Construction of dystrophin fusion proteins to raise targeted antibodies to different epitopes.
For the study of the structure and function relationship of dystrophin, defective in DMD, and for diagnostic purposes it is important to dispose of antibodies against different parts of the protein. We have made five different constructs for the expression of fusion proteins containing parts of the four domains of dystrophin. Two different recombinant expression vectors, pATH2 and pEX1, were used. Rabbits were immunized with the fusion products and several polyclonal antibodies were raised. At a later stage, monoclonal antibodies were also raised to some of the fusion proteins. One polyclonal antibody, named P20 AB, is directed against the region covering amino acid sequence 1749-2248 or the nucleotide sequence 5456-6953 of the mRNA, which corresponds to the major deletion-prone region of the DMD gene. We show the particular value, sensitivity and specificity of the P20 AB in dystrophin analysis.
Ginjaar
H B
HB
Department of Human Genetics, Sylvius Laboratory, Leiden, The Netherlands.
van Paassen
H B
HB
den Dunnen
J T
JT
Man
N T
NT
Morris
G E
GE
Moorman
A F
AF
van Ommen
G J
GJ
D000917:Antibody Formation; D000918:Antibody Specificity; D016189:Dystrophin / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D000939:Epitopes / Q000276:immunology*; D006801:Humans; D007150:Immunohistochemistry; D009132:Muscles / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000276:immunology; D015183:Restriction Mapping
D016189:Dystrophin; D000939:Epitopes; D011993:Recombinant Fusion Proteins