthalfanhour).

  • Dr.Gregory’spresentation


Ge l electrophoresis3

Gelelectrophoresis

  • Turn off thepower supply andremovecover of

  • electrophoresisapparatus.

  • Wearinggloves,pickup the gel andtrayand drain theliquidinto the electrophoresisapparatus.

  • Carefullytransferthegelon its trayto theUV

  • transilluminator.Turnoff theroomlights.

  • MAKESURETHE LIDTOTHETRANSILLUMINATOR ISCLOSEDand then turnon theUVtransilluminator. Viewthe gel andphotographusing a cellphone camera.

  • Turn off theUVtransilluminator,discardthe gelina wastebagprovidedbyyour teacherand wipe the transilluminatorwith papertowels.


What you need to know abo u t w o rki n g in a

TypicalPCR

Results

300bp

200bp


What you need to know abo u t w o rki n g in a

Housekeeping

  • EATINGandDRINKINGarenot permittedin labsthatcontain biological,chemicalorradiological materials

  • Disposeoftrashin theappropriate containerwhenitis generatedto preventaccumulation

  • Donotplaceemptybottlesor otherhazardsin walkways

  • Keepchemicalsandglassware awayfromtheedgesof countertops

  • Immediatelycleanupanyspills

GoodHousekeeping

PoorHousekeeping


What you need to know abo u t w o rki n g in a

FireSafety

  • Knowthelocationofthenearestfire extinguisherandemergencyexits

  • Duringa fire,usestairs,notelevators

  • Flamecabinetisrequiredforstorageof 10Gal.ormoreofanyflammable substance

  • Compressedgascylindersmustbe secured

  • Electricalcordsshouldbefreeof

  • breaksorfraying

  • Intheeventofanalarm:

    • Secureanymaterialthatleft unattendedwouldposeahazard

    • Exitproperly

Acceptable

Unacceptable

Acceptable

Unacceptable


What you need to know abo u t w o rki n g in a

EyewashandShowers

  • Knowwherethenearesteyewashandemergencyshower stationsare

  • Ifyou aren’tsureordon’tknowhowtousethem,askyour

  • supervisor


What you need to know abo u t w o rki n g in a

HazardInformation

  • Beawareofhazardlabelsonentrydoorsandonthe doors

  • ofrefrigerators,freezers,microwavesanddewars

  • YoushouldbewearingthecorrectPersonalProtective Equipment(PPE)beforeopeninganydoorwitha hazard labelonit

  • MSDS–MaterialSafetyDataSheets

  • Includechemicalandphysicalproperties

  • Toxicityandhealtheffects

  • Compatibility,safehandlingandstorage

  • Spillandfireresponse


What you need to know abo u t w o rki n g in a

  • Anagentofbiological originthathasthecapacitytoproduce deleteriouseffectsin humans,suchasmicroorganisms,toxinsand allergensderivedfromthosemicroorganisms,andallergensand toxinsderivedfromhigherplantsandanimals.

  • Examples

Bacteria

Fungi

Parasites

Toxins

Viruses

Prions

Rickettsials


What you need to know abo u t w o rki n g in a

A/BSL-1

A/BSL-2

A/BSL-3

A/BSL-4

Lowindividualrisk(non-infectioustohealthyadults)

Moderateindividualrisk

(Notgenerallysevere,

Highindividualrisk

(Treatmentmayor

Severeindividualrisk

(Treatmentoftennotavailable)

treatmentusuallyavailable)maynotbeavailable)

Low risktocommunity

Low risktocommunityLow risktocommunityHighrisktocommunity

Examples

E.Colilabstrains

(e.g.,DH5α,K12)

Mice Rats Rabbits

Humancells,fluids, tissues

NHPcells, fluids,tissues Lentiviralvectors RhesusMacaques Animalsinfectedwith someBSL2agents

M.tuberculosis WestNilevirus Francisellatularensis Yellowfevervirus Monkeypox virus

Animalsinfectedwith BSL3agents

Ebolavirus Lassavirus Marburgvirus

Animalsinfected withBSL4agents


What you need to know abo u t w o rki n g in a

A/BSL-1

A/BSL-2

A/BSL-3

A/BSL-4

Labcoats,gowns, etc.

Eye protection

Latex ornitrilegloves

  • Labcoats,gowns, etc.

