Imaging Of HeLa Cells Research Articles

Broadband stimulated Raman hyperspectral imaging of cells and semiconductors with synchronized fiber and solid-state sources

We demonstrate broadband hyperspectral stimulated Raman scattering (SRS) microscopy covering over 2000 cm−1, which is achieved by two tunable Stokes light sources synchronized with a pump light source. Specifically, a fiber optical parametric oscillator and a fiber laser, both equipped with automatic wavelength tuning capabilities, are employed to acquire SRS signals in the ranges of 270–2040 cm−1 and 2800–3100 cm−1, respectively. Using this system, we perform hyperspectral SRS imaging of live HeLa cells, covering the entire fingerprint and C–H stretching regions. Furthermore, the spectral coverage in the lower fingerprint region (<1000 cm−1) enables SRS signal acquisition of wide bandgap semiconductors. We succeed in obtaining the SRS spectra of 4H-silicon carbide (SiC) and gallium nitride and demonstrate SRS imaging of the longitudinal optical phonon-plasmon coupled mode of 4H-SiC. We anticipate that the present method will further expand the applications of SRS in various scientific fields.

A Fluorescent Probe Differentiates Apoptosis from Cysteine-Deprivation Ferroptosis through Imaging of Viscosity and Lipid Droplets.

Since death is an inevitable phenomenon, exploring cell deaths holds importance. During this process, the cellular microenvironment within cells such as pH, polarity, viscosity etc alter. One such microenvironment, viscosity elevates during different cell deaths. However, demarcating cell death processes solely based on viscosity sensing is challenging. Herein, we develop a unique fluorescent probe PS-NAP after careful investigation among three analogues for efficient viscosity imaging in HeLa cells. Cationic PS-NAP, a MMP independent molecular probe, can potentially target mitochondria and map elevated mitochondrial viscosity during apoptosis, starvation and drug induced ferroptosis processes. Notably, during ferroptosis induced by cysteine deprivation, PS-NAP exclusively colocalizes in newly generated lipid droplets (LDs) instead of mitochondria. Thus, the probe has a potential for demarcating cysteine deprivation-induced ferroptosis from other cellular stresses such as apoptosis.

Lens-Free On-Chip Quantitative Phase Microscopy for Large Phase Objects Based on a Biplane Phase Retrieval Method.

Lens-free on-chip microscopy (LFOCM) is a powerful computational imaging technology that combines high-throughput capabilities with cost efficiency. However, in LFOCM, the phase recovered by iterative phase retrieval techniques is generally wrapped into the range of -π to π, necessitating phase unwrapping to recover absolute phase distributions. Moreover, this unwrapping process is prone to errors, particularly in areas with large phase gradients or low spatial sampling, due to the absence of reliable initial guesses. To address these challenges, we propose a novel biplane phase retrieval (BPR) method that integrates phase unwrapping results obtained at different propagation distances to achieve accurate absolute phase reconstruction. The effectiveness of BPR is validated through live-cell imaging of HeLa cells, demonstrating improved quantitative phase imaging (QPI) accuracy when compared to conventional off-axis digital holographic microscopy. Furthermore, time-lapse imaging of COS-7 cells in vitro highlights the method's robustness and capability for long-term quantitative analysis of large cell populations.

Microwave-Assisted Synthesis of N, S Co-Doped Carbon Quantum Dots for Fluorescent Sensing of Fe(III) and Hydroquinone in Water and Cell Imaging.

The detection of heavy metal ions and organic pollutants from water sources remains critical challenges due to their detrimental effects on human health and the environment. Herein, a nitrogen and sulfur co-doped carbon quantum dot (NS-CQDs) fluorescent sensor was developed using a microwave-assisted carbonization method for the detection of Fe3+ ions and hydroquinone (HQ) in aqueous solutions. NS-CQDs exhibit excellent optical properties, enabling sensitive detection of Fe3+ and HQ, with detection limits as low as 3.40 and 0.96 μM. Notably, with the alternating introduction of Fe3+ and HQ, NS-CQDs exhibit significant fluorescence (FL) quenching and recovery properties. Based on this property, a reliable "on-off-on" detection mechanism was established, enabling continuous and reversible detection of Fe3+ and HQ. Furthermore, the low cytotoxicity of NS-CQDs was confirmed through successful imaging of HeLa cells, indicating their potential for real-time intracellular detection of Fe3+ and HQ. This work not only provides a green and rapid synthesis strategy for CQDs but also highlights their versatility as fluorescent probes for environmental monitoring and bioimaging applications.

