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Volume 31
Issue 18
10.3390/molecules31183307
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This is an early access version, the complete PDF, HTML, and XML versions will be available soon.
Article

A Novel Protein Recovery Method via Gel Separation and Silica Columns Capable of Rapid Antigen Purification and Deep Proteomic Identification

by
Dan Wang
1,†,
Jianhong Wu
2,†,
Xingmei Zheng
1,
Ziquan Fan
2,*,
Zhexuan Li
1,
Cunzhi Peng
1,
Bingqiang Xu
3 and
Zheng Tong
1,*
1
Key Laboratory of Biology and Genetic Resources of Tropical Crops, Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences & Hainan Institute for Tropical Agricultural Resources, Haikou 571101, China
2
Thermo Fisher Scientific (China) Co., Ltd., Shanghai, China., 2517 Jinke Road, Pudong New Area, Shanghai 201206, China
3
State Key Laboratory of Tropical Crop Breeding, Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences, Sanya 572024, China
*
Authors to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Molecules 2026, 31(18), 3307; https://doi.org/10.3390/molecules31183307 (registering DOI)
Submission received: 30 July 2026 / Revised: 9 September 2026 / Accepted: 16 September 2026 / Published: 17 September 2026
(This article belongs to the Special Issue Advances in the Mass Spectrometry of Chemical and Biological Samples, 2nd Edition)
Versions Notes

Abstract

Gel electrophoresis is widely used for protein and nucleic acid analysis. Nucleic acid recovery from agarose gels is convenient and efficient, but protein recovery is not. This study aimed to develop and validate a novel method for recovering proteins from gels. Methods: After protein separation on gels, excised gel slices containing target proteins are processed using the same silica-column workflow as that used for nucleic acid purification, but with buffers specifically adapted for protein recovery. This approach only requires brief centrifugation steps and avoids laborious extraction and specialized equipment. Results: Proteome-wide recovery showed a slight preference of the method for low-molecular-mass and acidic proteins, while individual protein recovery efficiencies ranged from 69.9% to 86.7% and showed no correlation with intrinsic protein properties. The method effectively supported antigen purification. A banana (Musa acuminata AAA group ‘Brazilian’) proteome library constructed from recovered fractions contained over 18,000 protein groups and 27,000 individual proteins, which significantly enhanced protein identification in library-based data-independent acquisition (DIA) compared to directDIA analysis of single samples. Conclusions: This method bridges gel-based protein separation- and column-based purification, making protein purification as simple, fast, and scalable as nucleic acid purification. It offers a practical alternative for proteomic sample preparation, antigen purification, and DIA workflows.
Keywords: gel separation-based protein recovery method; silica column; protein purification; antigen purification; molecular mass-based proteomic pre-fractionation; proteomic identification gel separation-based protein recovery method; silica column; protein purification; antigen purification; molecular mass-based proteomic pre-fractionation; proteomic identification

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MDPI and ACS Style

Wang, D.; Wu, J.; Zheng, X.; Fan, Z.; Li, Z.; Peng, C.; Xu, B.; Tong, Z. A Novel Protein Recovery Method via Gel Separation and Silica Columns Capable of Rapid Antigen Purification and Deep Proteomic Identification. Molecules 2026, 31, 3307. https://doi.org/10.3390/molecules31183307

AMA Style

Wang D, Wu J, Zheng X, Fan Z, Li Z, Peng C, Xu B, Tong Z. A Novel Protein Recovery Method via Gel Separation and Silica Columns Capable of Rapid Antigen Purification and Deep Proteomic Identification. Molecules. 2026; 31(18):3307. https://doi.org/10.3390/molecules31183307

Chicago/Turabian Style

Wang, Dan, Jianhong Wu, Xingmei Zheng, Ziquan Fan, Zhexuan Li, Cunzhi Peng, Bingqiang Xu, and Zheng Tong. 2026. "A Novel Protein Recovery Method via Gel Separation and Silica Columns Capable of Rapid Antigen Purification and Deep Proteomic Identification" Molecules 31, no. 18: 3307. https://doi.org/10.3390/molecules31183307

APA Style

Wang, D., Wu, J., Zheng, X., Fan, Z., Li, Z., Peng, C., Xu, B., & Tong, Z. (2026). A Novel Protein Recovery Method via Gel Separation and Silica Columns Capable of Rapid Antigen Purification and Deep Proteomic Identification. Molecules, 31(18), 3307. https://doi.org/10.3390/molecules31183307

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