Advancing the frontiers of IR microscopy
mIRage® sub-micron IR sub-micron IR spectroscopy
and imaging with FTIR quality data
Non-contact reflection-based IR measurement with FTIR transmission-like spectral quality and no IR spectral artifacts
Fast, easy to use with little to no sample preparation and spectroscopy in seconds
Overcomes the limitations of Raman spectroscopy
![mIRage product photo with logo 800px](https://www.photothermal.com/wp-content/uploads/2023/06/mIRage-product-photo-with-logo-800px.png)
The mIRage® IR microscope is an innovative new microscopy system uniquely providing sub-micron IR spectroscopy and imaging across a wide variety of applications.
Using a proprietary technique based upon Optical Photothermal IR (O-PTIR) spectroscopy, mIRage® breaks the diffraction limit of traditional IR spectroscopy and bridges the gap between conventional IR microspectroscopy and nanoscale IR spectroscopy.
Click here to read our latest applications note for life science applications
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Life science – cells
Co-located fluorescence + sub-micron IR of cells
Neuroglioma cells were stained with G3BP1 for protein stress granules, DAPI for nucleus and BODIPY for lipids.
The left image: RBG overlay widefield epi-fluorescence image with red showing protein stress granules, Blue showing nucleus and Green showing lipids. Square markers show locations of O-PTIR spectral collection.
The center image shows the brightfield image. Square markers show locations of O-PTIR spectral collection.
The right side O-PTIR spectra was collected in seconds from the marker locations shown in the left and middle panes. Clear spectral differences can be observed, consistent with the targeted sub-cellular features. Of particular note, is the subtle shift in the Amide I band of the protein stress granule indicating a likely different protein secondary structure to the other locations.
![Co-located FL + sub-micron IR of cells - final-3](https://www.photothermal.com/wp-content/uploads/2022/12/Co-located-FL-sub-micron-IR-of-cells-final-3.png)
![Co-located FL + sub-micron IR of cells - final-3](https://www.photothermal.com/wp-content/uploads/2022/12/Co-located-FL-sub-micron-IR-of-cells-final-3.png)
![Co-located FL + sub-micron IR of cells - final-3](https://www.photothermal.com/wp-content/uploads/2022/12/Co-located-FL-sub-micron-IR-of-cells-final-3.png)
Water dipping objective, 40x, 1.0NA
Spectra collected in seconds
Images in minutes
Images collected in single frequency mode with 50nm step size
Probe beam detected in single frequency mode
No water background compensation
Spectra show little to no water absorbances
![Cheek cells - final-1](https://www.photothermal.com/wp-content/uploads/2022/12/Cheek-cells-final-1.jpg)
![Cheek cells - final-1](https://www.photothermal.com/wp-content/uploads/2022/12/Cheek-cells-final-1.jpg)
![Cheek cells - final-1](https://www.photothermal.com/wp-content/uploads/2022/12/Cheek-cells-final-1.jpg)
![Ultra high NA water dipping objective2](https://www.photothermal.com/wp-content/uploads/2023/10/Ultra-high-NA-water-dipping-objective2.png)
![Ultra high NA water dipping objective2](https://www.photothermal.com/wp-content/uploads/2023/10/Ultra-high-NA-water-dipping-objective2.png)
oil immersion objective
Cheek cells on glass coverslip (170nm)
Oil immersion objective, 100x, NA 1.30
Spectra collected in seconds
Images in minutes
Images collected in single frequency mode with 50nm step size
Probe beam detected in single frequency mode
Spatial resolution of ~285nm
![O-PTIR of fixed cell with oil immersion objective - final-2](https://www.photothermal.com/wp-content/uploads/2022/12/O-PTIR-of-fixed-cell-with-oil-immersion-objective-final-2.jpg)
![O-PTIR of fixed cell with oil immersion objective - final-2](https://www.photothermal.com/wp-content/uploads/2022/12/O-PTIR-of-fixed-cell-with-oil-immersion-objective-final-2.jpg)
![Ultra high NA oil immersion objective](https://www.photothermal.com/wp-content/uploads/2023/09/Ultra-high-NA-oil-immersion-objective.png)
![Ultra high NA oil immersion objective](https://www.photothermal.com/wp-content/uploads/2023/09/Ultra-high-NA-oil-immersion-objective.png)
Submicron amyloid aggregate
imaging in neurons
Left; O-PTIR, single frequency ratio image of 1630/1656cm-1. Shows distribution of beta protein structures with separation of 282nm! Right; O-PTIR spectra from IR image (left) showing spectra on (#1) and off (#2) the beta protein structure. Spectral differences, clearly show the differences in the amide I band, typical of beta sheet structured proteins, despite these two locations only being separated by 282nm!
