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An in vitro Microscopy-based Assay for Microtubule-binding and Microtubule-crosslinking by Budding Yeast Microtubule-associated Protein

Bio Protoc.. 2018-12; 
Zhu Y, Tan W, Lee WL.
Products/Services Used Details Operation
THE™ alpha Tubulin Antibody, mAb, mouse (GenScript, catalog number: A01410) (store at −20 °C)

摘要

In this protocol, we describe a simple microscopy-based method to assess the interaction of a microtubule-associated protein (MAP) with microtubules. The interaction between MAP and microtubules is typically assessed by a co-sedimentation assay, which measures the amount of MAP that co-pellets with microtubules by centrifugation, followed by SDS-PAGE analysis of the supernatant and pellet fractions. However, MAPs that form large oligomers tend to pellet on their own during the centrifugation step, making it difficult to assess co-sedimentation. Here we describe a microscopy-based assay that measures microtubule binding by direct visualization using fluorescently-labeled MAP, solving the limitations of the co-se... More

关键词

Imaging chamber; Microscopy-based assay; Microtubule; Microtubule binding; Microtubule crosslinking; Microtubule-associated protein; Yeast