A near infrared fluorescent probe for hypoxia based on dicyanoisophorone and its application in Hela cells imaging

Hypoxia will accelerate tumors metastasis and deterioration, thereby limiting the effects of chemotherapy or radiotherapy. Thus, developing efficient techniques for detecting hypoxia in tumor cells is extremely important for cancer diagnosis and therapy. In this work, we reported a dicyanoisophorone-based probe (DCI-Azo) that specifically switched on its near infrared emission with hypoxia up-regulated azo-reductase (AzoR). In order to reduce the difficulty of synthesis and simplify the post-processing process, we adopted a one-pot-synthesis method to synthesized NIR fluorophore (DCI-Am) with yield 97 %. Based on the fluorophore, DCI-Azo was designed and synthesized. The sensitivity of DCI-Azo for hypoxia in vitro was evaluated with Na2S2O4 and rat liver microsomes. It exhibited near-infrared emission (λem = 650 nm), large Stokes Shift (>160 nm), high sensitivity (LOD 0.53 μg mL−1 rat liver microsomes), high selectivity, and low cytotoxicity (cell viability > 80 % after incubation for 24 h). Moreover, the probe was successfully used for detecting hypoxia (1% O2) in Hela cells and tumor tissue in mouse model. The fluorescence intensity in Hela cells has increased ∼ 26-fold when the oxygen level is reduced to 1 % from 21 % O2. The fluorescence intensity of the tumor area enhanced ∼ 5 folds compared to the normal area nearby. All these features demonstrated that the probe DCI-Azo was a versatile tool for in vivo assay and imaging for cancer diagnosis studies.

Bio-based fabrication of palladium nanoparticles by using Aegle marmelos leaf extract for biomedical applications: Cytotoxicity, cell imaging studies in HeLa cells and antioxidant activity

This paper presents the outcome of medicinal plant-derived synthesis of nanomaterial. The results in the present study provide a simple, novel, sustainable, and economically noble process for the synthesis of palladium nanoparticles using leaf extract Aegle marmelos (A. marmelos). Fourier transformed infrared spectroscopy (FTIR) was used to verify the surface layer of synthesized palladium (ALE@Pd) nanoparticles (NPs) enriched with several kinds of phytochemicals. Transmission electron microscopy (TEM) revealed that the synthesized NPs possess a ball shape and the average diameter was 4.31 ± 1.89 nm. The synthesized NPs possess stability confirmed by UV–visible (UV–Vis) spectrophotometry. The face-centered cubic (FCC) crystal structure of the fabricated nanoparticles was confirmed by powder X-ray diffraction (PXRD). Fabricated ALE@Pd NPs are subjected to further testing for their cytotoxicity in HeLa cell lines, as well as for a comparative study of antioxidant activity between the A. marmelos leaves and prepared ALE@Pd NPs. The anti-carcinogenic activity and cell imaging in HeLa cells were tested, and the minimal inhibition concentration (MIC or IC50) has been determined and found to be around 46 µg/mL for HeLa cell lines. Fabricated palladium nanoparticles derived from A. marmelos have the potential to be employed as an environmentally friendly and cost-effective anticancer agent. Furthermore, synthesized ALE@Pd NPs possess higher antioxidant activity than A. marmelos leaves extract (ALE) due to the increased phenolic content connected to the surface of produced palladium NPs as a capping agent. The minimal inhibition concentration of synthesized ALE@Pd NPs was found to be ∼5 µg/mL.

Polydimethylsiloxanes - Based Fluorescent Probe for H2S Detection in Living Cells.

The development of fluorescent probes for H2S detection especially in living cells is of great significance due to its fundamental role as signal molecule. A promising scaffold for the development of such probes is polydimethylsiloxanes (PDMS), which is cost-effectiveness, non-toxicity, flexibility, and biocompatibility and easy to post-functionalize. Surprisingly, fluorescent probes for H2S detection based on PDMS have not been investigated. Moreover, 4-nitro - 2,1,3-benzoxadiazole (NBD) derivates provides high fluorescence quantum yield, a large molar absorption coefficient, and sensitivity to environmental changes. Through reasonable design and adjustment of substituents on the NBD group, precise control of its fluorescence properties can be achieved. Herein, a novel H2S fluorescent probe, P-NBD, was designed and synthesized by a one-step aromatic ring nucleophilic substitution of Cl-NBD with a PDMS derivative. Due to the occurrence of the cleavage reaction strategy and the intramolecular charge transfer process, P-NBD can detect H2S via a colorimetric method. The limit of detection is down to 93 nM. P-NBD demonstrated considerable detection capability comparable to other reported types of H2S probes. Moreover, the probe can also be utilized for H2S imaging in HeLa cells. This work provides new insights into the design and synthesis of novel H2S probes while also offering experimental evidence for the application of PDMS in cellular imaging.