Published: Oxana Klementieva et al., “Super-resolution infrared imaging of polymorphic amyloid aggregates directly in neurons”, Adv Sci, Adv. Sci. 2020, 1903004 https://doi.org/10.1002/advs.201903004
O-PTIR image, 1630/1656 O-PTIR spectra
![Amyloid Aggregate 7 Data image of submicron amyloid aggregate imaging in neurons](https://www.photothermal.com/wp-content/uploads/2023/09/Amyloid-Aggregate-7.jpg)
![Amyloid Aggregate 7 Data image of submicron amyloid aggregate imaging in neurons](https://www.photothermal.com/wp-content/uploads/2023/09/Amyloid-Aggregate-7.jpg)
Targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes
Both IR images at the top were collected at 100nm pixel size. ~5 mins per image. At the bottom right is the spectra from markers (spectra are single scans, ~1sec measurement time, no processing. Data collected using the new “Dual range (C-H/FP)” QCL, with spectral range coverage of 3000-2700, 1800-950cm-1.
Sample courtesy of Prof Jose Sule-Suso, Keele University, UK.
Publication in preparation (Dec, 2020)
Lipid chain length image
2856 (CH2)/2874 (CH3)
Lipid relative to protein
2856 (CH2)/ 1658 (Protein)
![Targeted Imaging Mode 2 Data illustration showing targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes](https://www.photothermal.com/wp-content/uploads/2021/09/Targeted-Imaging-Mode-2.jpg)
![Targeted Imaging Mode 2 Data illustration showing targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes](https://www.photothermal.com/wp-content/uploads/2021/09/Targeted-Imaging-Mode-2.jpg)
![Targeted Imaging Mode 2 Data illustration showing targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes](https://www.photothermal.com/wp-content/uploads/2021/09/Targeted-Imaging-Mode-2.jpg)
Using fluorescence to localize O-PTIR measurements
An Alzheimer’s disease mouse model brain tissue section was stained with Amytracker 630 to highlight amyloid aggregates, AF488 to highlight proteins and DAPI for the nucleus.
In the figure to the right is shown a brightfield image, top left, of the stained sample. In the top right is the RBG composite fluorescence image, which highlights in red/orange the regions of amyloid aggregation. Note how some amyloid aggregates highlighted in the fluorescence image are not readily distinguishable in the brightfield image. At the bottom is an averaged O-PTIR spectra, from the line profile indicated in the fluorescence image, with spectra averaged on (in blue) and off (in red) the aggregate. The average spectrum of the aggregate shows distinct spectral differences in the amide I band with a significant spectral feature at 1631cm-1, typical of protein beta sheet structures. This clearly demonstrates the utility of combining fluorescence imaging to highlight regions of amyloid aggregation, some of which cannot be readily seen in brightfield microscopy, with submicrometer O-PTIR spectroscopy which can then provide the molecular compositional information, in this case, being particularly sensitive to protein secondary structure, a characteristic strength of IR spectroscopy.
![Using fluorescence to localize O-PTIR measurements-3 - final](https://www.photothermal.com/wp-content/uploads/2022/12/Using-fluorescence-to-localize-O-PTIR-measurements-3-final.jpg)
![Using fluorescence to localize O-PTIR measurements-3 - final](https://www.photothermal.com/wp-content/uploads/2022/12/Using-fluorescence-to-localize-O-PTIR-measurements-3-final.jpg)
(top left) The brightfield image of the stained sample. (top right) The RBG composite fluorescence image. (bottom) An averaged O-PTIR spectra, from the line profile shown in the RBG composite fluorescence image.
IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon
A: Spectra obtained with O-PTIR from control tendon fibrils on CaF2 window. B: Single frequency image at right recorded at 1655 cm-1 in perpendicular orientation. markers denote locations at which spectra were acquired. Scale bar = 1µm
C and D: Optical photothermal IR (O-PTIR) spectra from intact tendon, from ~500 nm measurement spots. (B) Individual spectra obtained from the two orientations of a section mounted on a CaF2 window, relative to the linearly polarized QCL. Inserted visual image shows the 6 locations, all of which lie within the region imaged with FTIR FPA; scale bar = 70 μm.
Colored markers (+) correspond to spectral colors. (C) Comparison of spectra obtained from CaF2 (top) and glass (bottom) substrates in parallel and perpendicular orientations to linearly polarized QCL.
Published: Gorker Bakir et al., “Orientation Matters: Polarization Dependent IR Spectroscopy of Collagen from Intact Tendon Down to the Single Fibril Level”, Molecules 2020, 25, 4295 https://www.mdpi.com/1420-3049/25/18/4295
![collagen orientation fibrils and tendon - square Data images showing IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon](https://www.photothermal.com/wp-content/uploads/2023/10/collagen-orientation-fibrils-and-tendon-square.png)
![collagen orientation fibrils and tendon - square Data images showing IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon](https://www.photothermal.com/wp-content/uploads/2023/10/collagen-orientation-fibrils-and-tendon-square.png)
![collagen orientation fibrils and tendon - square Data images showing IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon](https://www.photothermal.com/wp-content/uploads/2023/10/collagen-orientation-fibrils-and-tendon-square.png)
Breast tissue calcification – Demonstration of <1 micron spatial resolution with O-PTIR
A: Optical image (mosaic). Red box indicates IR image measurement area. B: Single frequency image at 1050cm-1 to highlight calcification locations. C: O-PTIR Spectra from colored circle markers in IR image (B).
