DOI: 10.5281/zenodo.14500235
title: Quantum Spin Resonance in Engineered Magneto-Sensitive Fluorescent Proteins Enables Multi-Modal Sensing in Living Cells

description: Quantum mechanical phenomena have been identified as fundamentally significant to an increasing number of biological processes. Simultaneously, quantum sensing is emerging as a cutting-edge technology for precision biosensing. However, biological based candidates for quantum-sensors have thus far been limited to in vitrosystems, are prone to light induced degradation, and require sophisticated experimental setups making high-throughput studies prohibitively complex. We recently created a new class of magneto-sensitive fluorescent proteins (MFPs) [1], which we now show overcome these challenges and represent the first biological quantum-based sensor that functions at physiological conditions and in living cells. Through directed evolution, we demonstrate the possibility of engineering these proteins to alter properties of their response to magnetic fields and radio frequencies. These effects are explained in terms of the spin correlated radical pair (SCRP) mechanism, involving the protein backbone and a bound flavin cofactor. Using this engineered system we demonstrate the first observation of a fluorescent protein exhibiting Optically Detected Magnetic Resonance (ODMR) in living bacterial cells at room temperature, at sufficiently high signal-to-noise to be detected in a single cell, paving the way for development of a new class of in vivo biosensors. Magnetic resonance measurements using fluorescent proteins enable unprecedented technologies, for instance 3D spatial localisation of the fluorescence using gradient fields (i.e. Magnetic Resonance Imaging but using an endogenous probe). We further demonstrate the use of multiple variants of MFPs for multiplexing or lock-in amplification of fluorescence signals, opening a new approach to combining or extracting multiple signals from a biological measurement. Taken together, our results represent a new intersection of imaging and perhaps actuation modalities for engineered biological systems, based on and designed around understanding the quantum mechanical properties of MFPs.
description: Three types of experiments are present in the dataset; compared to the associated article they are labelled using different terminology: magnetic field effect (MFE) measurements are labelled MARY (magnetically altered reaction yield) and optically detected magnetic resonance (ODMR) measurements are labelled RYDMR (reaction yield detected magnetic resonance) or RFE (radio field effect). Spectral data is stored in .mat (MATLAB) files which can be opened using the free python library scipy using the scipy.io.loadmat function. 

For RYDMR and MARY data, images are stored in multi-page tiff files (*.ome.tiff) which can be opened using python package tifffile, for instance:

from tifffile import TiffFile
def load_mary(basedir):
    datasets = glob.glob(basedir + "MARY_*.ome.tiff")
    sweeps = max([int(d.split("_")[-1].split(".")[0]) for d in datasets]) + 1
    all_imgs = []
    for i in range(sweeps):
        impath = basedir + f"MARY_{i}.ome.tiff"
        file = TiffFile(impath)
        imgs = [series.asarray() for series in file.series]
        imgs = np.stack(imgs)
        all_imgs.append(imgs)
    return all_imgs

Meta-data is stored in files named *config.json, and further experimental data (such as electromagnet control sequence or RF frequencies at each point) are stored in *data.npz files. In some cases pre-processed data is stored in *pl.npz files.

During the course of writing the manuscript, the chosen names to refer to the protein variants changed from the names used in the data meta data.

For data for Figures 1 and 2, all the MagLOV samples are MagLOV, and the numbering in the sample name refers to biological replicates (i.e. MagLOV 1, MagLOV 2, MagLOV 3 are all MagLOV).

For other figures where variants are compared, the conversion table is:




Manuscript
Data Files


AsLOV2 R3
MagLOV 5


MagLOV
MagLOV 8


MagLOV 2
MagLOV 11


MagLOV 2 R11 f
MagLOV 14




In some cases a dot is used to denote replicate experiments, e.g. MagLOV 14.1 is MagLOV 14, the .1 indicates we did multiple repeats and this was the first.

For assistance with loading the data, please do not hesistate to contact Gabriel Abrahams (gabriel.abrahams@eng.ox.ac.uk).
description: Version update: previous version had 5 second data for Figure 4, uploaded 10 second data as appears in manuscript.