非接触亚微米分辨红外拉曼同步测量系统—mIRage(材料领域)

非接触亚微米分辨红外拉曼同步测量系统—mIRage(材料领域)

    一款划时代的新型红外光谱系统!


美国PSC (Photothermal Spectroscopy Corp, 前身Anasys公司)最新发布的一款应用广泛的非接触式亚微米分辨红外拉曼同步测量系统。基于PSC独家专利的光热诱导共振(PTIR)技术,mIRage显微红外光谱仪突破了传统红外的光学衍射极限,其空间分辨率高达500 nm,可以帮助科研人员更全面地了解亚微米尺度下样品表面微小区域的化学信息。

O-PTIR (Optical Photothermal Infrared) 光谱是一种快速简单的非接触式光学技术,克服了传统IR衍射的极限。与传统FTIR不同,不依赖于残留的IR 辐射分析,而通过检测由于本征红外吸收引发的样品表面快速的光热膨胀或收缩,来反映微小样品区域的化学信息。


可以快速准确的对半导体微电子器件可能存在的有机缺陷进行化学成分解析, 为优化器件质量提供实验依据

mIRage应用领域:


故障和缺陷分析

聚合物:多层薄膜,相位分布

生命科学:活细胞、组织、骨骼

微塑料:颗粒、纤维


mIRage工作原理:


• 可调的脉冲式中红外激光汇聚于样品表面,并同时发射与红外激光共线性的532 nm的可见探测激光;
• 当IR吸收引发样品材料表面的光热效应,并被可见的探测激光所检测到;
• 反射后的可见探测激光返回探测器,IR信号被提取出来;

• 通过额外地检测样品表面返回的拉曼信号,可以实现同时的拉曼测量。


DisCover IR


DisCover IR+IRman

 




O-PTIR克服了传统红外光谱的诸多不足: 

• 空间分辨率受限于红外光光波长,只有10-20 µm
• 透射模式需要复杂的样品准备过程,且只限于薄片样品

• 无传统ATR模式下的散射像差和接触污染

 

O-PTIR的优势之处在于: 

• 亚微米空间分辨的IR光谱和成像(~500 nm),且不依赖于IR波长
• 与透射模式相媲美的反射模式下的图谱效果
• 非接触测量模式——使用简单快捷,无交叉污染风险
• 很少或无需样品制备过程 (无需薄片), 可测试厚样品
• 可透射模式下观察液体样品

• 实现同时同地相同分辨率的IR和Raman测试,无荧光风险


mIRage 技术参数: 


波谱范围模式探针激光样品台最小步长样品台X-Y移动范围
IR (1850-800 cm-1)反射532 nm+785 nm100 nm110*75 mm
IR (3600-2700 cm-1)透射
Raman (3900-200 cm-1)反射

Customizable to 3000-2800 cm-1 + 1800-1000cm-1

Customizable to 2250-1950 cm-1 + 1800-1000cm-1


Applications

The advancement of analytical methods and systems are paving the way for innovation in academic, industrial and government sectors. As a result of the fast-paced progress, more research questions are asked that surpass the limits of these technologies. The mIRage IR Microscope is now answering these questions using Optical Photothermal IR (O-PTIR) spectroscopy, as well as IR+Raman – simultaneous submicron IR and Raman microscopy. With more chemical characterization requirements being met, mIRage is ushering in the next generation of applications and technology developments in numerous industries.

污染

故障/缺陷分析

聚合物

相位分布

生命科学

活细胞、组织、病毒

微塑料

颗粒、纤维

Contamination

Successful identification of contamination is a critical step in ensuring product or process quality is maintained. With stricter control standards and the decreasing size of high-tech products, confidently identifying smaller features is becoming increasingly important. With submicron spatial resolution using a non-contact reflection mode, O-PTIR easily resolves the most challenging of contamination issues.


Failure analysis of high technology components

Data illustration showing failure analysis of high technology components

Left; visible image showing location of 6 µm defect, Upper Right; Comparison of unknown O-PTIR spectrum to nearest library match, Lower Right: Comparison of unknown Raman spectrum to nearest library match


Data illustration with failure analysis of high technology components

Upper Left; Schematic representation of sample and measurement, Lower Left; Visible camera image of defect, Right; O-PTIR spectra from on and off the defect. Colors correspond to markers on visible image



Submicron line scan of polystyrene beads embedded in epoxy

Data images showing submicron line scan of polystyrene beads embedded in epoxy
Left: Demonstration of ~400nm spatial resolution as determined from a line scan (at 100nm steps) across 1 µm diameter polystyrene beads embedded in epoxy and sectioned to ~300nm thick. Right: A sharp boundary of only ~400nm is observed on both sides of the polystyrene beads as the IR spectral features transition between the two components.