IR image area 200×200 microns at 500nm step size. Image time, ~10mins.
Calcification IR image at 1050cm-1, clearly resolves calcifications averaging only a few microns in size, many even <1 micron. At 1050cm-1, traditional FTIR has a spatial of ~12microns, which is much larger than the actual features, which is why such small an localized calcifications had not been seen before.
Sample courtesy of Prof Nick Stone, Exeter University, UK. Publication in preparation (Dec, 2020)
![Breast Tissue Calcification vertical copy Data images of breast tissue calcification, demonstration of sub micron spatial resolution with O-PTIR](https://www.photothermal.com/wp-content/uploads/2023/10/Breast-Tissue-Calcification-vertical-copy.png)
![Breast Tissue Calcification vertical copy Data images of breast tissue calcification, demonstration of sub micron spatial resolution with O-PTIR](https://www.photothermal.com/wp-content/uploads/2023/10/Breast-Tissue-Calcification-vertical-copy.png)
![Breast Tissue Calcification vertical copy Data images of breast tissue calcification, demonstration of sub micron spatial resolution with O-PTIR](https://www.photothermal.com/wp-content/uploads/2023/10/Breast-Tissue-Calcification-vertical-copy.png)
Single bacterial cell O-PTIR microscopy with deuterium labelled E. coli
![QCL options CH and silent region with bacteria-web size](https://www.photothermal.com/wp-content/uploads/2023/10/QCL-options-CH-and-silent-region-with-bacteria-web-size.jpg)
![QCL options CH and silent region with bacteria-web size](https://www.photothermal.com/wp-content/uploads/2023/10/QCL-options-CH-and-silent-region-with-bacteria-web-size.jpg)
![QCL options CH and silent region with bacteria-web size](https://www.photothermal.com/wp-content/uploads/2023/10/QCL-options-CH-and-silent-region-with-bacteria-web-size.jpg)
Single bacterial cell simultaneous submicron IR+Raman microscopy
SNR of the OPTIR (~500nm spot) is ~4000:1 (RMS, taking amide band intensity as the peak and the baseline noise at the amide I position measured on a CaF2 blank) with ~20 sec accumulations.
![Simultaneous IR+Raman of bacteria-final real](https://www.photothermal.com/wp-content/uploads/2022/12/Simultaneous-IRRaman-of-bacteria-final-real.jpg)
![Simultaneous IR+Raman of bacteria-final real](https://www.photothermal.com/wp-content/uploads/2022/12/Simultaneous-IRRaman-of-bacteria-final-real.jpg)
![Simultaneous IR+Raman of bacteria-final real](https://www.photothermal.com/wp-content/uploads/2022/12/Simultaneous-IRRaman-of-bacteria-final-real.jpg)
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Microplastics
Sub-micron IR shape and size independent spectra
As a demonstration of O-PTIR capability to measure various particle shapes and sizes with artifact-free data collection in the presence of highly scattering salt crystals, we created a mixed model sample consisting of Polystyrene (PS) beads (900nm, 2μm, 4.5μm and 10μm) and Poly(methyl methacrylate) (PMMA). When using an FT-IR/QCL system, not only would these particles sizes be too small to measure, but the range of different particle sizes and the nearby presence of salt crystals would generate dispersive scatter artifacts which can significantly alter the spectra making accurate identification more difficult. To further demonstrate the relative immunity to scattering artifacts, in the figure to the right is shown these mixed polymer bead samples suspended in salt water and deposited on to a CaF2 substrate, so that the polymer beads would be interspersed with salt crystals.
It is noteworthy that despite the spectra being collected from differently sized beads the spectra all look very consistent with their polymer type (i.e., no dispersive scatter artifacts are present). This ensures that correct identification can be determined, regardless of particle shape and size.
![Microplastics IR+Raman in seconds](https://www.photothermal.com/wp-content/uploads/2023/10/Microplastics-IRRaman-in-seconds.png)
![Microplastics IR+Raman in seconds](https://www.photothermal.com/wp-content/uploads/2023/10/Microplastics-IRRaman-in-seconds.png)
![Microplastics IR+Raman in seconds](https://www.photothermal.com/wp-content/uploads/2023/10/Microplastics-IRRaman-in-seconds.png)
Fluorescence imaging + O-PTIR of microplastics
Fluorescence tagging of polymeric beads can help to isolate the polymer particles from other particles for measurement with O-PTIR, thus dramatically speeding up analysis
![Fluorescence imaging of microplatics web](https://www.photothermal.com/wp-content/uploads/2023/10/Fluorescence-imaging-of-microplatics-web.png)
![Fluorescence imaging of microplatics web](https://www.photothermal.com/wp-content/uploads/2023/10/Fluorescence-imaging-of-microplatics-web.png)
![Fluorescence imaging of microplatics web](https://www.photothermal.com/wp-content/uploads/2023/10/Fluorescence-imaging-of-microplatics-web.png)
Sub-micron IR+Raman microplastics
![microplastics spectra and images-1](https://www.photothermal.com/wp-content/uploads/2023/10/microplastics-spectra-and-images-1.png)
![microplastics spectra and images-1](https://www.photothermal.com/wp-content/uploads/2023/10/microplastics-spectra-and-images-1.png)
![microplastics spectra and images-1](https://www.photothermal.com/wp-content/uploads/2023/10/microplastics-spectra-and-images-1.png)
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Polymers
Polymer laminates analysis with O-PTIR
- Key peaks at 1642 cm-1 (Nylon) and 1142cm-1 are used for single frequency imaging
- Image collected at 100nm steps (~3mins per image)
- Central EVOH layer of 1.6microns clearly visible!