Altered Endoplasmic Reticulum Integrity and Organelle Interactions in Living Cells Expressing INF2 Variants.

The cytoskeleton mediates fundamental cellular processes by organizing inter-organelle interactions. Pathogenic variants of inverted formin 2 (INF2) CAAX isoform, an actin assembly factor that is predominantly expressed in the endoplasmic reticulum (ER), are linked to focal segmental glomerulosclerosis (FSGS) and Charcot-Marie-Tooth (CMT) neuropathy. To investigate how pathogenic INF2 variants alter ER integrity, we used high-resolution live imaging of HeLa cells. Cells expressing wild-type (WT) INF2 showed a predominant tubular ER with perinuclear clustering. Cells expressing INF2 FSGS variants that cause mild and intermediate disease induced more sheet-like ER, a pattern similar to that seen for cells expressing WT-INF2 that were treated with actin and microtubule (MT) inhibitors. Dual CMT-FSGS INF2 variants led to more severe ER dysmorphism, with a diffuse, fragmented ER and coarse INF2 aggregates. Proper organization of both F-actin and MT was needed to modulate the tubule vs. sheet conformation balance, while MT arrays regulated spatial expansion of tubular ER in the cell periphery. Pathogenic INF2 variants also induced mitochondria fragmentation and dysregulated mitochondria distribution. Such mitochondrial abnormalities were more prominent for cells expressing CMT-FSGS compared to those with FSGS variants, indicating that the severity of the dysfunction is linked to the degree of cytoskeletal disorganization. Our observations suggest that pathogenic INF2 variants disrupt ER continuity by altering interactions between the ER and the cytoskeleton that in turn impairs inter-organelle communication, especially at ER-mitochondria contact sites. ER continuity defects may be a common disease mechanism involved in both peripheral neuropathy and glomerulopathy.

Ultrabroadband Optical Diffraction Tomography.

Optical diffraction tomography (ODT) is a powerful noninvasive 3D imaging technique, but its combination with broadband light sources is difficult. In this study, we introduce ultrabroadband ODT, covering over 150 nm of visible spectral bandwidth with a lateral spatial resolution of 150 nm. Our work addresses a critical experimental gap by enabling the measurement of broadband refractive index changes in 3D samples, crucial information that is difficult to assess with existing methodologies. We present broadband, spectrally resolved ODT images of HeLa cells, obtained via pulse-shaping-based Fourier transform spectroscopy. The spectral observations enabled by ultrabroadband ODT, combined with material-dependent refractive index responses, allow for precise three-dimensional identification of nanoparticles within cellular structures. Our work represents a crucial step toward time and spectrally resolved tomography of complex 3D structures with implications for life and materials science applications.

Designing a Hypoxia-Activated Sensing Platform Using an Azo Group-Triggered Reaction with the Formation of Silicon Nanoparticles.

Hypoxia is known as a specific signal of various diseases, such as liver fibrosis. We designed a hypoxia-sensitive fluorometric approach that cleaved the azo bond (N═N) in the presence of hypoxia-controlled agents (sodium dithionite and azoreductase). 4-(2-Pyridylazo) resorcinol (Py-N═N-RC) bears a desirable hypoxia-responsive linker (N═N), and its azo bond breakup can only occur in the presence of sodium dithionite and azoreductase and leads to the release of 2,4-dihydroxyaniline, which can react with 3-[2-(2-aminoethylamino)ethylamino]propyltrimethoxysilane to generate yellow fluorescent silicon nanoparticles. This approach exhibited high selectivity and sensitivity toward both sodium dithionite and azoreductase over other potential interferences. The mouse liver microsome, which is known to contain azoreductase, was applied and confirmed the feasibility of the designed platform. Py-N═N-RC is expected to be a practical substrate for hypoxia-related biological analyses. Furthermore, silicon nanoparticles were successfully applied for Hela cell imaging owing to their negligible cytotoxicity and superb biocompatibility.

