Recombinant Antibody Fragments with Specificity for Toxic Oligomeric Forms of Disease Related Proteins

Michael Sierks (Inventor)

Research output: Patent

Abstract

Altered protein processing, including misfolding and aggregation, is a pathological aspect of numerous neurodegenerative diseases including Alzheimers, Parkinsons and related synucleionopathies, and Huntingtons Disease (HD). Aggregation of the protein huntingtin (htt) into fibrils has been associated with HD. The exon 1 portion of htt has been shown to form aggregates in vitro and in vivo. Htt can form a number of different morphologies including branched fibrils and buds. The roles of the various morphologies in the progression of HD are not known. Here we demonstrate a novel technique that can be used to isolate morphology specific antibodies expressed on the surface of bacteriophage from a nave phage display antibody library by panning against oligomeric forms of the protein alpha-synuclein. Alpha-synuclein was chosen as a model protein to develop this protocol since it aggregates more slowly than htt. Our results demonstrate that phage displayed antibodies against the target protein can be isolated after only two rounds of selection. Moreover, using this AFM based technique we can isolate antibodies to the oligomeric morphology of alpha-synuclein. After only 2 rounds of selection, 20 out of 48 clones showed positive binding as indicated by ELISA analysis, and 11of the 20 positive clones contained full length scFv as indicated by PCR. The specificity of the selected scFv-phages against oligomeric alpha-synuclein was verified by Atomic Force Microscopy.
Original languageEnglish (US)
StatePublished - May 10 2006

Fingerprint

alpha-Synuclein
Immunoglobulin Fragments
Poisons
Bacteriophages
Huntington Disease
Antibodies
Proteins
Clone Cells
Atomic Force Microscopy
Neurodegenerative Diseases
Exons
Enzyme-Linked Immunosorbent Assay
Polymerase Chain Reaction

Cite this

@misc{ec86147cedbf4eed9c6ef0e5583efdec,
title = "Recombinant Antibody Fragments with Specificity for Toxic Oligomeric Forms of Disease Related Proteins",
abstract = "Altered protein processing, including misfolding and aggregation, is a pathological aspect of numerous neurodegenerative diseases including Alzheimers, Parkinsons and related synucleionopathies, and Huntingtons Disease (HD). Aggregation of the protein huntingtin (htt) into fibrils has been associated with HD. The exon 1 portion of htt has been shown to form aggregates in vitro and in vivo. Htt can form a number of different morphologies including branched fibrils and buds. The roles of the various morphologies in the progression of HD are not known. Here we demonstrate a novel technique that can be used to isolate morphology specific antibodies expressed on the surface of bacteriophage from a nave phage display antibody library by panning against oligomeric forms of the protein alpha-synuclein. Alpha-synuclein was chosen as a model protein to develop this protocol since it aggregates more slowly than htt. Our results demonstrate that phage displayed antibodies against the target protein can be isolated after only two rounds of selection. Moreover, using this AFM based technique we can isolate antibodies to the oligomeric morphology of alpha-synuclein. After only 2 rounds of selection, 20 out of 48 clones showed positive binding as indicated by ELISA analysis, and 11of the 20 positive clones contained full length scFv as indicated by PCR. The specificity of the selected scFv-phages against oligomeric alpha-synuclein was verified by Atomic Force Microscopy.",
author = "Michael Sierks",
year = "2006",
month = "5",
day = "10",
language = "English (US)",
type = "Patent",

}

TY - PAT

T1 - Recombinant Antibody Fragments with Specificity for Toxic Oligomeric Forms of Disease Related Proteins

AU - Sierks, Michael

PY - 2006/5/10

Y1 - 2006/5/10

N2 - Altered protein processing, including misfolding and aggregation, is a pathological aspect of numerous neurodegenerative diseases including Alzheimers, Parkinsons and related synucleionopathies, and Huntingtons Disease (HD). Aggregation of the protein huntingtin (htt) into fibrils has been associated with HD. The exon 1 portion of htt has been shown to form aggregates in vitro and in vivo. Htt can form a number of different morphologies including branched fibrils and buds. The roles of the various morphologies in the progression of HD are not known. Here we demonstrate a novel technique that can be used to isolate morphology specific antibodies expressed on the surface of bacteriophage from a nave phage display antibody library by panning against oligomeric forms of the protein alpha-synuclein. Alpha-synuclein was chosen as a model protein to develop this protocol since it aggregates more slowly than htt. Our results demonstrate that phage displayed antibodies against the target protein can be isolated after only two rounds of selection. Moreover, using this AFM based technique we can isolate antibodies to the oligomeric morphology of alpha-synuclein. After only 2 rounds of selection, 20 out of 48 clones showed positive binding as indicated by ELISA analysis, and 11of the 20 positive clones contained full length scFv as indicated by PCR. The specificity of the selected scFv-phages against oligomeric alpha-synuclein was verified by Atomic Force Microscopy.

AB - Altered protein processing, including misfolding and aggregation, is a pathological aspect of numerous neurodegenerative diseases including Alzheimers, Parkinsons and related synucleionopathies, and Huntingtons Disease (HD). Aggregation of the protein huntingtin (htt) into fibrils has been associated with HD. The exon 1 portion of htt has been shown to form aggregates in vitro and in vivo. Htt can form a number of different morphologies including branched fibrils and buds. The roles of the various morphologies in the progression of HD are not known. Here we demonstrate a novel technique that can be used to isolate morphology specific antibodies expressed on the surface of bacteriophage from a nave phage display antibody library by panning against oligomeric forms of the protein alpha-synuclein. Alpha-synuclein was chosen as a model protein to develop this protocol since it aggregates more slowly than htt. Our results demonstrate that phage displayed antibodies against the target protein can be isolated after only two rounds of selection. Moreover, using this AFM based technique we can isolate antibodies to the oligomeric morphology of alpha-synuclein. After only 2 rounds of selection, 20 out of 48 clones showed positive binding as indicated by ELISA analysis, and 11of the 20 positive clones contained full length scFv as indicated by PCR. The specificity of the selected scFv-phages against oligomeric alpha-synuclein was verified by Atomic Force Microscopy.

M3 - Patent

ER -