![Polymers laminates analysis - attribution](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis-attribution.jpg)
![Polymers laminates analysis - attribution](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis-attribution.jpg)
![Polymers laminates analysis - attribution](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis-attribution.jpg)
![Polymer laminates analysis with O-PTIR Data image of polymer laminates analysis with O-PTIR](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis.jpg)
![Polymer laminates analysis with O-PTIR Data image of polymer laminates analysis with O-PTIR](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis.jpg)
![Polymer laminates analysis with O-PTIR Data image of polymer laminates analysis with O-PTIR](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis.jpg)
O-PTIR – polymer (PLA-ACM)
phase dispersions
Clear spectral differences attributable to the expected chemical domains of PLA and ACM were observed.
IR image: 20x20um, 100nm step size, ~3min/image
Sample courtesy of Dr Rudiger Berger, Max Planck Inst Polymer Research, Mainz, Germany
![Polymers PLA-ACM-2 Data illustrating O-PTIR – polymer (PLA-ACM) phase dispersions](https://www.photothermal.com/wp-content/uploads/2022/08/Polymers-PLA-ACM-2.jpg)
![Polymers PLA-ACM-2 Data illustrating O-PTIR – polymer (PLA-ACM) phase dispersions](https://www.photothermal.com/wp-content/uploads/2022/08/Polymers-PLA-ACM-2.jpg)
![Polymers PLA-ACM-2 Data illustrating O-PTIR – polymer (PLA-ACM) phase dispersions](https://www.photothermal.com/wp-content/uploads/2022/08/Polymers-PLA-ACM-2.jpg)
Imaging and spectroscopy of bioplastic laminates
O-PTIR scan of bioplastic laminate
Linear sampling scan spanning 8.0 µm measured every 100 nm apart (plotted only every 200 nm and across 2 µm for clarity) across the boundary of the bioplastic laminate, moving from the pure PHBHx layer to the pure PLA layer.
Gradual spectral changes over the space much greater than the optical resolution suggest the mixed distribution of PLA and PHBHx without any sharp boundary.
No clear isosbestic point indicates that the system is not a simple binary mixture.
PLA and PHBHx contributions are overlapped and mingled in the fingerprint region
Composite (red/green) single frequency images
![bio-Polymers-singleFrequencyDiagram - vertical3](https://www.photothermal.com/wp-content/uploads/2024/01/bio-Polymers-singleFrequencyDiagram-vertical3.png)
![bio-Polymers-singleFrequencyDiagram - vertical3](https://www.photothermal.com/wp-content/uploads/2024/01/bio-Polymers-singleFrequencyDiagram-vertical3.png)
Spatial resolution breakthrough with O-PTIR
Theoretical spatial resolution comparisons
(FTIR, QCL and O-PTIR microscopes)
![Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes](https://www.photothermal.com/wp-content/uploads/2020/12/comparison-chart-2.jpg)
![Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes](https://www.photothermal.com/wp-content/uploads/2020/12/comparison-chart-2.jpg)
![Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes](https://www.photothermal.com/wp-content/uploads/2020/12/comparison-chart-2.jpg)
New “Dual Range (C-H/FP)” QCL
![Spectra showing new “Dual Range (C-H/FP)” QCL Spectra showing new “Dual Range (C-H/FP)” QCL](https://www.photothermal.com/wp-content/uploads/2020/12/Wide-QCL-spectra-2.jpg)
![Spectra showing new “Dual Range (C-H/FP)” QCL Spectra showing new “Dual Range (C-H/FP)” QCL](https://www.photothermal.com/wp-content/uploads/2020/12/Wide-QCL-spectra-2.jpg)
![Spectra showing new “Dual Range (C-H/FP)” QCL Spectra showing new “Dual Range (C-H/FP)” QCL](https://www.photothermal.com/wp-content/uploads/2020/12/Wide-QCL-spectra-2.jpg)
Library (Wiley KnowItAll) search results delivered >95% match
O-PTIR spectra collected in reflection mode. Displayed spectra are raw and unprocessed (<5sec collection time, ~500nm spot size)
O-PTIR spectra are measured off thick polymers (mm’s), vs library FTIR references data off thin films (~10microns)
Publications
Brochure
Datasheet
Advancing the frontiers of IR microscopy
mIRage® sub-micron IR sub-micron IR spectroscopy and imaging with FTIR quality data
Non-contact reflection-based IR measurement with FTIR transmission-like spectral quality and no IR spectral artifacts
Fast, easy to use with little to no sample preparation and spectroscopy in seconds
Overcomes the limitations of Raman spectroscopy
![mIRage product photo with logo 800px](https://www.photothermal.com/wp-content/uploads/2023/06/mIRage-product-photo-with-logo-800px.png)
![mIRage product photo with logo 800px](https://www.photothermal.com/wp-content/uploads/2023/06/mIRage-product-photo-with-logo-800px.png)
![mIRage product photo with logo 800px](https://www.photothermal.com/wp-content/uploads/2023/06/mIRage-product-photo-with-logo-800px.png)
The mIRage® IR microscope is an innovative new microscopy system uniquely providing sub-micron IR spectroscopy and imaging across a wide variety of applications.
