Premium
Biospecific irreversible fishing coupled with atomic force microscopy for detection of extremely low‐abundant proteins
Author(s) -
Archakov Alexander,
Ivanov Yurii,
Lisitsa Andrey,
Zgoda Victor
Publication year - 2009
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200800598
Subject(s) - atomic force microscopy , chemistry , microscopy , biophysics , nanotechnology , computational biology , biology , materials science , physics , optics
Abstract In the absence of an analog of PCR for proteins, the concentration detection limit (DL) becomes a real challenge. The problem may be solved by means of a combination of biospecific irreversible fishing with atomic force microscopy (AFM). AFM offers the ability to register individual molecules and their complexes, while biospecific fishing takes advantage of an affine interaction between analyte molecules spread over a large volume of biomaterial and ligand molecules immobilized on the chip surface. Fishing may be conducted in Kd‐dependent reversible mode and in Kd‐independent irreversible mode. In this study, the DLs of two previously applied proteomic approaches were determined and compared to the DL of a newly developed analytical method. The first approach, based on MS analysis of biomaterial after 2‐DE or LC separation of proteins, attained a DL at the level of 10 −8 –10 −10 M. The second approach, based on the optical biosensor analysis of molecular interactions in the format of proteomic microarrays, had a DL of 10 −9 –10 −10 M. Our proposed method which combines biospecific fishing with AFM allowed us to attain DL values of 10 −11 M under reversible binding conditions and 10 −16 M under irreversible binding conditions.