Human-specific cystic fibrosis transmembrane conductance regulator antibodies detect in vivo gene transfer to ovine airways.
A panel of 11 human cystic fibrosis transmembrane conductance regulator (hCFTR) antibodies were tested in ovine nasal, tracheal, and bronchial epithelial brushings. Two of these, G449 (polyclonal) and MATG1104 (monoclonal), recognized hCFTR but did not cross react with endogenous sheep CFTR. This specificity allows immunologic detection of hCFTR expressed in gene transfer studies in sheep against the background of endogenous ovine CFTR, thus enhancing the value of the sheep as a model animal in which to study CFTR gene transfer. Studies on mixed populations of human and sheep nasal epithelial cells showed that detection of hCFTR by these two antibodies was possible even at the lowest proportion of human cells (1:100). The hCFTR gene was delivered in vivo by local instillation using polyethylenimine-mediated gene transfer to the ventral surface of the ovine trachea and hCFTR mRNA and protein levels scored in a blinded fashion. Despite abundant hCFTR mRNA expression, the number of cells expressing hCFTR protein detectable by G449 was low (approximately 0.006-0.05%). Immunohistochemistry for hCFTR in animals treated by whole-lung aerosol demonstrated positive cells in sections of tracheal epithelium and in distal conducting airways. The strategic use of hCFTR-specific antibodies supports the utility of the normal sheep as a model for hCFTR gene transfer studies.
Medical Sciences (Medical Genetics), University of Edinburgh, Molecular Medicine Centre, Western General Hospital, Edinburgh EH4 2XU, United Kingdom. H.Davidson@ed.ac.uk
Heather
H
Davidson
Gerry
G
McLachlan
Abigail
A
Wilson
A Christopher
AC
Boyd
Ann
A
Doherty
Gordon
G
MacGregor
Lee
L
Davies
Hazel A
HA
Painter
Rebecca
R
Coles
Stephen C
SC
Hyde
Deborah R
DR
Gill
Margarida D
MD
Amaral
David D S
DD
Collie
David J
DJ
Porteous
Deborah
D
Penque
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D000918:Antibody Specificity; D002452:Cell Count; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry / Q000276:immunology*; D004847:Epithelial Cells / Q000166:cytology; D005260:Female; D015870:Gene Expression; D018014:Gene Transfer Techniques; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012137:Respiratory System / Q000378:metabolism*; D016415:Sequence Alignment; D012756:Sheep / Q000378:metabolism*; D013045:Species Specificity
D000906:Antibodies; D012333:RNA, Messenger; D019005:Cystic Fibrosis Transmembrane Conductance Regulator