Instrumentation
Laser Raman Molecular Microprobe (MOLE) P. Dhamelincourt and F. Wallart
M. Leclercq and A. T. N'Guyen
D. O. Landon
Laboratoire de Spectrochemie Infrarouge et Raman, C.N.R.S., Université de Lille, B.P. 36, 59650 Villeneuve d'Ascq, France
I.S.A. Nord 35 rue Inkermann 59000 Lille, France
I.S.A. Inc. 173 Essex Avenue Metuchen, N.J. 08840
This new microanalysis technique for detecting, identi fying, and obtaining the distribution of the different com ponents of an heterogeneous sample is similar to electron or ion microprobe methods. However, an important advan tage is the ability to study samples in air, under a con trolled atmosphere, or even inside transparent media rath er than just in a vacuum
Most microanalysis physical methods are based on atomic properties and only indirectly provide information on how atoms link together to form polyatomic structures. This is the case for electron or ion microprobe methods, which are based on the analysis of electron X-rays or secondary ions emitted by samples under electron or ion beam impact. These methods often fail to yield information on chemical bonds and conformation of molecules. In the laser molecular microprobe (1, 2), photons generated by a laser are used to excite the sample and cause the emission of Raman lines of different components. By use of these Raman lines, each component can be detected, identified, and then located by forming a micrographie image that gives the " m a p " of its distribution in the sample. This is the principle of the nondestructive microanalytical method described here.
Instrument Description T h e basic instrument (3-6) combines a conventional optical micro-
scope (with bright and dark field illumination), an optical filter possessing a very low stray light level, and a multichannel and/or monochannel detection system. This configuration gives a very versatile instrument because it permits different modes of operation without disturbing the sample. A block diagram of the instrument is shown in Figure 1. A laser (argon, krypton, dye laser, etc.) is the monochromatic source of light used for irradiating the sample. A conventional optical microscope provides an illuminatory stage and forms part of the imaging system. T h e sample under study is placed on the microscope stage. All modes of observation available in a standard optical microscope are available in the instrument; therefore, the samples can be studied in air, liquid, or transparent media. A double monochromator spectrometer with two concave aberration corrected gratings is the central part of the instrument. This device permits spectral separation of the chosen spectral line, which corresponds to a particular vibrational mode, and it trans-
414 A · ANALYTICAL CHEMISTRY, VOL. 51, NO. 3, MARCH 1979
mits good images in a wide spectral range without changing components. T h e detection system is composed of two separate assemblies. One, which has a photomultiplier, records the spectrum on a chart recorder. T h e other has a three-stage image intensifier, a TV-type detector, and a visualization system t h a t displays micrographic images. A computer and disc or tape memory can be attached to the system for information processing and storage. Figure 2 gives the optical scheme of the instrument.
Modes of Operation T h e two modes of operation currently used depend on the type of irradiation of the sample. Punctual Illumination with Monochannel or Multichannel D e t e c tion. By use of the bright field illumination system of the microscope, the same objective (50X, ΝΑ, 0.85-100Χ, N/A, 0.90) focuses the laser beam into a spot on the component of the sample to be identified and collects the scat tered light. In this case, the top aper ture of the objective, the grating sur faces, the sample, and entrance slit are simultaneously optically aligned by the transfer optic. To obtain the Raman spectrum, the optical filter can function as a Raman spectrometer when the detector is a photomultiplier followed by an amplifier and a chart recorder, or as a Raman spectrograph when the detector is an intensifier phototube, followed by a low-level TV camera (SIT or SEC tube). Global Illumination and Imaging System. Selecting, in the Raman spec trum, radiation characterizing one particular component forms a micro0003-2700/79/0351-414A$01.00/0 © 1979 American Chemical Society
graphic image that gives the distribu tion of this component. The aperture of the microscope objective (O) (Fig ure 2) is optically aligned with the three slits (Oi, 0 2 , 0 3 ) of the optical filter. The image (Si) of the sample (S) given by the objective is formed on the gratings (S2, S3) and goes through the aperture of the exit slit (O3) to the photocathode of the inten sifier tube (S4). A rotating laser beam feeding the objective annular illumi nator (dark field illuminator) homoge neously illuminates the sample and suppresses the "speckle noise" of the images. The spatial resolution of the images is about 1 μηι. The Spectral Mode. In the monochannel spectral mode classical Raman spectra can be recorded of the entire sample or of a specific portion selected to identify positively the na ture of that portion. In the multichannel spectral mode the Raman spectra can be observed in spectrographic form on an oscillo scope in real time in up to 100-cm -1 segments. This mode is particularly useful for monitoring the evolution of a sample. The Imaging Mode. To locate one of the components of the sample, an image of the surface obtained from a characteristic line of this component can be directly observed on the TV monitor screen. This allows "map ping" the distribution of a given sub stance in a heterogeneous sample.