Novel pyrene-based Schiff base ligand as a fluorescent sensor for Al3+ and colorimetric sensor for Ni2+ ions detection: Synthesis, photo-physical and cell imaging studies

A simple fluorescent and colorimetric pyrene-based ((Z)-4-hydroxy-3-((pyren-1-ylmethylene)amino)benzoic acid (HPMB) chemosensor was synthesized through the condensation reaction between pyrene-1-carboxaldehyde and 3-amino-4-hydroxybenzoic acid. The structure of HPMB was elucidated using 1H NMR, MS, and UV–Visible spectroscopic techniques. The receptor HPMB displayed a detectable, colorimetric colour shift in the presence of Ni2+ and the colour of the solution was observed to be light brown. On the other hand, it also exhibited a fluorescent “turn on” response when Al3+ ions were added in DMSO solution (HEPES 0.01 M, pH = 7.4) at room temperature. Job’s plot showed that HPMB binding ratio to Ni2+ and Al3+ ions was in 1:1 ratio. The detection limit for Ni2+ and Al3+ ions was found to be 0.33 × 10-6 M and 0.26 × 10-6 M, respectively. In addition, HPMB was utilised to create a molecular logic gate, and the fluorescence imaging of HeLa cells was investigated because of HPMB’s low cytotoxicity, indicating that it could be used to detect Al3+ and Ni2+ in living cells.

Synthesis, Fluorescence, and Bioactivity of Novel Isatin Derivatives.

The isatin group is widespread in nature and is considered to be a privileged building block for drug discovery. In order to develop novel SHP1 inhibitors with fluorescent properties as tools for SHP1 biology research, this work designed and synthesized a series of isatin derivatives. The presentive compound 5a showed good inhibitory activity against SHP1PTP with IC50 of 11 ± 3 μM, displayed about 92% inhibitory rate against MV-4-11 cell proliferation at the concentration of 20 μM, exhibited suitable fluorescent properties with a long emission wavelength and a large Stokes shift, and presented blue fluorescent imaging in HeLa cells with low cytotoxicity. This study could offer chemical tool to further understand SHP1 biology and develop novel SHP1 inhibitors in therapy.

Polymorphic strategies for encapsulating polyindole@nido-carborane fluorescent polymeric nanocapsules: Their release, imaging, and biological evaluation in the gastrointestinal microenvironment

In this experiment, by combining water-soluble nido-carborane with indole dyes, and used non-toxic and non-irritating sodium alginate and acrylic resin as coating materials to obtain four fluorescent complexes, indol-nido-carborane-sodium alginate- Polymer-A (INS-A) 、indol-nido-carborane-sodium alginate-Polymer-B (INS-B)、indol-nido-carborane-sodium alginate-Polymer-C (INS-C) and indol-nido-carborane-sodium alginate-Polymer-D (INS-D).The transmission electron microscopy (TEM) results showed that the four complexes have different morphologies, with INS-A showing irregular clusters, INS-B being uniformly spherical, INS-C demonstrating a cashew-nut shape and INS-D manifesting a semi-encapsulated state. The spectroscopic properties and in vitro release experiments of the four complexes were performed in different simulated gastrointestinal environments. The results indicated that INS-A and INS-C had excellent photophysical properties and high cumulative release, so INS-A and INS-C were selected for subsequent experiments. Atomic Force Microscope (AFM) test manifested that the surfaces of INS-A and INS-C were relatively smooth. Imaging of HeLa cells with different channels was performed, the Pearson correlation coefficients of both INS-A and INS-C could be calculated to be distributed between 0.21 and 0.60, indicating that the complexes exhibited good imaging performance and could enter the cell and distribute around the nucleus. The results of cell proliferation toxicity (CCK8) experiments showed that at the concentration of 125 μg/mL, the inhibition rates of INS-A and INS-C on HCT-116 and HeLa cells were 47.15 %, 36.70 % and 42.81 %, 35.52 %, respectively, demonstrating that the complexes had an inhibitory effect on cancer cells. In summary, the fluorescent complexes prepared in this paper have good imaging ability and tumor targeting, and are potential fluorescent anticancer reagent.

