Electron Spin Resonance Scanning Probe Spectroscopy for Ultra-Sensitive Biochemical Studies
Jason P. Campbell, Jason T. Ryan, Pragya R. Shrestha, Zhanglong Liu, Canute I. Vaz, Jihong Kim, Vasileia Georgiou, Kin P. Cheung
Electron spin resonance (ESR) spectroscopys affinity for detecting paramagnetic free radicals, or spins, has been increasingly employed to examine a large variety of biochemical interactions. Such paramagnetic species are broadly found in nature and can be intrinsic (defects in semiconductors, electron/hole pairs, stable radicals in proteins) or, more often, purposefully introduced into the material of interest (doping/attachment of paramagnetic spin labels to biomolecules of interest). Using ESR to trace the reactionary path of paramagnetic spins or spin-active proxy molecules provides detailed information about the reactions transient species and the labels local environment. For many biochemical systems, like those involving membrane proteins, synthesizing the necessary quantity of spin-labeled biomolecules (typically 50 pmol to 100 pmol) is quite challenging and often limits the possible biochemical reactions available for investigation. Quite simply, ESR is too insensitive. Here, we demonstrate an innovative approach that greatly enhances ESRs sensitivity by developing a near-field X-band ESR spectrometric method that does not require a microwave resonator. Sensitivity improvement is confirmed via measurement of 140 amol of the most common nitroxide spin label in a ≈ 593 fL liquid cell at ambient temperature and pressure. This embodiment eliminates many of the typical ESR sample restrictions and renders the technique feasible for spatially resolved measurements on a wider variety of biochemical samples. Thus our approach broadens the pool of possible biochemical and structural biology studies as well as greatly enhances the analytical power of existing ESR applications.
, Ryan, J.
, Shrestha, P.
, Liu, Z.
, Vaz, C.
, Kim, J.
, Georgiou, V.
and Cheung, K.
Electron Spin Resonance Scanning Probe Spectroscopy for Ultra-Sensitive Biochemical Studies, Analytical Chemistry, [online], https://tsapps.nist.gov/publication/get_pdf.cfm?pub_id=917622
(Accessed September 21, 2023)