Using a proprietary technique based upon Optical Photothermal IR (O-PTIR) spectroscopy, mIRage® breaks the diffraction limit of traditional IR spectroscopy and bridges the gap between conventional IR microspectroscopy and nanoscale IR spectroscopy.
Applications
Click here to read our latest applications note for life science applications
Life science – cells
Co-located fluorescence + sub-micron IR of cells
![Co-located FL + sub-micron IR of cells - final-3](https://www.photothermal.com/wp-content/uploads/2022/12/Co-located-FL-sub-micron-IR-of-cells-final-3.png)
![Co-located FL + sub-micron IR of cells - final-3](https://www.photothermal.com/wp-content/uploads/2022/12/Co-located-FL-sub-micron-IR-of-cells-final-3.png)
![Co-located FL + sub-micron IR of cells - final-3](https://www.photothermal.com/wp-content/uploads/2022/12/Co-located-FL-sub-micron-IR-of-cells-final-3.png)
O-PTIR of fixed cell with oil immersion objective
![O-PTIR of fixed cell with oil or water immersion objective - vertical 2](https://www.photothermal.com/wp-content/uploads/2023/12/O-PTIR-of-fixed-cell-with-oil-or-water-immersion-objective-vertical-2-803x1024.png)
![O-PTIR of fixed cell with oil or water immersion objective - vertical 2](https://www.photothermal.com/wp-content/uploads/2023/12/O-PTIR-of-fixed-cell-with-oil-or-water-immersion-objective-vertical-2-803x1024.png)
• Cheek cells on glass coverslip (170nm)
• Oil immersion objective, 100x, NA1.30
• Spectra collected in seconds
• Images in minutes
• Images collected in single frequency mode with 50nm steps size
• Probe beam detected in reflection mode
• Spatial resolution of ~285nm
Submicron amyloid aggregate imaging in neurons
![Amyloid Aggregate 7 C Data image of submicron amyloid aggregate imaging in neurons](https://www.photothermal.com/wp-content/uploads/2023/11/Amyloid-Aggregate-7-C.jpg)
![Amyloid Aggregate 7 C Data image of submicron amyloid aggregate imaging in neurons](https://www.photothermal.com/wp-content/uploads/2023/11/Amyloid-Aggregate-7-C.jpg)
Left; O-PTIR, single frequency ratio image of 1630/1656cm-1. Shows distribution of beta protein structures with separation of 282nm! Right; O-PTIR spectra from IR image (left) showing spectra on (#1) and off (#2) the beta protein structure. Spectral differences, clearly show the differences in the amide I band, typical of beta sheet structured proteins, despite these two locations only being separated by 282nm!
Published: Oxana Klementieva et al., “Super-resolution infrared imaging of polymorphic amyloid aggregates directly in neurons”, Adv Sci, Adv. Sci. 2020, 1903004 https://doi.org/10.1002/advs.201903004
O-PTIR measurement of cell in water (H2O) with water dipping objective
![Cheek cells - final-1](https://www.photothermal.com/wp-content/uploads/2022/12/Cheek-cells-final-1.jpg)
![Cheek cells - final-1](https://www.photothermal.com/wp-content/uploads/2022/12/Cheek-cells-final-1.jpg)
![Cheek cells - final-1](https://www.photothermal.com/wp-content/uploads/2022/12/Cheek-cells-final-1.jpg)
• Water dipping objective, 40x, 1.0NA
• Spectra collected in seconds
• Images in minutes
• Images collected in single frequency mode with 50nm steps size
• Probe beam detected in transmission mode
• No water background compensation
• Spectra show little to no water absorbances
Targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes
Lipid chain length image
2856 (CH2)/2874 (CH3)
Lipid relative to protein
2856 (CH2)/ 1658 (Protein)
![Targeted Imaging Mode 2 Data illustration showing targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes](https://www.photothermal.com/wp-content/uploads/2021/09/Targeted-Imaging-Mode-2.jpg)
![Targeted Imaging Mode 2 Data illustration showing targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes](https://www.photothermal.com/wp-content/uploads/2021/09/Targeted-Imaging-Mode-2.jpg)
![Targeted Imaging Mode 2 Data illustration showing targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes](https://www.photothermal.com/wp-content/uploads/2021/09/Targeted-Imaging-Mode-2.jpg)
Top Left: Lipid Chain length image (2856cm-1 (CH2)/ 2874cm-1 (CH3). Top Right: : Lipid relative to protein image (2856cm-1) (CH2)/ 1658cm-1). Both IR images collected at 100nm pixel size. ~5 mins per image. Bottom Right: O-PTIR Spectra from markers in images (spectra are single scans, ~1sec measurement time, no processing. Bottom Left: Optical image.