  • Eye protection

  • Latex ornitrilegloves

  • Changewhen contaminated

  • Double glovewhen necessary

  • Removeglovesand washhandsafter working

  • Donotre-usegloves

Allmanipulations performedinsidea BSC

Fullprotective clothingthatmust not leavethelab

Eye protection

Latex ornitrilegloves

•BSL-3workpractices Appropriate respiratoryprotection

AllPPE indicatedup toand including BSL-3

Positive-pressure suit

Special facility engineering features


What you need to know abo u t w o rki n g in a

  • Laboratoryhasdoorstolimittraffic

  • Handwashingsinkis available

  • Worksurfacesareeasytodisinfect


What you need to know abo u t w o rki n g in a

  • Limitaccesswhenworking

  • Noeating,drinking,applying cosmeticsorhandlingcontact lenses

  • Nomouthpipetting

  • Glovesmustbewornandlab coatsandprotectiveeyewearare recommended

  • Minimizesplashesandcreation

  • ofaerosols

  • Disinfectwasteandwork surfaces

  • Biologicalwasteshouldbe placedin abiohazarddisposal box,labeled,andplacedoutside forpickupwhen¾full


What you need to know abo u t w o rki n g in a

  • All BSL-1requirements,plus:

  • Autoclaveis available

  • Eyewashis present

  • Signageis posted

  • Biologicalwaste streamis separate


What you need to know abo u t w o rki n g in a

  • AllBSL-1practices,plus:

  • Asupervisormustlimitaccessto thosewhoaretrainedand approved

  • Policyforhandlingsharpsmustbe

  • implemented

  • Laboratoryequipmentmustbe routinelydecontaminated

  • Protectivelabcoatsordisposable

  • gownsmustbe worn

  • Laboratory-specificBiosafety manualmustbeavailable inthelab


What you need to know abo u t w o rki n g in a

  • Hand Washing

  • Alllaboratoriesare requiredtohavea sink availableforhand washing

  • Washhandsfor15 secondsusingwarm waterandmild– preferablyliquid–soap

  • Rinsewithwarm

  • runningwater

  • Drywith disposable papertowel


What you need to know abo u t w o rki n g in a

  • Hand Washing

  • Alcohol-basedhand sanitizersarean alternativetohand washing

  • Sanitizersareeffective againstcommon clinicalmicrobes,but havenotbeentested againstlaboratory pathogens

  • Handwashingis preferred


What you need to know abo u t w o rki n g in a

PPE

Gloves

  • Latexornitrileglovesshouldbe usedforall handling ofbiological materials

  • Doubleglovesmaybeneededin somecircumstancesin orderto avoidexposureorcontamination

  • Theytypeofgloves necessary andthefrequencyofchangingis specifictoyourworkandshould beindicatedin lab-specific training

  • Glovesmustneverbe worn

  • outsideoftheworkarea


What you need to know abo u t w o rki n g in a

PPE

EyeandFaceProtection

  • Wearprotectiveeyewearwhen conductingproceduresthathave thepotentialtocreatesplashesof microorganismsorother hazardousmaterials

  • Peoplewhowearcontactlenses shouldalso weareyeprotection

  • AtBSL2 andabove,eyeandface protectionmustbe usedfor anticipatedsplashesandsprays ofinfectiousmaterialswhenthe microorganismishandledoutside ofa biosafetycabinetorother containmentdevice


What you need to know abo u t w o rki n g in a

  • Sharpsprecautions

  • Sharpsareanyinstrumentthatcanpuncture,cutorscrape

  • UseEXTREME cautionwhenworkingwith sharps

  • Wheneverpossible,alternativestosharps–suchas

  • plasticware–shouldbe used

  • Whensharpsarenecessary,safetysharpsshouldbe selectedwhenevertheyareavailable


What you need to know abo u t w o rki n g in a

WorkPractices

  • ExamplesofSafety Sharps


What you need to know abo u t w o rki n g in a

  • Sharpsprecautions

  • Brokenglasswaremustneverbepicked upbyhand

  • Pickup brokenglassmechanically,using forceps,a brushanddustpan,tongs,etc

  • SharpsDisposal

  • Useddisposableneedlesmustnotbe bent,altered,broken,recapped, removedfromdisposablesyringes,or otherwisemodified

  • Alwaysdisposeofcontaminatedsharps inanapproved,puncture-resistant sharpscontainer

  • Disposeofcontainerwhenit is¾fullby sealingthecontainerandplacingin a biologicalwastebox


What you need to know abo u t w o rki n g in a

  • SharpsDisposal

  • Uncontaminatedordecontaminatedglassmaybe disposedofin adesignated,labeledcardboardbox

  • Boxshouldbesturdyandin goodcondition

  • Takecarenottooverloadthebox–itshouldbekepttoa

  • reasonableweight,approximately25lbs.


