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.
Davidson
Heather
H
Medical Sciences (Medical Genetics), University of Edinburgh, Molecular Medicine Centre, Western General Hospital, Edinburgh EH4 2XU, United Kingdom. H.Davidson@ed.ac.uk
McLachlan
Gerry
G
Wilson
Abigail
A
Boyd
A Christopher
AC
Doherty
Ann
A
MacGregor
Gordon
G
Davies
Lee
L
Painter
Hazel A
HA
Coles
Rebecca
R
Hyde
Stephen C
SC
Gill
Deborah R
DR
Amaral
Margarida D
MD
Collie
David D S
DD
Porteous
David J
DJ
Penque
Deborah
D
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