Data collected using the new “Dual range (C-H/FP)” QCL, with spectral range coverage of 3000-2700, 1800-950cm-1.
Sample courtesy of Prof Jose Sule-Suso, Keele University, UK.
Publication in preparation (Dec, 2020)
Life science: tissue
Using fluorescence to localize O-PTIR measurements
![Using fluorescence to localize O-PTIR measurements-3 - final](https://www.photothermal.com/wp-content/uploads/2022/12/Using-fluorescence-to-localize-O-PTIR-measurements-3-final-1024x873.jpg)
![Using fluorescence to localize O-PTIR measurements-3 - final](https://www.photothermal.com/wp-content/uploads/2022/12/Using-fluorescence-to-localize-O-PTIR-measurements-3-final-1024x873.jpg)
An Alzheimer’s disease mouse model brain tissue section was stained with Amytracker 630 to highlight amyloid aggregates, AF488 to highlight proteins and DAPI
for the nucleus. In the figure above, on the top left, is shown a bright field image of the stained sample. In the top right is the RBG composite fluorescence image, which highlights in red/orange the regions of amyloid aggregation. Note how some amyloid aggregates highlighted in the fluorescence image are not readily distinguishable in the bright field image. At the bottom is an averaged O-PTIR spectra, from the line profile indicated in the fluorescence image, with spectra averaged on (in blue) and off (in red) the aggregate. The average spectrum of the aggregate shows distinct spectral differences in the amide I band with a significant spectral feature at 1631cm-1, typical of protein beta sheet structures. This clearly demonstrates the utility of combining fluorescence imaging to highlight regions of amyloid aggregation, some of which cannot be readily seen in brightfield microscopy, with submicrometer O-PTIR spectroscopy which can then provide the molecular compositional information, in this case, being particularly sensitive to protein secondary structure, a characteristic strength of IR spectroscopy.
IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon
![collagen orientation fibrils and tendon - square Data images showing IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon](https://www.photothermal.com/wp-content/uploads/2023/10/collagen-orientation-fibrils-and-tendon-square.png)
![collagen orientation fibrils and tendon - square Data images showing IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon](https://www.photothermal.com/wp-content/uploads/2023/10/collagen-orientation-fibrils-and-tendon-square.png)
![collagen orientation fibrils and tendon - square Data images showing IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon](https://www.photothermal.com/wp-content/uploads/2023/10/collagen-orientation-fibrils-and-tendon-square.png)
A: Spectra obtained with O-PTIR from control tendon fibrils on CaF2 window. B: Single frequency image at right recorded at 1655 cm-1 in perpendicular orientation. markers denote locations at which spectra were acquired. Scale bar = 1µm
C and D: Optical photothermal IR (O-PTIR) spectra from intact tendon, from ~500 nm measurement spots. (B) Individual spectra obtained from the two orientations of a section mounted on a CaF2 window, relative to the linearly polarized QCL. Inserted visual image shows the 6 locations, all of which lie within the region imaged with FTIR FPA; scale bar = 70 μm.
Colored markers (+) correspond to spectral colors. (C) Comparison of spectra obtained from CaF2 (top) and glass (bottom) substrates in parallel and perpendicular orientations to linearly polarized QCL.
Published: Gorker Bakir et al., “Orientation Matters: Polarization Dependent IR Spectroscopy of Collagen from Intact Tendon Down to the Single Fibril Level”, Molecules 2020, 25, 4295 https://www.mdpi.com/1420-3049/25/18/4295
Breast tissue calcification – Demonstration of <1 micron spatial resolution with O-PTIR
![Breast Tissue Calcification vertical copy Data images of breast tissue calcification, demonstration of sub micron spatial resolution with O-PTIR](https://www.photothermal.com/wp-content/uploads/2023/10/Breast-Tissue-Calcification-vertical-copy.png)
![Breast Tissue Calcification vertical copy Data images of breast tissue calcification, demonstration of sub micron spatial resolution with O-PTIR](https://www.photothermal.com/wp-content/uploads/2023/10/Breast-Tissue-Calcification-vertical-copy.png)
![Breast Tissue Calcification vertical copy Data images of breast tissue calcification, demonstration of sub micron spatial resolution with O-PTIR](https://www.photothermal.com/wp-content/uploads/2023/10/Breast-Tissue-Calcification-vertical-copy.png)
A: Optical image (mosaic). Red box indicates IR image measurement area. B: Single frequency image at 1050cm-1 to highlight calcification locations. C: O-PTIR Spectra from colored circle markers in IR image (B).