/
Image Intensifier and TV Camera
Double Spectrometer with Concave Holographic Gratings Mode Commutation Control
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Monochannel Detection Mode Control Module
^ O .•s"
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Display Module
System
Micrographie Image
Spectrometer Control Input Optical System Microscope
To Computer
Multichannel Detection Mode Control Module
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Laser
Figure 1 . Block diagram of laser Raman microprobe
Double Concave Holographic Grating Filter
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,* Semitransparent Laser l\ . p!ate Beam * 7 r i r * / ^ c tli \ y Laser jTJ' 1 1 I 11· Roam 7N ' '
S4 - . Image Intensifier Phototube
Photomultiplier
SIT or SEC Camera
Some Applications of the Laser Raman Microprobe To show the many possibilities opened by the laser Raman microprobe, we now present some examples of applications of the instrument. Geochemistry and Geophysics. The first example is a fragment of a natural stone of celestine (collection of the Faculty of Science of Lille, France). The stone has been cut and roughly polished to make it approxi mately plane. By examination under an optical microscope (dark field illu mination by reflected white light), an area comprising crystal inclusion is se lected. In the monochannel spectral mode, using punctual illumination (spot di ameter: 2 μπι), we obtain a character istic Raman spectrum for each of the. two areas of interest (Figures 3 and 4). In the multichannel spectral mode, using global illumination, we isolate the 473-cm -1 wavelength characteris tic of sulfur, and the image is observe/1 at this wavelength in the imaging mode (Figure 5). Similarly, the 1000cm _ 1 wavelength characteristic of SrSC>4 is isolated, and the image is ob served at this wavelength (Figure 6).
Monochannel Detection (Photomultiplier)
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Multichannel
- Sample -
Punctual Illumination
Global Illumination
Illumination
Amplitude
θ
TV Oscilloscope Monitor Multichannel Detector v
_/U_^_ Chart Recorder Monochannel Detector
Detection Figure 2. Optical scheme of instrument
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Figure 3. Raman spectrum of Celestine crystal inclusion: sulfur ANALYTICAL CHEMISTRY, VOL. 5 1 , NO. 3, MARCH 1979 · 415 A
1000 450
100
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r4i 700
1000
1100 AScrrr 1
' Figure 4. Raman spectrum of area around same inclusion: Strontium sulfate
5 0 μΠΊ Figure 5. Raman image of sulfur inclusion obtained with 4 7 3 - c m _ 1 line 416 A · ANALYTICAL CHEMISTRY, VOL. 51, NO. 3, MARCH 1979
When the image at the characteris tic line of SrSC>4 is observed, the sul fur is no longer obvious but its trace is still visible. T h e sulfur crystal in the plane of the surface is thin and there fore transparent to the radiation emit ted by S r S 0 4 . Further impurities at the surface appear as dark spots on the micrographie image. Fluid inclu sions can also be directly analyzed (7) provided that the matrix is sufficient ly transparent to the laser beam and scattered light. Archeology and Gemnology. Knowing the exact nature of a stone object without taking a sample (even a microscopic one) would be greatly beneficial in any museum of art and archeology. With the laser Raman mi croprobe, it is possible to identify di rectly the nature of the stone. Examining the precise location of a small area of the stone can prevent contact with fluorescent impurities, which are often present in natural ma terials and which, in most cases, pre vent the recording of Raman spectra with a classical Raman spectrometer. Figure 7 shows the comparison be tween Raman spectra obtained from
50 μϊΥ\ Figure 6. Raman image of same area obtained with 1 0 0 0 - c m _ 1 characteristic line of sulfate
a precious Chinese vase and from a sample of serpentine (8). Another important application is the determination of authenticity and geographical origins of gems by identifying in situ the solid inclusions present in precious stones. Investigation of Defects in Industrial Materials. The formation of bubble-like inclusions is a problem currently encountered in the commercial production of glass. The in situ analysis of bubble content aids in properly selecting materials and process parameters which will inhibit their formation. Figure 8 represents one example of the investigation of solid deposits on the wall of a bubble inside a sample of industrial glass (in collaboration with the Inorganic Chemistry Laboratory E.N.S.C. Strasbourg, France). Spectra a, b, and c represent sulfur, polysulfide, and sulfate of sodium, respectively. Each deposit can be imaged by use of a characteristic Raman line. Pollution Analysis. This new nondestructive analytical tool is also perfectly suited to the study and identification of urban and industrial dusts. Figure 9 shows the results obtained from dust collected on a highway. The two broad maxima near 1400 and 1600 c m - 1 (Figure 9c) are often observed
Figure 7. Raman spectra obtained from a) precious Chinese vase, b) specimen of serpentine
on the spectra of air particulate dust. They can be attributed to degraded organic compounds or soot coating (9). Industrial Quality Control. In the industrial field, it is very important to control the quality of manufactured products. Local defects in synthetic films or fibers impair their clarity and can cause rending. The compounds responsible for these defects can be identified with MOLE (molecular optics laser examiner) by simply recording in situ the Raman spectra of the
defects. Generally, the defects are caused by atmospheric dusts, local differences of crystallinity, degraded areas, or local concentration of copolymers. These studies are useful for quality control of semiconductors and integrated circuits. Figure 10 shows the spectra of different parts of an integrated circuit and the pollutants discovered on its surface. The major pollutant found was lead acetate mixed with organic compounds.
