About

Ralph Jimenez is a Professor and Adjunct Institute Fellow at JILA, University of Colorado Boulder, in the Department of Chemistry. He earned his Ph.D. from the University of Chicago in 1996 and completed postdoctoral work at the University of California, San Diego, from 1997 to 1998. His research focuses on the interface of quantum optics with physical chemistry, particularly manipulating light at the single-photon or few-photon level to enhance spectroscopy through quantum-engineered light, utilizing properties such as entanglement, superposition, and coherence to increase sensitivity and information content. Jimenez's work also involves the photophysics of fluorescent proteins, aiming to develop molecular probes with improved brightness, photostability, and other properties for cellular imaging and single-molecule measurements. His contributions include establishing methods to increase fluorescent protein brightness and applying machine-learning approaches to improve photostability, addressing longstanding challenges in the field of genetically encoded biomarkers.

Research topics

• Physics
• Chemistry
• Optics
• Quantum mechanics
• Atomic physics
• Biology
• Biophysics

Selected publications

• Enabling robust, stable, and accurate nonlinear optical measurements from squeezed light generated in hot rubidium vapor

  2026-03-04

  article
  PublisherDOI

• Supplementary document for Long-term stabilization of intensity-difference squeezing from four-wave mixing in rubidium vapor - 7891812.pdf

  Figshare · 2026-05-15

  articleOpen access

  Supplemental document

  PublisherDOI

• Dark-State-Mediated Photobleaching in mCherry-Based Red Fluorescent Proteins

  The Journal of Physical Chemistry Letters · 2026-03-16

  articleOpen accessSenior author

  Developing bright and photostable red fluorescent proteins (RFPs) is one of the "holy grails" of the protein engineering community. Despite several attempts, such fluorescent proteins (FPs) have remained elusive. One bottleneck to engineering next-generation RFPs is our lack of understanding of nonfluorescent or dark-state properties in such constructs. Here, we develop a theoretical and experimental framework that describes how photobleaching decays in FPs relate to dark-state conversion and ground-state recovery. Our systematic photophysical investigation of mCherry and mCherry-d, an RFP with enhanced dark-state behavior, showed the presence of photodestructive dark states in such FPs. Molecular dynamics simulations reveal enhanced fluctuation around the imidazolinone end of the chromophore in mCherry-d, potentially facilitating conversion to nonfluorescent states. Collectively, this work quantifies dark-state kinetics and provides insights into engineering dark states in RFPs to develop bright, yet photostable, molecular probes.

  PublisherDOI

• Two-photon-excited fluorescence spectroscopy of Rb atoms in a magneto-optical trap

  ArXiv.org · 2026-02-06

  articleOpen accessSenior author

  We report the results of two-photon-excited fluorescence (TPEF) measurements of the $5\mathrm{S}_{1/2} \rightarrow 5\mathrm{D}_{1/2}$ transition of $^{85}$Rb and $^{87}$Rb cooled in a magneto-optical trap (MOT). We observe TPEF at excitation powers as low as 1 $μ$W or fluxes as low as $4.30 \pm 0.22 \times 10^{18}\ \text{photons}\,\text{cm}^{-2}\,\text{s}^{-1}$ ($^{85}$Rb) and $5.17 \pm 0.26 \times 10^{18}\ \text{photons}\,\text{cm}^{-2}\,\text{s}^{-1}$ ($^{87}$Rb). Our results show that Rb, with additional benefits due to its ability to be optically cooled to the point where Doppler-broadening is negligible, is a promising platform for observing sensitive two-photon spectral signatures at low photon fluxes.

  PublisherOA PDF

• Supplementary document for Long-term stabilization of intensity-difference squeezing from four-wave mixing in rubidium vapor - 7891812.pdf

  Figshare · 2026-05-15

  articleOpen access

  Supplemental document

  PublisherDOI

• Long-term stabilization of intensity-difference squeezing from four-wave mixing in rubidium vapor

  Figshare · 2026-05-15

  otherOpen access

  Generation of quantum states of light through off-resonance four-wave mixing in rubidium vapor is a straightforward and well-established technique. However, achieving a sufficiently high and consistent level of intensity difference squeezing (IDS) and intrinsic quantum correlations between photon pairs over an extended timescale, necessary for quantum-light-based nonlinear optical spectroscopy and microscopy, remains challenging and largely unexplored. Here, we report a simple stabilization method combining an active periodic laser frequency retuning with an automatic control algorithm based on quantitatively assessing the factors affecting the IDS level and squeezed light intensity. Validation of our method was performed by acquiring data over a 5-hour period and the results demonstrate a remarkably stable squeezing level of -7.8 dB in combination with a > 4× reduction of the standard deviation of IDS level from 0.46 dB to 0.10 dB. The achieved stabilization further enables us to quantitatively assess the IDS reduction due to the scattering in a polystyrene bead suspension as a function of sample transmission. Our approach should enable a variety of applications requiring an extended squeezing stability over multiple hours, especially for those following biological processes and chemical reactions in real time.

