Proteomics

Dataset Information

15

Hit-Gel: Streamlining in-gel protein digestion for high-throughput proteomics experiments.


ABSTRACT: Traditional in gel-digestion procedures require extensive sample handling, are prone to contamination and not compatible with high-throughput sample preparation. To address these shortcomings, we have modified the conventional in-gel digestion procedure for high-throughput proteomics studies. The modified method, termed “High Throughput in Gel digestion” (HiT-Gel), is based on a 96-well plate format which results in a drastic reduction in labour intensity and sample handling. Direct comparison revealed that HiT-Gel reduces technical variation and significantly decreases sample contamination over the conventional in-gel digestion method. HiT-Gel also produced superior results when a single protein band was excised from a gel and processed by in-gel digestion. Moreover, we applied Hit-Gel for a mass spectrometry analysis of Arabidopsis thaliana protein complexes separated by native PAGE in 24 fractions and four biological replicates. We show that the high throughput capacity of HiT-Gel facilitates large scale studies with high sample replication or detailed fractionation. Our method can easily be implemented as it does not require specialised laboratory equipment.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Arabidopsis thaliana  

TISSUE(S): Leaf

DISEASE(S): Not Available

SUBMITTER: Michal Gorka  

LAB HEAD: Dr. Alexander Graf

PROVIDER: PXD009510 | Pride | 2018-10-16

REPOSITORIES: Pride

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Publications

Hit-Gel: Streamlining in-gel protein digestion for high-throughput proteomics experiments.

Swart Corné C   Martínez-Jaime Silvia S   Gorka Michal M   Zander Kerstin K   Graf Alexander A  

Scientific reports 20180605 1


In-gel digestion has been used as a standard method for the preparation of protein samples for mass spectrometry analysis for over 25 years. Traditional in gel-digestion procedures require extensive sample handling, are prone to contamination and not compatible with high-throughput sample preparation. To address these shortcomings, we have modified the conventional in-gel digestion procedure for high-throughput proteomics studies. The modified method, termed "High Throughput in Gel digestion" (H  ...[more]

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