IR image area 200×200 microns at 500nm step size. Image time, ~10mins.
Calcification IR image at 1050cm-1, clearly resolves calcifications averaging only a few microns in size, many even <1 micron. At 1050cm-1, traditional FTIR has a spatial of ~12microns, which is much larger than the actual features, which is why such small an localized calcifications had not been seen before.
Sample courtesy of Prof Nick Stone, Exeter University, UK. Publication in preparation (Dec, 2020)
Life science: bacteria
Single bacterial cell O-PTIR microscopy with deuterium labelled E. coli
![QCL options CH and silent region with bacteria-web size](https://www.photothermal.com/wp-content/uploads/2023/10/QCL-options-CH-and-silent-region-with-bacteria-web-size.jpg)
![QCL options CH and silent region with bacteria-web size](https://www.photothermal.com/wp-content/uploads/2023/10/QCL-options-CH-and-silent-region-with-bacteria-web-size.jpg)
![QCL options CH and silent region with bacteria-web size](https://www.photothermal.com/wp-content/uploads/2023/10/QCL-options-CH-and-silent-region-with-bacteria-web-size.jpg)
Single bacterial cell simultaneous submicron IR+Raman microscopy
![Simultaneous IR+Raman of bacteria-final real](https://www.photothermal.com/wp-content/uploads/2022/12/Simultaneous-IRRaman-of-bacteria-final-real.jpg)
![Simultaneous IR+Raman of bacteria-final real](https://www.photothermal.com/wp-content/uploads/2022/12/Simultaneous-IRRaman-of-bacteria-final-real.jpg)
![Simultaneous IR+Raman of bacteria-final real](https://www.photothermal.com/wp-content/uploads/2022/12/Simultaneous-IRRaman-of-bacteria-final-real.jpg)
SNR of the OPTIR (~500nm spot) is ~4000:1 (RMS, taking amide band intensity as the peak and the baseline noise at the amide I position measured on a CaF2 blank) with ~20 sec accumulations.
Microplastics
![Microplastics IR+Raman in seconds](https://www.photothermal.com/wp-content/uploads/2023/10/Microplastics-IRRaman-in-seconds.png)
![Microplastics IR+Raman in seconds](https://www.photothermal.com/wp-content/uploads/2023/10/Microplastics-IRRaman-in-seconds.png)
![Microplastics IR+Raman in seconds](https://www.photothermal.com/wp-content/uploads/2023/10/Microplastics-IRRaman-in-seconds.png)
Fluorescence imaging + O-PTIR of microplastics
Fluorescence tagging of polymeric beads can help to isolate the polymer particles from other particles for measurement with O-PTIR, thus dramatically speeding up analysis
![Fluorescence imaging of microplatics web](https://www.photothermal.com/wp-content/uploads/2023/10/Fluorescence-imaging-of-microplatics-web.png)
![Fluorescence imaging of microplatics web](https://www.photothermal.com/wp-content/uploads/2023/10/Fluorescence-imaging-of-microplatics-web.png)
![Fluorescence imaging of microplatics web](https://www.photothermal.com/wp-content/uploads/2023/10/Fluorescence-imaging-of-microplatics-web.png)
Sub-micron IR+Raman microplastics
![microplastics spectra and images-1](https://www.photothermal.com/wp-content/uploads/2023/10/microplastics-spectra-and-images-1.png)
![microplastics spectra and images-1](https://www.photothermal.com/wp-content/uploads/2023/10/microplastics-spectra-and-images-1.png)
![microplastics spectra and images-1](https://www.photothermal.com/wp-content/uploads/2023/10/microplastics-spectra-and-images-1.png)
mIRage locates PS (0.9 µm, 2.0 µm, 4.5 µm and 10 µm) and PMMA beads (3.0 µm) in salt crystal mixture in hi-res IR images at key absorption bands. Distortion free spectra, even amongst salt crystals at hotspots, confirm the identity of the microplastics and readily searched against IR database. Importantly, and unlike traditional FTIR/QCL systems, spectra are consistent, regardless of particle shape or size when measured in reflection mode – no dispersive scatter artefacts.
Polymers
Polymer laminates analysis with O-PTIR
- Key peaks at 1642 cm-1 (Nylon) and 1142cm-1 are used for single frequency imaging
- Image collected at 100nm steps (~3mins per image)
- Central EVOH layer of 1.6microns clearly visible!
