Tandem immuno-purification of affinity-tagged proteins from mouse testis extracts for MS analysis
Naoki Tani, Nobuhiro Tanno, Kei‐ichiro Ishiguro
Abstract
Here, we describe a protocol for extraction and tandem immunoprecipitation of the cytoplasmic and nuclear proteins from mouse testis for mass spectrometry. This protocol has been applied to knockin mice that express a meiotic protein of interest tagged with 3xFLAG-HA in the testis. The protocol is optimized for salt extraction of cytoplasmic and nuclear proteins from mouse frozen testes and thus can be used for a variety of proteins. For complete details on the use and execution of this protocol, please refer to Ishiguro et al. (2020) and Tanno et al. (2022).
Topics & Concepts
ImmunoprecipitationCytoplasmTandem mass spectrometryCell biologyTandemBiologyExtraction (chemistry)Molecular biologyTandem affinity purificationMeiosisProtein purificationChemistryComputational biologyMass spectrometryAntibodyBiochemistryChromatographyGeneticsGeneAffinity chromatographyComposite materialEnzymeMaterials scienceAdvanced Proteomics Techniques and ApplicationsMolecular Biology Techniques and ApplicationsMass Spectrometry Techniques and Applications