  PublisherDOI

• Two-photon-excited fluorescence spectroscopy of Rb atoms in a magneto-optical trap

  Open MIND · 2026-02-06

  preprintSenior author

  We report the results of two-photon-excited fluorescence (TPEF) measurements of the $5\mathrm{S}_{1/2} \rightarrow 5\mathrm{D}_{1/2}$ transition of $^{85}$Rb and $^{87}$Rb cooled in a magneto-optical trap (MOT). We observe TPEF at excitation powers as low as 1 $μ$W or fluxes as low as $4.30 \pm 0.22 \times 10^{18}\ \text{photons}\,\text{cm}^{-2}\,\text{s}^{-1}$ ($^{85}$Rb) and $5.17 \pm 0.26 \times 10^{18}\ \text{photons}\,\text{cm}^{-2}\,\text{s}^{-1}$ ($^{87}$Rb). Our results show that Rb, with additional benefits due to its ability to be optically cooled to the point where Doppler-broadening is negligible, is a promising platform for observing sensitive two-photon spectral signatures at low photon fluxes.

  DOI

• Long-term stabilization of intensity-difference squeezing from four-wave mixing in rubidium vapor

  Figshare · 2026-05-15

  otherOpen access

  Generation of quantum states of light through off-resonance four-wave mixing in rubidium vapor is a straightforward and well-established technique. However, achieving a sufficiently high and consistent level of intensity difference squeezing (IDS) and intrinsic quantum correlations between photon pairs over an extended timescale, necessary for quantum-light-based nonlinear optical spectroscopy and microscopy, remains challenging and largely unexplored. Here, we report a simple stabilization method combining an active periodic laser frequency retuning with an automatic control algorithm based on quantitatively assessing the factors affecting the IDS level and squeezed light intensity. Validation of our method was performed by acquiring data over a 5-hour period and the results demonstrate a remarkably stable squeezing level of -7.8 dB in combination with a > 4× reduction of the standard deviation of IDS level from 0.46 dB to 0.10 dB. The achieved stabilization further enables us to quantitatively assess the IDS reduction due to the scattering in a polystyrene bead suspension as a function of sample transmission. Our approach should enable a variety of applications requiring an extended squeezing stability over multiple hours, especially for those following biological processes and chemical reactions in real time.

  PublisherDOI

• Dark state-mediated photobleaching in mCherry-based red fluorescent proteins

  bioRxiv (Cold Spring Harbor Laboratory) · 2026-01-23

  articleOpen accessSenior author

  ABSTRACT Developing bright and photostable red fluorescent proteins (RFPs) is one of the ‘holy grails’ of the protein engineering community. Despite several attempts, finding such fluorescent proteins (FPs) has remained elusive. One bottleneck to engineering next generation RFPs is our lack of understanding of non-fluorescent or dark state properties in such constructs. Here, we develop a theoretical and experimental framework that describes how photobleaching decays in FPs relates to dark state conversion and ground state recovery. Our systematic photophysical investigation of mCherry and mCherry-d, an RFP with enhanced dark state behavior, showed the presence of photodestructive dark states in such FPs. Molecular dynamics simulations reveal enhanced fluctuation around the imidazolinone-end of the chromophore in mCherry-d, potentially facilitating conversion to non-fluorescent states. Collectively, this work quantifies dark state kinetics and gives insights into engineering dark states in RFPs to develop bright yet photostable molecular probes.

  PublisherOA PDFDOI

• Long-term stabilization of intensity-difference squeezing from four-wave mixing in rubidium vapor

  Optics Express · 2026-05-04

  articleOpen access

  Generation of quantum states of light through off-resonance four-wave mixing in rubidium vapor is a straightforward and well-established technique. However, achieving a sufficiently high and consistent level of intensity difference squeezing (IDS) and intrinsic quantum correlations between photon pairs over an extended timescale, necessary for quantum-light-based nonlinear optical spectroscopy and microscopy, remains challenging and largely unexplored. Here, we report a simple stabilization method combining an active periodic laser frequency retuning with an automatic control algorithm based on quantitatively assessing the factors affecting the IDS level and squeezed light intensity. Validation of our method was performed by acquiring data over a 5-hour period, and the results demonstrate a remarkably stable squeezing level of -7.8 dB in combination with a > 4× reduction of the standard deviation of IDS level from 0.46 dB to 0.10 dB. The achieved stabilization further enables us to quantitatively assess the IDS reduction due to the scattering in a polystyrene bead suspension as a function of sample transmission. Our approach should enable a variety of applications requiring an extended squeezing stability over multiple hours, especially for those following biological processes and chemical reactions in real time.

  PublisherDOI

Recent grants

• Collaborative Research: Development of a Novel Multiphoton Microscope for Measuring Biomolecular Dynamics Over 15 Orders of Magnitude in Time

  NSF · $345k · 2005–2009

• NIH Grant R01GM083849

  NIH · $1.5M · 2013

• Collaborative Research: Development of a High Speed Cell Mechanical Property Testing Cytometer

  NSF · $282k · 2009–2013

Frequent coauthors

• Srijit Mukherjee

  Stanford University

  67 shared

• Alexander Mikhaylov

  57 shared

• Amy E. Palmer

  52 shared

• Kristen M. Parzuchowski

  50 shared

• Emily A. Gibson

  University of Colorado Anschutz Medical Campus

  49 shared

• Michael D. Mazurek

  46 shared

• Ryan N. Wilson

  Joint Institute for Laboratory Astrophysics

  43 shared

• Premashis Manna

  Massachusetts Institute of Technology

  40 shared

Education

• Ph.D.

  University of Chicago

  1996

• Other

  University of California, San Diego

  1998

• Other

  The Scripps Research Institute

  2003

Awards & honors

• Arthur S. Flemming Award 2017
• U.S. Department of Commerce Gold Medal 2017
• U.S. Department of Commerce Bronze Medal 2016
• U.S. Department of Commerce Bronze Medal 2023