Beads-free protein immunoprecipitation for a mass spectrometry-based interactome and posttranslational modifications analysis.
Beads-free protein immunoprecipitation for a mass spectrometry-based interactome and posttranslational modifications analysis.
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用于基于质谱的相互作用组和翻译后修饰分析的无珠蛋白免疫沉淀。
DOI：
10.1186/s12953-015-0079-0
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发表时间：
2015
期刊：
影响因子：
2
通讯作者：
Ostrowski J
中科院分区：
文献类型：
--
作者：
Mikula M;Rubel T;Karczmarski J;Statkiewicz M;Bomsztyk K;Ostrowski J
蛋白质免疫沉淀（IP）与MS结合提供了询问蛋白质复合物及其翻译后修饰（PTM）的方法。在典型的蛋白IP测定中，抗体与蛋白A/G珠缀合，需要大量抗体、试管转移和离心。作为替代方案，我们提出了Matrix-IP，无珠微板为基础的平台与表面固定的抗体。试验使用标准96孔聚丙烯PCR板，该板是用UV-C光实验室制造的，然后在IP反应前包被蛋白A/G。我们展示了Matrix-IP平台在异质核核糖核蛋白K（hnRNP K）相互作用组和PTM的MS分析中的应用。Matrix-IP是一种节省时间、易于使用的高通量方法，适用于低样品量和自动化。本文的在线版本（doi：10.1186/s12953-015-0079-0）包含补充材料，可供授权用户使用。
Protein immunoprecipitation (IP) coupled with MS provides means to interrogate protein complexes and their posttranslational modifications (PTMs). In a typical protein IP assay antibodies are conjugated to protein A/G beads requiring large amounts of antibodies, tube transfers and centrifugations. As an alternative, we present Matrix-IP, beads-free microplate-based platform with surface-immobilized antibodies. Assay utilizes standard 96-well polypropylene PCR plates that are laboratory-fabricated with UV-C light and then protein A/G coated prior to IP reaction. We demonstrate application of Matrix-IP platform in MS analysis of heterogeneous nuclear ribonucleoprotein K (hnRNP K) interactome and PTMs. Matrix-IP is time-saving, easy to use high throughput method adaptable for low sample amounts and automation. The online version of this article (doi:10.1186/s12953-015-0079-0) contains supplementary material, which is available to authorized users.