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MISTIC-fusion proteins as antigens for high quality membrane protein antibodies.

ABSTRACT: Lack of high-quality antibodies against transmembrane proteins is a widely recognized hindrance in biomedical and cell biological research. Here we present a robust pipeline for the generation of polyclonal antibodies employing full-length membrane proteins as immunogens to overcome this "antibody bottleneck". We express transmembrane proteins fused to a MISTIC fragment that enhances expression of eukaryotic membrane proteins in E. coli. Purified membrane proteins are used as immunogen for rabbit injection employing standard immunizing protocols. The raised antibodies against membrane proteins of the endoplasmic reticulum and the nuclear envelope, which we use as test cases, function in a wide range of applications and are superior to ones produced against soluble domains as immunogens.

SUBMITTER: Alves NS

PROVIDER: S-EPMC5288655 | biostudies-literature | 2017 Feb

REPOSITORIES: biostudies-literature

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MISTIC-fusion proteins as antigens for high quality membrane protein antibodies.

Alves Natalia Silva NS Astrinidis Susanne Adina SA Eisenhardt Nathalie N Sieverding Cornelia C Redolfi Josef J Lorenz Michael M Weberruss Marion M Moreno-Andrés Daniel D Antonin Wolfram W

Scientific reports 20170202

Lack of high-quality antibodies against transmembrane proteins is a widely recognized hindrance in biomedical and cell biological research. Here we present a robust pipeline for the generation of polyclonal antibodies employing full-length membrane proteins as immunogens to overcome this "antibody bottleneck". We express transmembrane proteins fused to a MISTIC fragment that enhances expression of eukaryotic membrane proteins in E. coli. Purified membrane proteins are used as immunogen for rabbi ...[more]

PMID: 28148968

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