What you need to know abo u t w o rki n g in a

  • BiologicalWasteDisposal

  • Tobeusedforallitemscontaminatedwith

  • humanoranimalblood,fluid ortissue

  • Alsostocks,culturesorwastefrom infectiousmaterialsormicroorganisms

  • Allmaterialsthatmaybecontaminated

  • withrecombinantmolecules

  • Donotplacesharpsin thebiological wastebox.

    • Sealedsharpscontainersonlymaybe

  • placedinthebiologicalwastebox

  • Whenboxis¾fullorreaches25lbs. closeandtieliner,securelycloselid,label withPInameandroomnumberandplace inhallwayforpickup


What you need to know abo u t w o rki n g in a

  • Disinfectionand

  • Decontamination

  • Disinfectionis theprocessof reducinga contaminantload

  • Canbeaccomplishedinthe laboratoryusinga 70%solution ofethanol(EtOH)ora 10% solutionofbleach(sodium hypochlorite)

  • Allworkssurfacesandmaterials shouldbe disinfectedbeforeand afteruse

  • SOPsforroutinedecontamination areavailable attheBiological SafetypageoftheEH&Swebsite


What you need to know abo u t w o rki n g in a

  • Disinfectionand Decontamination

  • Decontaminationis theprocessof

  • removingbiohazardousagents

  • Canbeaccomplishedbyphysicalor chemicalmeans

  • Istypically doneusinganautoclave,

  • utilizinghigh temperatureandpressure

  • Aqueoussolutionssuchasblood,urine,or microbialculturesmustbeautoclaved priortodisposal

  • Placeitemsina secondarycontainer madeofstainlesssteelorautoclaveable plastic

  • Largerloadsrequiremoretime andshould bearrangedinawaythatallowsforsteam penetration(i.e.nottoodenselypacked)

  • Donotcapvesselsoraddexcessiveliquid totheload


What you need to know abo u t w o rki n g in a

  • BiologicalSafety Cabinets(BSCs)

  • UsesHigh Efficiency

  • ParticulateAir (HEPA) filters

  • Doesnotprotectagainstvapor orfumes,whichmaydamage HEPAfilters

  • ClassI:

  • Inwardairflowprotectspersonnel

  • Exhauststooutside ClassII:

  • Fourdifferenttypes

  • Protectspersonnel,materialsandenvironmentwith

  • directionalairflowandmultipleHEPAfilters,aspictured

  • ClassIII:

  • BothinletandexhaustairareHEPAfiltered(pictured)


What you need to know abo u t w o rki n g in a

  • LaminarFlowHood

  • Isnota BSCanddoesnot providepersonnel protection

  • Typicallyusedfornucleic acidmanipulationorother proceduresthatarevery sensitivetocontamination, butthatdonotposearisk topersonnel

  • Airflowsouttowardthe user

  • Nottobeusedforwork withinfectiousor potentiallyinfectious materials


What you need to know abo u t w o rki n g in a

  • ChemicalFumeHood

  • Isnota BSCbutdoes providepersonnel protectionfromchemical fumesbyexternal ventilation

  • IsnotHEPAfiltered

  • Nottobeusedforwork withinfectiousor potentiallyinfectious materials

  • Exhaustcontaining infectiousmaterials createsanexposurerisk fortheimmediate environment


What you need to know abo u t w o rki n g in a

  • SafeOperationofBSCs

  • Disinfectcabinetsbeforeand aftereachusewith 70% ethanolor10%bleachsolution

  • Afterdisinfectingthecabinet, loadsuppliesandallowthe cabinettorunfor10-15 minutesbeforebeginningwork

  • Suppliesshouldincludea smallautoclavebag,sharps containerandbeakerwith disinfectantforliquidwaste

  • YourBSC shouldhavea

  • currentcertificationlabel


What you need to know abo u t w o rki n g in a

  • SafeOperationofCentrifuges

  • Checktubesforcracks,leaksor

  • chips

  • Usematchingsetsoftubesand bucketstoensurethatthe centrifugeis properlybalanced

  • Checkthattubesandcupsare sealedandthattherotoris locked andbucketsareproperlyseated

  • Closelidfirmly

  • Whenthecycleis finished,allow therotortocometoa complete stopbeforeopeninglid


What you need to know abo u t w o rki n g in a

  • (LN2)is extremelycold,-320Fº

  • Displacesoxygen

  • Expandsrapidly

  • Protecteyesandskinwith

  • goggles,glovesandshoes

  • DonotallowLINtotouchbare skin;itwillburn

  • Donotseal containers

  • Takecarewhentransferringliquid ortransportingcontainers


Blo o d b orne pathogens

BloodbornePathogens

  • BloodbornePathogens

  • pathogenicmicroorganismspresentin humanbloodandother bodyfluids thatcancausediseaseinhumans,including hepatitisBvirus(HBV),hepatitisC virus(HCV),andhuman immunodeficiencyvirus(HIV).

