A new approach for selective rat endolymphatic sac epithelium collection to obtain pure specific RNA.
The endolymphatic sac (ES) is an organ that is located in the temporal bone. Its anatomical location makes ES tissue collection without any contamination very difficult, and sometimes accurate molecular analyses of the ES are prevented due to this matter. In the present study, a new selective ES epithelial tissue collection method was attempted using laser capture microdissection to obtain pure ES RNA without any contamination. The validity of this method was demonstrated by RT-PCR with three specific primer pairs against osteocalcin, calponin H1, and NKCC2, which are specific proteins in bone, smooth muscle, and kidney/ES cells, respectively. From the RT-PCR results, the high specificity and sufficient sensitivity of the new method was indicated. It is considered that the new method is optimal for ES collection without contamination and it will be able to contribute to future analyses of the ES.
Akiyama
Kosuke
K
Department of Otolaryngology, Faculty of Medicine, Kagawa University, Kita-gun Miki-cho Ikenobe 1750-1, Kita-gun, Kagawa-ken 761-0793, Japan. kakaka@med.kagawa-u.ac.jp
Miyashita
Takenori
T
Matsubara
Ai
A
Mori
Terushige
T
Inamoto
Ryuhei
R
Nishiyama
Akira
A
Mori
Nozomu
N
D000818:Animals; D002135:Calcium-Binding Proteins / Q000235:genetics; D004712:Endolymphatic Sac / Q000737:chemistry* / Q000166:cytology*; D004848:Epithelium / Q000737:chemistry; D007834:Lasers; D042282:Microdissection / Q000379:methods*; D008840:Microfilament Proteins / Q000235:genetics; D015675:Osteocalcin / Q000235:genetics; D012333:RNA, Messenger / Q000737:chemistry / Q000302:isolation & purification*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D028021:Sodium-Potassium-Chloride Symporters / Q000235:genetics; D064466:Solute Carrier Family 12, Member 1; D000096985:Calponins
D002135:Calcium-Binding Proteins; D008840:Microfilament Proteins; D015675:Osteocalcin; D012333:RNA, Messenger; D028021:Sodium-Potassium-Chloride Symporters; D064466:Solute Carrier Family 12, Member 1; D000096985:Calponins; C578716:Slc12a1 protein, rat