title: Step-by-step protocol to perfuse and dissect the mouse parotid gland and isolation of high-quality RNA from murine and human parotid tissue.
abstract: Macroscopic identification and surgical removal of the mouse parotid gland is demanding because of its anatomic location and size. Moreover, the mouse parotid gland contains high concentrations of RNases, making it difficult to isolate high-quality RNA. So far, appropriate methods for optimal perfusion-fixation and dissection of mouse parotid glands, as well as the isolation of high quality RNA from this tissue, are not available. Here we present a simple, optimized, step-by-step surgical method to perfuse and isolate murine parotid glands. We also compared two common RNA extraction methods (RNeasy Mini Kit versus TRIzol) for their yields of high-quality, intact RNA from human and murine parotid gland tissues that were either snap-frozen or immersed in RNAlater stabilization solution. Mouse parotid tissue that was perfused and immersed in RNAlater and human samples immersed in RNAlater exhibited the best RNA quality, independent of the isolation method.
authors.affiliation: Institute for Anatomy and Cell Biology, Medical Cell Biology, Justus-Liebig-University, Giessen, Germany.
authors.forename: Christoph
authors.initials: C
authors.lastname: Watermann
authors.affiliation: Institute for Anatomy and Cell Biology, Medical Cell Biology, Justus-Liebig-University, Giessen, Germany.
authors.forename: Klaus Peter
authors.initials: KP
authors.lastname: Valerius
authors.affiliation: Department of Otorhinolaryngology, Head and Neck Surgery, Justus-Liebig-University, Giessen, Germany.
authors.forename: Steffen
authors.initials: S
authors.lastname: Wagner
authors.affiliation: Department of Otorhinolaryngology, Head and Neck Surgery, Justus-Liebig-University, Giessen, Germany.
authors.forename: Claus
authors.initials: C
authors.lastname: Wittekindt
authors.affiliation: Department of Otorhinolaryngology, Head and Neck Surgery, Justus-Liebig-University, Giessen, Germany.
authors.forename: Jens Peter
authors.initials: JP
authors.lastname: Klussmann
authors.affiliation: Institute for Anatomy and Cell Biology, Medical Cell Biology, Justus-Liebig-University, Giessen, Germany.
authors.forename: Eveline
authors.initials: E
authors.lastname: Baumgart-Vogt
authors.affiliation: Institute for Anatomy and Cell Biology, Medical Cell Biology, Justus-Liebig-University, Giessen, Germany.
authors.forename: Srikanth
authors.initials: S
authors.lastname: Karnati
mesh_terms: D000818:Animals; D004210:Dissection / Q000379:methods*; D006801:Humans; D051379:Mice; D010306:Parotid Gland / Q000378:metabolism / Q000648:ultrastructure*; D010477:Perfusion / Q000379:methods*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D016707:Tissue Fixation / Q000379:methods
chemical_list: D012313:RNA
keywords: RNA isolation; human; mouse; parotid gland; perfusion-fixation; qRT-PCR; surgical preparation