The detection of the mRNAs of procollagen types I, II and III in human fetal fingers by in situ hybridization using digoxigenin-labelled oligonucleotide probes.
Messenger RNAs (mRNAs) encoding procollagen alpha 1 type I,alpha 1 type II and alpha 1 type III have been localized in paraffin sections of human fetal fingers using digoxigenin-labelled synthetic oligonucleotide probes. The probe-mRNA hybrids were visualized using an anti-digoxin antibody amplified with sandwich techniques. These protocols provided an excellent hybridization signal with minimal background noise. The sensitivity of the protocols was nearly equivalent to that seen when using isotopic cDNA probes. In human fetal fingers, intense hybridization signals for procollagen alpha 1 type I mRNA were detected in the osteoblasts and the fibroblasts of periosteum and perichondrium, the tenocytes of tendons, fibroblasts of ligaments, the synovial membrane and deeper layers of the dermis. In contrast, positive hybridization signals for procollagen alpha 1 type II mRNA were visualized in chondrocytes and the cambial layer of perichondrium. The signals for procollagen alpha 1 type III mRNA were detected in the fibroblasts of the dermis and perichondrium. The probes which have lower melting temperatures (Tm) could not detect the corresponding mRNAs.
Hamada
K
K
Department of Orthopaedic Surgery, Tokai University Oiso Hospital, Kanagawa, Japan.
Okawara
Y
Y
Fryer
J N
JN
Tomonaga
A
A
Fukuda
H
H
D001483:Base Sequence; D004076:Digoxigenin; D005260:Female; D005385:Fingers / Q000196:embryology / Q000502:physiology*; D006801:Humans; D017403:In Situ Hybridization; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016612:Paraffin Embedding; D011247:Pregnancy; D011347:Procollagen / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000378:metabolism*
D015345:Oligonucleotide Probes; D011347:Procollagen; D012333:RNA, Messenger; D004076:Digoxigenin