Film defect identification

Data image showing film defect material identification

Left: Optical image of defect in a 240 µm thick two layer film. Markers on image represent the location of subsequent O-PTIR spectral collection. Right: Spectra collected in the defect-free (red) and defect (blue) region of the sample. The spectra display peaks indicative of isotactic polypropylene (998 cm-1). Insert: In the plot of the varying intensities for the isotactic polypropylene peak, both on the defect and off, The film region shows consistent signal intensity, while the defect region shows significant variability.









Polymers

Polymers are present in virtually all products we interact with daily. With increasing environmental awareness, polymer science is looking at more novel and complex solutions to improve functionality and reduce environmental impact. These requirements often exceed the limits of traditional IR microscopy, especially when it comes to spatial resolution. The mIRage IR Microscope, with its unique submicron spatial resolution using a non-contact reflection mode technique, is able to meet even the most demanding of analytical and sample characterization needs.



Submicron IR+Raman Microplastics

O-PTIR image and spectra of polystyrene and Polymethylmethacrylate dispersed in saline solution.

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.



O-PTIR – polymer (PLA-ACM) phase dispersions

Data illustrating O-PTIR – polymer (PLA-ACM) phase dispersions

High quality spectra were collected in seconds, with high spatial resolution images collected in minutes. Image on the right shows image resolution of an inclusion of ACM as small as 249nm! 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


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!

Data image of polymer laminates analysis with O-PTIR

IR spectroscopy for direct fiber characterization


Data image showing the use of mIRage for direct fiber characterization

O-PTIR spectra of PP-based nanofibers with 800 nm diameter




Imaging and spectroscopy of bioplastic laminates


Composite (red/green) single frequency images
Image showing mIRage imaging and spectra 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




Little to no sample preparation of a multilayer film

Image showing a sliced piece of polymer multilayer film that required little to no sample preparation
Left: A multilayer packaging film block face sample with manually selected markers for subsequent O-PTIR spectra collection. Right: The spectra easily show difference in composition of each layer.

Submicron spatial resolution between film layers

Image demonstrating the submicron spatial resolution between film layers
Left: An optical image of a food multilayer film sample. Right: Corresponding O-PTIR spectra spaced 500 nm apart, with clear spectral distinction.


Life science

From plant biology to medical research, life science is an ever expanding research field that has impact in numerous industries. Providing submicron spatially resolved chemical analysis on biological samples, in a label free and objective approach, has proven itself to be a difficult result to obtain. The mIRage IR Microscope has accomplished this with its non-contact reflection mode O-PTIR technique, and is unlocking numerous applications capabilities.


Single bacterial cell O-PTIR microscopy with deuterium labelled E. coli

Single bacterial cell O-PTIR microscopy with deuterium labelled E. coli

A: O-PTIR image at 1655cm-1 (protein) at 200nm step size. B: O-PTIR image at 2195cm-1 (C-D stretch) at 200nm step size. Both images took 3 min to acquire each. C: Single E. Coli cell (2.6×1.3 microns) imaged at 1655cm-1 with 50nm steps. Image time, ~1 min. D: Four submicron (~500nm spot) O-PTIR spectra were acquired from the single bacterial cell image above (Upper Right), with corresponding colors. Spectra are normalized to 1655cm-1. Intracellular differences are apparent with the Amide I band position and shape indicating intracellular chemical (protein secondary structural) differences being detected. Each spectrum is 10 averages (~15 secs). You can see the C-D absorbances at around 2195cm-1 and 2100cm-1.

Single bacterial cell simultaneous submicron IR+Raman microscopy

Single bacterial cell simultaneous submicron IR+Raman microscopy

A: Visible image of bacterial cells. Orange box indicate region of IR imaging. B: O-PTIR infrared image at 1655cm-1, with 50nm step size. Collection time ~1 min. C: Simultaneous, submicron IR and Raman spectra collected from the indicated spot on the single bacterial cell. Spectra are normalized to the most intense band spectra are ~20sec accumulations. O-PTIR spectra are collected with a Dual Range (C-H/FP) QCL, covering 3000-2700, 1800-950cm-1 in a single unit. O-PTIR spectra are raw (no processing). Raman spectra are baseline corrected.
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.

Targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes

Lipid relative to protein
2856 (CH2)/ 1658 (Protein)
Lipid chain length image
2856 (CH2)/2874 (CH3)
Data illustration showing targeted imaging mode (chemically specific imaging) Intra-cellular imaging, off glass slide, at 100nm step sizes

A: Lipid Chain length image (2856cm-1 (CH2)/ 2874cm-1 (CH3). B: : Lipid relative to protein image (2856cm-1) (CH2)/ 1658cm-1). Both IR images collected at 100nm pixel size. ~5 mins per image. D: O-PTIR Spectra from markers in images (spectra are single scans, ~1sec measurement time, no processing. C: 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)

IR Polarized O-PTIR to study collagen orientation in individual fibrils and tendon

Data images showing 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

Breast tissue calcification – Demonstration of<1 micron spatial resolution with O-PTIR

Data images of breast tissue calcification, demonstration of sub 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)

Submicron amyloid aggregate
imaging in neurons

O-PTIR image, 1630/1656                       O-PTIR spectra

Data image of 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

Single mammalian cell analysis – submicron O-PTIR off glass slide with no dispersive scatter artefacts
Data image showing single mammalian cell analysis – submicron O-PTIR off glass slide with no dispersive scatter artefacts

Left; Optical image of a cells deposited on regular glass slide. Markers show replicate spectra locations per cell.
Right; Average spectrum per three different cell lines (normal and two cancerous). Shaded area are 1 standard deviation from replicate spectra. Spectra are collected in reflection mode off regular glass slides. Other than averaging (per cell line) and area normalization to the amide I and II bands, no other pre-processing (eg baseline corrections etc) were performed. Variability in the glass spectral region from 1300-900cm-1) are due to cell thickness differences.
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)

Submicron O-PTIR imaging of live cells in water

Image of data showing submicron O-PTIR imaging of live cells in water
Left: Optical image of hydrated epithelial cheek cells in water. Middle: Key macromolecules are easily spectrally discerned and spatially isolated, with the lipid inclusion as small as 0.5-1 µm being easily resolved. Spectra are not corrected for water and therefore inclusive of water absorbances. Images were collected at 500 nm step size. Right: The measurements
were collected using a 0.5 µm step size in transmission mode.

IR+Raman analysis of red blood cells

Data illustration showing the results from IR+Raman analysis of red blood cells

Left: Optical image with selected 70 x 70 µm area for subsequent Raman imaging. Middle: Subsequent Raman image at 1583 cm-1. Right: IR+Raman spectra collected off of a selected red blood cell (~500 nm resolution).


Microplastics contamination in oceans and waterways

An emerging environmental concern especially for ocean/aquatic life is microplastics pollution. With research continuing to increase to assess the potential harmful affects to aquatic life and the food chain, accurate characterization and identification of microplastics is becoming critical. Microplastics are small plastic microscopic particles and fibers that range from 1micron to 5mm. Microplastics have been found in virtually all environments, from waterways, to wastewater, to the air we breathe and the food we eat. Owing to their small sizes (1 micron to 5mm) their accurate identification and morphological characterization can be challenging. O-PTIR, with its ability to measure submicron particles and fibers of varied sizes in far field reflection mode to yield FTIR transmission-like spectral quality regardless of particle shape and size, makes it the ideal technique for microplastics/particulate characterization. When coupled with simultaneous Raman (IR+Raman), it becomes an even more powerful tool that brings together the best of IR and Raman into a single measurement, for more thorough and accurate characterization for microplastics contaminants.



O-PTIR image and spectra of PS and PMMA dispersed in saline

O-PTIR image and spectra of polystyrene and Polymethylmethacrylate dispersed in saline solution.

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.


1、多层薄膜


 

高光谱成像: 1 sec/spectra. 1 scan/spectra
样品区域尺寸:20 µm x 85 µm size. 1 µm spacing. 
图谱中可以明显看出在不同区域上的羰基,氨基以及CH2 拉伸振动的分布

很少或无需样品制备的多层高分子膜的O-PTIR分析


高分子薄膜层间的亚微米空间分辨O-PTIR分析



2、高分子



 

高分子膜缺陷。左:尺寸为240 µm的两层薄层上缺陷的光学图像;

右:在无缺陷处(红色)和缺陷处(蓝色)的样品的IR谱图,998 cm-1处为of isotactic polypropylene 的特征红外吸收峰

环氧树脂包埋聚苯乙烯球的亚微米分辨O-PTIR线扫描



PS和PMMA微塑料混合物的亚微米红外拉曼同步O-PTIR

光谱和成像分析




3、生命科学 


 

