Highly sensitive protein detection by combination of atomic force microscopy fishing with charge generation and mass spectrometry analysis

An approach combining atomic force microscopy (AFM) fishing and mass spectrometry (MS) analysis to detect proteins at ultra‐low concentrations is proposed. Fishing out protein molecules onto a highly oriented pyrolytic graphite surface coated with polytetrafluoroethylene film was carried out with an...

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Bibliographic Details
Published in:The FEBS journal 2014-10, Vol.281 (20), p.4705-4717
Main Authors: Ivanov, Yuri D, Pleshakova, Tatyana, Malsagova, Krystina, Kozlov, Andrey, Kaysheva, Anna, Kopylov, Arthur, Izotov, Alexander, Andreeva, Elena, Kanashenko, Sergey, Usanov, Sergey, Archakov, Alexander
Format: Article
Language:English
Subjects:
AFM
atomic force microscopy
Biophysics
Cytochromes b5 - analysis
electric field
graphene
human serum albumin
Humans
Mass spectrometry
Microscopy
Microscopy, Atomic Force - instrumentation
Microscopy, Atomic Force - methods
Molecular biology
Peptide Fragments - analysis
polytetrafluoroethylene
protein fishing
Proteins
Serum Albumin - analysis
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - methods
ultra‐low protein concentration
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Summary:An approach combining atomic force microscopy (AFM) fishing and mass spectrometry (MS) analysis to detect proteins at ultra‐low concentrations is proposed. Fishing out protein molecules onto a highly oriented pyrolytic graphite surface coated with polytetrafluoroethylene film was carried out with and without application of an external electric field. After that they were visualized by AFM and identified by MS. It was found that injection of solution leads to charge generation in the solution, and an electric potential within the measuring cell is induced. It was demonstrated that without an external electric field in the rapid injection input of diluted protein solution the fishing is efficient, as opposed to slow fluid input. The high sensitivity of this method was demonstrated by detection of human serum albumin and human cytochrome b₅ in 10⁻¹⁷–10⁻¹⁸ m water solutions. It was shown that an external negative voltage applied to highly oriented pyrolytic graphite hinders the protein fishing. The efficiency of fishing with an external positive voltage was similar to that obtained without applying any voltage.
ISSN:1742-464X
1742-4658
DOI:10.1111/febs.13011