Abstract
A solid phase conjugation method is described based on the preadsorption of proteins to aluminium hydroxide adjuvant followed by activation of the adsorbed carrier proteins with iodoacetic acid N-hydroxysuccinimidester or other conjugation reagents. Cysteine-containing peptides were coupled to the iodoacetic acid-activated carrier-adjuvant particles through their SH groups. No dialysis is required since the reaction product is isolated at each step of the procedure by a simple centrifugation and can easily be extensively washed between individual manipulations. The method generates peptide-carrier-adjuvant particles with sterically defined presentation of the peptides at the surface of the particles. When used for immunization of mice and rabbits the conjugates elicited high-titered specific anti-peptide sera, which reacted well with the parent protein in ELISA. The strongest reactions were with the denatured form of the parent protein. On immunoblots antisera to the N- and C-terminus of calreticulin recognized the same M, 52,000 protein.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adsorption
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Aluminum Hydroxide
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Amino Acid Sequence
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Animals
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Antibody Formation
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Calcium-Binding Proteins / administration & dosage
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Calcium-Binding Proteins / immunology
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Calcium-Binding Proteins / metabolism
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Calreticulin
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Carrier Proteins / administration & dosage
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Carrier Proteins / immunology*
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Carrier Proteins / metabolism*
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Immunoblotting
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Mice
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Molecular Sequence Data
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Ovalbumin / administration & dosage
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Ovalbumin / immunology
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Ovalbumin / metabolism
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Peptide Fragments / administration & dosage
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Peptide Fragments / immunology*
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Rabbits
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Ribonucleoproteins / administration & dosage
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Ribonucleoproteins / immunology
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Ribonucleoproteins / metabolism
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Tuberculin / administration & dosage
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Tuberculin / immunology
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Tuberculin / metabolism
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Vaccines, Synthetic / administration & dosage
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Vaccines, Synthetic / immunology
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Vaccines, Synthetic / metabolism
Substances
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Calcium-Binding Proteins
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Calreticulin
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Carrier Proteins
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Peptide Fragments
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Ribonucleoproteins
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Tuberculin
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Vaccines, Synthetic
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Aluminum Hydroxide
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Ovalbumin