Multifunctional Eu-doped carbon dots nanoprobe for highly sensitive and selective determination of glutathione in biological fluid and cell imaging

Glutathione (GSH) holds a vital role in biological systems, serving diverse cellular functions. Selective determination of GSH over cysteine (Cys) and homocysteine (Hcy) stays challenging due to the shared functional groups among these biothiols. In this study, a versatile Eu-doped carbon dots (Eu-CDs) nanoprobe has been developed for the exceptionally sensitive and selective detection of GSH in biological fluids. The synthesized Eu-CDs exhibited high solubility in water, stability to salt and pH variations, and consistent fluorescence emission, boasting a quantum yield of 40.67 %. Following the optimization of detection parameters, a robust linear relationship for GSH was established in the concentration range of 0.0–50 μM, with a detection limit of 0.03 μM. The viability of the technique in actual samples was evaluated by successfully measuring the GSH in urine and human serum samples with a range recovery from 90.6 % to 98.5 %. Furthermore, Eu-CDs proved to be exceptionally suitable for imaging Hela cells owing to their low cytotoxicity and remarkable biocompatibility. As a result, the developed fluorescent biosensor demonstrated significant potential for widespread application in clinical diagnostic analyses.

Smartphone-based sensing and in vivo and in vitro imaging of Mn(VII) based on nitrogen-doped red fluorescent carbon dots.

Smartphone-assisted visual assay platform provides novel insight for the real-time in-field quantitation of intended analytes in resource-insufficient areas. Herein, nitrogen-doped red fluorescent (FL) carbon dots (R-CDs) were developed for the timely on-site quantitation of Mn(VII) using the smartphone-assisted assay platform. R-CDs, possessing a desirable bright red FL at 616 nm under a 470 nm excitation, were fabricated through hydrothermal treatment adopting passion fruit and neutral red as precursors. Interestingly, bright red FL at 616 nm are gradually quenched upon introducing Mn(VII) based on the inner filter effect, concurrently accompanying with significant FL color variation from bright red to dark red. Inspired by the above-mentioned phenomena, hue-saturation-values (HSV) of real-time captured images could be precisely quantified through a color recognition APP within the smartphone, of which the V/S values could be employed to quantify Mn(VII) with a linear range of 50-400 μM. Furthermore, confocal fluorescence imaging of HeLa cells and zebrafish larvae demonstrates that R-CDs could be employed for the visual determination of Mn(VII) in vivo and in vitro, illustrating that R-CDs possess powerful practical application prospect in biosystem.

A “Turn-Off” Pyrene-Based Ligand as a Fluorescent Sensor for the Detection of Cu2+ and Fe2+ Ions: Synthesis and Application in Real Water Samples, Logic Gate Construction, and Bio-Imaging

Herein, we report the synthesis and characterization of a novel Schiff base ligand, (Z)-5-((pyren-1-ylmethylene)amino)-2,4-dihydro-3H-pyrazol-3-one (PMDP). The characterization of ligand PMDP was carried out using ESI-MS, 1H NMR, and UV–Visible spectroscopic techniques. As a probe, PMDP displayed a detectable, colorimetric colour shift in the presence of Cu2+ and Fe2+ ions. The solution was seen to have a light brown colour and to exhibit a fluorometric “turn off” response when Cu2+ and Fe2+ ions were present in a DMSO solution (HEPES 0.01 M, pH = 7.4) at room temperature. Job’s plot revealed that the PMDP binding ratio to Cu2+ and Fe2+ ions was in 1:2 ratio. In contrast to the other metal ions (Cd2+, Mn2+, Co2+, Na+, Ni2+, Cu+, Fe3+, Hg2+, Mg2+, Zn2+, K+, and V5+), the synthesised probe showed exceptional sensitivity and selectivity for detecting Cu2+ and Fe2+ metal ions. The results indicate that the detection limits for Cu2+ and Fe2+ are 0.42 μM and 0.51 μM, respectively. Furthermore, PMDP was efficiently utilised for the quantitative analysis of Cu2+ and Fe2+ in real water samples, RGB colour values in smart phones, logic gate construction, and cell imaging in HeLa cells.

Japanese encephalitis virus hijacks ER-associated degradation regulators for its replication.