  • OtherPotentiallyInfectiousMaterial(OPIM)

  • materialsotherthanhumanbloodthatcancontainbloodborne

  • pathogensandmaypotentiallybeinfectious

  • includesHIV-containingcellortissueculturesororgancultures andHIV-orHBV-containingculturemediumorothersolutions

  • UniversalPrecautions

  • bloodandcertainbodyfluids ofallpatientsaretreatedas potentiallyinfectious

  • Allresearchinvolvingtheuseofhumanblood,bodyfluids,and OPIMiscarriedoutusingBSL2 practicesandprocedures, becauseitis unknownif thesematerialscontainbloodborne pathogens


What you need to know abo u t w o rki n g in a

  • Anexposureis contactwith blood orotherinfectiousorpotentially infectiousmaterials

    • Forexample,needlesticksor scrapesandcutswith contaminatedsharps

    • Ifyou’renotsureif you’ve had anexposure,checkfor puncturesin yourglove.Ifthe gloveisbroken,assumean exposurehasoccurred

    • Contactwithbrokenskin throughcutsorrashes

    • Splashestotheeyes,noseor mouth


What you need to know abo u t w o rki n g in a

  • Ifyouhavean exposure:

    • Stopwhatyouaredoing

    • Thoroughlywashtheaffectedareawith soapandwarm waterfor15minutesusingamassagingmotion

    • Foreyesplashes,go tothenearesteyewashstationand

  • rinsewithplainwaterfor15minutes

  • Ifyouneedmedicalattention:

  • Call911andtell theoperatoryourlocation,name,nature oftheinjury

  • Thencall theUniversityPolice at568-8999

  • Afterreceivingtheneededmedicalattention:

    • Notifyyoursupervisor

    • ContacttheDepartmentofHumanResource Managementat568-3916


What you need to know abo u t w o rki n g in a

  • BiologicalSpillResponse

    • Alertothersin thearea,includingyour supervisor

    • PutonappropriatePPE,then

  • Coverthespillwith papertowel(s)

  • Disinfectbypouringa disinfectant aroundtheperimeterofthespilland allowingtostandfor20minutes

  • Cleanbywipingupwithpapertowel(s)

  • Disinfectbysprayingandwiping downwith disinfectantand papertowel

  • Ifyou areunsureofwhattodo oruncomfortableperforming theclean-up,orifthespill islargerthatyoucanrespondto, calltheUniversityPolice at568-8999


What you need to know abo u t w o rki n g in a

Ifyouhaveanyquestionsorconcerns, pleasecontactDepartmentof EnvironmentalHealth & Safety

LSUHSC:

Phone:(504)568-6585

Emergency:568-8999 Email:[email protected]


What you need to know abo u t w o rki n g in a

HOWTOMAINTAINALAB

NOTEBOOK-

RECORDKEEPINGANDHIPAA

FernTsien,PhD Department ofGenetics LSUHSC


Typ e an d format

TypeandFormat

  • Checkwithyourmentorifhe/sherequiresaspecificformatdepending onthetypeof researchyouaredoing

  • Boundbook

  • Looseleafsheetsandfolders

  • Computer/spreadsheet

  • Keepaseparatefileforprintouts,photographs, X-rays,andanythingthatcannotbetapedonto thenotebook.


Content

Content

  • Everypageshouldinclude:

    • Dateof the startof theexperimenton everypage(includingyear)

    • Brieftitleofexperimentortypeofclinic

    • Statementof purpose:extensionof the titlewithsome basicbackground information


Content continued

Content,continued

  • Everypageshouldalso include:

  • Descriptionof experiment(protocol)

  • Listof patients seen(ifthereisprivateinformationsuch as names,etc. keepthis notebooklocked-moreon thislater)

  • Atthe endof the entire day,a summaryof what youaccomplished.Didthe experimentwork?How canyou correctit? Is thereanyinformationthat youneedto lookup?


Tr y t o recor d everythin g you wil l no t b e abl e t o remember

Trytorecordeverything-you

willnotbeabletoremember

everything

  • Where thingsare kept

  • Serumlotnumber

  • Antibodytiter

  • Centrifugespeed,model, temperature

  • Incubationtimesandtemperatures

  • Numberofcells

  • Agaroseor acylamidepercentageofa gel

  • Cellcultureconditions

  • Calculations

  • BufferpH

  • Whoisinchargeofeachlargeequipment(autoclave,etc.)


