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bioRxiv · 10.1101/2023.11.09.566381

Single protein molecules separation, tracking and counting in ultra-thin silicon channels

Ohayon, S.·Taib, L.·Verma, N. C.·Iarossi, M.·Bhattacharya, I.·Marom, B.·Huttner, D.·Meller, A.
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Abstract

Emerging single-molecule protein sensing techniques are ushering in a transformative era in biomedical research. Nevertheless, challenges persist in realizing ultra-fast full-size protein sensing, including loss of molecular integrity due to protein fragmentation, biases introduced by antibodies affinity, identification of proteoforms and low throughputs. Here, we introduce a single-molecule method for parallel protein separation and tracking, yielding multi-dimensional molecular properties used for their identification. Proteins are tagged by dual amino-acid specific labels and are electrophoretically separated by their mass/charge in custom-designed silicon nano-channel. This approach allows us to analyze thousands of individual proteins within a few minutes by tracking their motion during the migration. We demonstrate the power of the method by quantifying a cytokine panel for host-response discrimination between viral and bacterial infections. Moreover, we show that two clinically-relevant splice isoforms of VEGF can be accurately quantified from human serum samples. Being non-destructive and compatible with full-length intact proteins, this method opens up new ways for antibody-free single protein molecule quantification.

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Ohayon, S., Taib, L., Verma, N. C., Iarossi, M., Bhattacharya, I., Marom, B., Huttner, D., Meller, A.. 2023-11-13. Single protein molecules separation, tracking and counting in ultra-thin silicon channels. https://doi.org/10.1101/2023.11.09.566381

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