Label-free observation of individual solution phase molecules

Needham, L.-M.; Saavedra, C.; Rasch, J. K.; Sole-Barber, D.; Schweitzer, B. S.; Fairhall, A. J.; Vollbrecht, C. H.; Mehlenbacher, B.; Zhang, Z.; Tenbrake, L.; Pfeifer, H.; Chapman, E. R.; Goldsmith, R. H.

doi:10.1101/2023.03.24.534170

bioRxiv · 10.1101/2023.03.24.534170

Label-free observation of individual solution phase molecules

Needham, L.-M.·Saavedra, C.·Rasch, J. K.·Sole-Barber, D.·Schweitzer, B. S.·Fairhall, A. J.·Vollbrecht, C. H.·Mehlenbacher, B.·Zhang, Z.·Tenbrake, L.·Pfeifer, H.·Chapman, E. R.·Goldsmith, R. H.

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Abstract

The vast majority of chemistry and biology occurs in solution, and new label-free analytical techniques that can help resolve solution-phase complexity at the single-molecule level can provide new microscopic perspectives of unprecedented detail. Here, we use the increased light-molecule interactions in high-finesse fiber Fabry-Perot microcavities to detect individual biomolecules as small as 1.2 kDa with signal-to-noise ratios >100, even as the molecules are freely diffusing in solution. Our method delivers 2D intensity and temporal profiles, enabling the distinction of sub-populations in mixed samples. Strikingly, we observe a linear relationship between passage time and molecular radius, unlocking the potential to gather crucial information about diffusion and solution-phase conformation. Furthermore, mixtures of biomolecule isomers of the same molecular weight can also be resolved. Detection is based on a novel molecular velocity filtering and dynamic thermal priming mechanism leveraging both photo-thermal bistability and Pound-Drever-Hall cavity locking. This technology holds broad potential for applications in life and chemical sciences and represents a major advancement in label-free in vitro single-molecule techniques.

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Needham, L.-M., Saavedra, C., Rasch, J. K., Sole-Barber, D., Schweitzer, B. S., Fairhall, A. J., Vollbrecht, C. H., Mehlenbacher, B., Zhang, Z., Tenbrake, L., Pfeifer, H., Chapman, E. R., Goldsmith, R. H.. 2023-03-25. Label-free observation of individual solution phase molecules. https://doi.org/10.1101/2023.03.24.534170

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