Maintenance

Maintenance

  • Recordeverythingassoonasyou can.Donot wait

  • until theendof the weekto recorddata.

  • Attachall data, printouts,andX-raysto the appropriateexperiments.Ifthey don’t fit, put away everythingina file,properlylabeled.

  • Maketablesandgraphs.Thiswillhelp you andyour mentoranalyze the data.It canalsobeusedforlab meetingsand your finalpresentation.

  • Makea plan for thefollowingweek.Thinkabout

  • what thedatameansand whatyou needto donext.

  • Askforhelpto interpretthe data beforecontinuing with theexperiment.


What you need to know abo u t w o rki n g in a

NotebookEthics

  • The labnotebookbelongstothelab.Thismeansthat yourmentorwillreadit,so do notputanythingtoo personalor embarrassing.

  • Donot “peek” atanotherlab member’snotebookwithout asking first.Only youandyour mentor shouldhave accessto it.

  • Donot beoffendedifthementor looksatyournotebook.He/sheneeds tolookatit sometimestohelp troubleshootanexperiment.


Noteboo k ethics cont

NotebookEthics,cont.

  • Donot disposeofoldnotebooksyoufindin thelab,datafoundina drawer oron the computer.

  • Ifyoumadeamistakeor ifyouget negative results,recordthis.Donot beembarrassed andomitrecordingthedata.

  • Your dataisyourresponsibility.Recordit accuratelyandresponsibly.


Hi p a a

HIPAA

  • HealthInsurancePortability

  • andAccountabilityAct

  • FederallawpassedbyCongress in1996 toensurepatient

  • confidentiality

  • Regulationsenforcedby theUSDeptofHealthandHumanServices

  • GuidelinesimplementedinApril, 2003

  • Whatpartdo studentsplayin implementingHIPAA?

  • Howdoesthis lawaffectyourroleas a

  • student?


What you need to know abo u t w o rki n g in a

HIPAAregulationswere

designedto:

protectindividuals’rights to privacyand

confidentiality

and

assurethe security ofelectronictransferof

personalinformation


Workin g wit h patien t data

Workingwithpatientdata

  • Onlythepeoplein yourlaborclinicalgroupcan

  • discussthepatientsortheirdata.

  • Takeallreasonablestepstomakesurethat individualswithoutthe‘needtoknow’donotoverhearconversations.

  • DONOTconductdiscussionswithpatient names inelevatorsorcafeterias.

  • Donotletothersseeyour computerscreenwhile youareworkingonpatientdata.Besuretologoutwhendonewithanycomputerfile.


What you need to know abo u t w o rki n g in a

HIPAAappliestousall--in

allsettings

  • Thatmeansdonotdiscuss patientnamesatschool,at home,ontheshuttlebuses,as wellasthehospitalsand clinics.

  • Ifyourecognizesomeoneintheclinic(evenifitisyourbest friend),orseeanameyou recognize intheresearchor clinicaldatabase,keepittoyourself!!!


Wh y hipaa

WhyHIPAA?

  • Geneticadvancements-as moreisknown about our geneticpredispositionto diseases,HIPAA willensure that,forexample,anindividualis notdeniedinsurance because thecompanyknows thatshe mayeventually develop cancer.

  • Marketing- asinformationismoreeasilycaptured concerning,forexample,theprescriptionswepurchase,HIPAAisdesignedto preventmarketingofunsolicited productsor servicesbasedon harvestedmarketingdata.

  • Technology-asinformationis quickly andsometimeslooselymoved aroundnetworks,HIPAA standardswill holdviolatorsaccountableforaccidentalorintentional ‘interception’ofprotected healthinformation(PHI).


Wh y hipaa1

WhyHIPAA?

  • The latetennisstarArthurAshe’spositive HIV status wasdisclosedby ahealthcareworker and publishedby anewspaperwithouthispermission.


Hipa a regulations

HIPAARegulations

  • Give patientsmorecontrolovertheirhealthinformation.

  • Setboundariesontheuseand disclosureofhealth

  • records.

  • Establishappropriatesafeguardsforall peoplewho participateinorareassociatedwiththeprovisionofhealthcaretoensurethattheyhonorpatients’rightstoprivacy.

  • Hold violators accountablethroughcivilandcriminal penalties.

  • Strike abalancewhenpublicresponsibilityrequires disclosureofsomeformsofdata--forexample,toprotectpublichealth.


Conclusions

Conclusions

  • Write downeverything!

  • Maintain goodnotes!

  • Protect patientdata!


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