![Polymer laminates analysis with O-PTIR Data image of polymer laminates analysis with O-PTIR](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis.jpg)
![Polymer laminates analysis with O-PTIR Data image of polymer laminates analysis with O-PTIR](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis.jpg)
![Polymer laminates analysis with O-PTIR Data image of polymer laminates analysis with O-PTIR](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis.jpg)
![Polymers laminates analysis - attribution](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis-attribution.jpg)
![Polymers laminates analysis - attribution](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis-attribution.jpg)
![Polymers laminates analysis - attribution](https://www.photothermal.com/wp-content/uploads/2020/12/Polymers-laminates-analysis-attribution.jpg)
O-PTIR – polymer (PLA-ACM)
phase dispersions
![Polymers PLA-ACM-2 Data illustrating O-PTIR – polymer (PLA-ACM) phase dispersions](https://www.photothermal.com/wp-content/uploads/2022/08/Polymers-PLA-ACM-2.jpg)
![Polymers PLA-ACM-2 Data illustrating O-PTIR – polymer (PLA-ACM) phase dispersions](https://www.photothermal.com/wp-content/uploads/2022/08/Polymers-PLA-ACM-2.jpg)
![Polymers PLA-ACM-2 Data illustrating O-PTIR – polymer (PLA-ACM) phase dispersions](https://www.photothermal.com/wp-content/uploads/2022/08/Polymers-PLA-ACM-2.jpg)
Clear spectral differences attributable to the expected chemical domains of PLA and ACM were observed.
IR image: 20x20um, 100nm step size, ~3min/image
Sample courtesy of Dr Rudiger Berger, Max Planck Inst Polymer Research, Mainz, Germany
Imaging and spectroscopy of bioplastic laminates
![bio-Polymers-singleFrequencyDiagram](https://www.photothermal.com/wp-content/uploads/2019/08/bio-Polymers-singleFrequencyDiagram.png)
![bio-Polymers-singleFrequencyDiagram](https://www.photothermal.com/wp-content/uploads/2019/08/bio-Polymers-singleFrequencyDiagram.png)
![bio-Polymers-threeSpectras](https://www.photothermal.com/wp-content/uploads/2019/08/bio-Polymers-threeSpectras.png)
![bio-Polymers-threeSpectras](https://www.photothermal.com/wp-content/uploads/2019/08/bio-Polymers-threeSpectras.png)
Linear sampling scan spanning 8.0 µm measured every 100 nm apart (plotted only every 200 nm and across 2 µm for clarity) across the boundary of the bioplastic laminate, moving from the pure PHBHx layer to the pure PLA layer.
Gradual spectral changes over the space much greater than the optical resolution suggest the mixed distribution of PLA and PHBHx without any sharp boundary.
No clear isosbestic point indicates that the system is not a simple binary mixture.
PLA and PHBHx contributions are overlapped and mingled in the fingerprint region
Spatial resolution breakthrough with O-PTIR
Theoretical spatial resolution comparisons
(FTIR, QCL and O-PTIR microscopes)
![Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes](https://www.photothermal.com/wp-content/uploads/2020/12/comparison-chart-2.jpg)
![Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes](https://www.photothermal.com/wp-content/uploads/2020/12/comparison-chart-2.jpg)
![Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes Chart showing theoretical spatial resolution comparisons of FTIR, QCL and O-PTIR microscopes](https://www.photothermal.com/wp-content/uploads/2020/12/comparison-chart-2.jpg)
New “Dual Range (C-H/FP)” QCL
![Spectra showing new “Dual Range (C-H/FP)” QCL Spectra showing new “Dual Range (C-H/FP)” QCL](https://www.photothermal.com/wp-content/uploads/2020/12/Wide-QCL-spectra-2.jpg)
![Spectra showing new “Dual Range (C-H/FP)” QCL Spectra showing new “Dual Range (C-H/FP)” QCL](https://www.photothermal.com/wp-content/uploads/2020/12/Wide-QCL-spectra-2.jpg)
![Spectra showing new “Dual Range (C-H/FP)” QCL Spectra showing new “Dual Range (C-H/FP)” QCL](https://www.photothermal.com/wp-content/uploads/2020/12/Wide-QCL-spectra-2.jpg)
Library (Wiley KnowItAll) search results delivered >95% match
O-PTIR spectra collected in reflection mode. Displayed spectra are raw and unprocessed (<5sec collection time, ~500nm spot size)
O-PTIR spectra are measured off thick polymers (mm’s), vs library FTIR references data off thin films (~10microns)
![mIRage-logo-2017-900px](https://www.photothermal.com/wp-content/uploads/2018/10/mIRage-logo-2017-900px.png)
![mIRage-logo-2017-900px](https://www.photothermal.com/wp-content/uploads/2018/10/mIRage-logo-2017-900px.png)
![image of the mIRage: Combines FTIR and Raman infrared spectroscopy material analysis](https://www.photothermal.com/wp-content/uploads/2018/10/mIRage-product-1000px.png)
![image of the mIRage: Combines FTIR and Raman infrared spectroscopy material analysis](https://www.photothermal.com/wp-content/uploads/2018/10/mIRage-product-1000px.png)
Photothermal Spectroscopy Corp.
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Phone: (805) 845-6568
Email: info [at] photothermal.com