左:70*70 µm范围的血红细胞的光学照片;中:红色条框区域在1583cm-1处的Raman照片;右:红血细胞选择区域的同步的IR和Raman图谱



矿物质的红外成像:小鼠骨骼中的蛋白质分布分析

 

上左:水中上皮细胞的光学照片;
上右:目标分子能够在红外光谱上很容易的区分和空间分离,可以明显看到0.5-1.0 µm的脂肪包体;
下:原理示意图:红外光谱测量使用透射模式,步长为0.5 µm

PLA/PHBHx生物塑料薄片的O-PTIR光谱和成像分析

 


4、医药领域

 

 

左:PLGA高分子和Dexamethasone药物分子的混合物表面的光学照片
中:在1760 cm-1 出的高光谱图像,显示了 PLGA在混合物中的分布,图像尺寸40 µm * 40 µm 

右:在1666 cm-1 出的高光谱图像,显示了 Dexamethasone在混合物中的分布,图像尺寸40 µm *40 µm

 

 

5、法医鉴定

 

 

左:800 nm纤维的光学照片

右:纳米纤维不同区域的O-PTIR图谱

 

6、其他领域

 

•  故障分析和缺陷

•  微电子污染

•  食品加工

•  地质学 

•  考古和文物鉴定


[1] Depth-resolved mid-infrared photothermal imaging of living cells and organisms with submicrometer spatial resolution, Ji-Xin Cheng et al., Sci. Adv. 2016, 2, e1600521.

[2] Mid-Infrared Photothermal Imaging of Active Pharmaceutical Ingredients at Submicrometer Spatial Resolution, Ji-Xin Cheng et al., Anal. Chem. 2017, 89, 4863-4867.

[3] Label-Free Super-Resolution Microscopy. Springer, Biological and Medical Physics, Biomedical Engineering.

[4] Advances in Infrared Microspectroscopy and Mapping Molecular Chemical Composition at Submicrometer Spatial Resolution, Spectroscopy 2018.

[5] Evolution of a Radical-Triggered Polymerizing High Internal Phase Emulsion into an Open-Cellular Monolith, Macromolecular Chemistry and Physics, 2019.

[6] A Global Perspective on Microplastics, Journal of Geophysical Research: Ocean, 2019.

[7] Super-Resolution Infrared Imaging of Polymorphic Amyloid Aggregates Directly in Neurons (Front Cover), Advanced Science, 2020.

[8] Self-formed 2D/3D Heterostructure on the Edge of 2D Ruddlesden-Popper Hybrid Perovskites Responsible for Intriguing Optoelectronic Properties and Higher Cell

Efficiency, Applied Physics, 2020.

[9] Two-Dimensional Correlation Analysis of Highly Spatially Resolved Simultaneous IR and Raman Spectral Imaging of Bioplastics Composite Using Optical Photothermal Infrared and Raman Spectroscopy, The Journal of Molecular Structure, 2020.

[10] Super resolution correlative far-field submicron simultaneous IR and Raman microscopy: a new paradigm in vibrational spectroscopy, Advanced Chemical Microscopy for Life Science and Translational Medicine, 2020.

[11] Submicron-resolution polymer orientation mapping by optical photothermal infrared spectroscopy, International Journal of Polymer Analysis and Characterization, 2020.

[12] Bulk to nanometre-scale infrared spectroscopy of pharmaceutical dry powder aerosols, Analytical Chemistry, 2020.

[13] Optical Photothermal Infrared Micro-Spectroscopy – A New Non-Contact Failure Analysis Technique for Identification of<10mm Organic Contamination in the Hard drive and other Electronics Industries. Microscopy Today, 2020.

[14] Spontaneous Formation of 2D-3D Heterostructures on the edges of 2D RuddlesdenPopper Hybrid Perovskite Crystals, Chemistry of Materials, 2020.

[15] Simultaneous Optical Photothermal Infrared (OPTIR) and Raman Spectroscopy of Submicrometer Atmospheric Particles, Analytical Chemistry, 2020.

[16] Detection of high explosive materials within fingerprints by means of optical-photothermal infrared spectromicroscopy, Analytical Chemistry, 2020.

[17] Polarized O-PTIR of collagen and individual fibril strands reveals orientation, Molecules Special Edition: “Biomedical Raman and Infrared Spectroscopy: Recent Advancement and Applications, 2020.



科学研究


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生物医学应用




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mIRage Demo演示-Microplastics

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