FIXATIONWITHFORMALDEHYDE-ACETONE

1.Dulbecco"sPBS,37^oC.

2.Fixationbuffer,i.e.PBS,PHEM,ormodifiedHanks("Cytoskeletonbuffer"),1.33x.

3.Formaldehyde,16%,EMSciences.Storeatroomtemperatureinthefumehood.Markthedatewhenyouopenanewampule.SealtightlywithParafilm.Handlewithcaution.

4.Fresh4%formaldehydesolution.Mix3parts1.33xfixationbufferwith1part16%formaldehyde(e.g.9mland3mlfor1coverslip),inabottledesignatedforfixatives.Captightlyandwarmupinawaterbath.

5.Acetone,chilledto-20^oCinafreezer.Transferinto100mlbeakersdesignatedforfixation,eachcoverslipneedsaseparatebeaker.

6.Forcepsforfixation.

7.Plasticfixationboxes.

1.Transferthewarmfixationsolutionintoaplasticfixationboximmediatelybeforefixation.

2.Drainallthemediumfromthecoverslipdish.Rinseout2timeswithwarmPBS.

3.Quicklyandcarefullyremovethecoverslip.

4.Placeonthesurfaceofthefixationsolution,cellsidefacingdown.Thecoverslipshouldfloatonthesurfaceofthesolution.Rocktheboxintermittently.Fixfor10minatroomtemperature.

5.Removethecoverslipwithapairoffixationforceps.Rinsewithdistilledwater2times.

6.Dipthecoverslipintothecoldacetone.Rockgently.Letsitfor5min.

7.RemovethecoverslipfromacetoneandrinseimmediatelywithPBS.Donotletthesurfacedryout.

8.Proceedwithstaining.

9.Usedacetoneshouldbeemptiedintoadesignatedcontainer.

10.Note:Theacetonestepmaybereplacedbyadding0.1%TritonX-100tosolution4anddofixationandextractioninonestep.

================  蚂蚁淘在线  ================

免责声明：本文仅代表作者个人观点，与本网无关。其创作性以及文中陈述文字和内容未经本站证实，对本文以及其中全部或者部分内容、文字的真实性、完整性、及时性本站不做任何保证或承诺，请读者仅作参考，并请自行核实相关内容

版权声明：未经蚂蚁淘在线授权不得转载、摘编或利用其他方式使用上述作品。已经经本网授权使用作品的，应该授权范围内使用，并注明“来源：蚂蚁淘在线”。违反上述声明者，本网将追究其相关法律责任。