Flaviviruses target their replication on membranous structures derived from the ER, where both viral and host proteins play crucial structural and functional roles. Here, we have characterized the involvement of the ER-associated degradation (ERAD) pathway core E3 ligase complex (SEL1L-HRD1) regulator proteins in the replication of Japanese encephalitis virus (JEV). Through high-resolution immunofluorescence imaging of JEV-infected HeLa cells, we observe that the virus replication complexes marked by NS1 strongly colocalize with the ERAD adapter SEL1L, lectin OS9, ER-membrane shuttle factor HERPUD1, E3 ubiquitin ligase HRD1 and rhomboid superfamily member DERLIN1. NS5 positive structures also show strong overlap with SEL1L. While these effectors show significant transcriptional upregulation, their protein levels remain largely stable in infected cells. siRNA mediated depletion of OS9, SEL1L, HERPUD1 and HRD1 significantly inhibit viral RNA replication and titres, with SEL1L depletion showing the maximum attenuation of replication. By performing protein translation arrest experiments, we show that SEL1L, and OS9 are stabilised upon JEV infection. Overall results from this study suggest that these ERAD effector proteins are crucial host-factors for JEV replication.

Folate Receptor Targeting Mn(II) Complex Encapsulated Porous Silica Nanoparticle as an MRI Contrast Agent for Early-State Detection of Cancer.

Cancer is recognized as one of the major causes of mortality, however, early-stage detection can increase the survival chance greatly. It is recognized that folate receptors are gradually overexpressed in the cellular membrane with the progress of cancer from stage 1 to stage 4. Utilizing the fact, herein, developed a porous silica nanoparticle system C1@SiO2-FA-NP; A) impregnated with thermodynamically stable Mn(II) complex (1) molecules within the core of the nanoparticle, and B) surface functionalized with folate units. It exhibited a high longitudinal relaxivity value r1 = 21.45 mM-1s-1 that substantially increased to r1 = 40.97 mM-1s-1 in the presence of 0.67mM concentration of BSA under the physiological condition. The in vitro fluorescent images after surface conjugation of C1@SiO2-FA-NP with FITC (fluorescein isothiocyanate) buttressed the inclusion of the nanoparticle exclusively within the cancerous HeLa cells than that of healthy HEK293 cells. The importance of the surface-bound folate unit in the nanoparticle is further established by comparing the fluorescent images of HeLa cells in the absence of the group. Finally, the applicability of C1@SiO2-FA-NP as the T1-weighted MRI contrast agent for early-stage cancer diagnosis is established within C57BL/6 mice after infecting the mice with HeLa cells.

Recognition and detection of histamine in foods using aptamer modified fluorescence polymer dots sensors

Histamine has been known as a momentous cause of biogenic amine poisoning. Therefore, the content of histamine in foods is strictly required to be controlled within a certain range. Here, an aptamer fluorescent sensor was developed for detection of histamine. Poly [(9, 9-di-n-octylfluorenyl-2, 7-diyl)-alt-(benzo [2,1,3] thiadia-zol-4, 8-diyl)] (PF8BT) and the styrene maleic anhydride copolymer (PSMA) were used for the preparation of PF8BT-Polymer dots (PF8BT-Pdots). PF8BT-Pdots and the cyanine3-phosphoramidite (Cy3) were linked through aptamer to achieve the ratiometric detection for histamine. PF8BT-Pdots were partly quenched by Cy3 due to the fluorescence resonance energy transfer (FRET), when the histamine molecule was recognized by aptamer on the surface of PF8BT-Pdots. A linear range (3–21 μmol/L) was obtained for histamine detection with a low limit of detection (LOD = 0.38 μmol/L). PF8BT aptamer Pdots (PF8BT-A) were used to detect histamine in simply treated aquaculture water and tuna. The cell imaging of HeLa cells presented a good biosecurity and outstanding fluorescent imaging capability of PF8BT-A. The aptamer fluorescent sensors provided a new platform for rapid and accurate detection of histamine in aquatic products and had great potential for the application in food safety and quality control.

Widely tunable fiber optical parametric oscillator synchronized with a Ti:sapphire laser for stimulated Raman scattering microscopy

Stimulated Raman scattering (SRS) microscopy is a powerful vibrational imaging technique with high chemical specificity. However, the insufficient tuning range or speed of light sources limits the spectral range of SRS imaging and, hence, the ability to identify molecular species. Here, we present a widely tunable fiber optical parametric oscillator with a tuning range of 1470 cm−1, which can be synchronized with a Ti:sapphire laser. By using the synchronized light sources, we develop an SRS imaging system that covers the fingerprint and C–H stretching regions, without balanced detection. We validate its broadband imaging capability by visualizing a mixed polymer sample in multiple vibrational modes. We also demonstrate SRS imaging of HeLa cells, showing the applicability of our SRS microscope to biological samples.