Figure 8. Identification of solid deposits contained in a bubble inside a commercial glass Different areas appear on wall of bubble when it is observed through a microscope. By use of punctual illumination a characteristic Raman spectrum can be recorded from each of these areas. Raman spectrum characteristic of a) sulfur, b) sodium polysulfide, c) sodium sulfate
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Figure 9. Raman spectra of some particles ( 2 - 1 0 μ in size) removed from surface of a collection filter placed near a highway Raman spectrum of a) quartz particle ((V-Si02), b) calcite particle (CaC03), c) Dolomite particle [CaMg(C03)2], d) Feldspar particle (Orthoclase)
Figure 11. Micrographie image (X300) of part of an histologi cal section of Blatella germanica L
Figure 10. Analysis of pollutants found at surface of a com mercial integrated circuit 420 A · ANALYTICAL CHEMISTRY, VOL. 51, NO. 3, MARCH 1979
from reference materials (uric acid, potassium urate) and fat body sphero crystals. A comparison between these spectra shows that fat body is mainly composed of uric acid with a small amount of potassium urate. Conclusions
Figure 12. Raman spectra obtained from a) commercial sample of uric acid (parti cle ~ 5 μ in size), b) fat body spherocrystal (~5 μ in diameter), c) commercial sam ple of potassium urate (particle ~ 5 μ in size)
Biology. Important information unattainable by electron or ion microprobe can be obtained on biological samples. For example, Figure 11 shows spherocrystals (10) (1-12 μ) of fat body of Blatella germanica L. (Insecta Dictyoptera). Fat body can be roughly considered as the liver of in vertebrates, and puric catabolism is
important. T h e samples are histologi cal sections of fixed and paraffin-em bedded material. Traditional methods of study have given the following re sults (10, 11): biochemistry, uric acid; X-ray, amorphous structure; and Castaing microprobe, abundant Κ and small quantities of Ca and P. Figure 12 represents the spectra obtained
T h e purpose of the results present ed here is simply to show the wide spread interest in this new microanalytical technique. Several other fields will be investi gated, and studies are already in prog ress (i.e., corrosion, and photochemi cal and electrochemical reactions). We expect the laser Raman microprobe to find application in laboratories using such microprobe techniques as Auger (AES), low-energy electron dif fraction (LEED), X-ray photoelectron spectroscopy (ESCA), secondary ion mass spectrometry (SIMS), and elec tron microscopy. T h e high-energy in tensity required in some of these tech niques and sample treatment prevent their use on delicate samples. The Raman microprobe probably provides the only approach to surface analysis on living materials. In addition, the instrument performs nondestructive analysis in various media (air, con trolled atmosphere or liquid) for small quantities of sample down to a few picograms. The versatility of this new instru ment should greatly facilitate micro analysis and structural determination, and complement other techniques. References
From left to right: P. Dhamelincourt has played a major role in the conception and realization of the molecular microprobe. He specializes in micro Raman spectroscopy. M. Leclercq was in charge of the Mole Laboratory at Lirinord Division Instrument S.A. He is now technical manager at D.I.L.O.R. (rue des Bois Blancs à Lille, France). A. T. N'Guyen worked in the Mole Laboratory at Lirinord Division Instrument S.A. He is now in charge of the Application Laboratory at D.I.L.O.R. F. Wallart is assistant professor at the University of Lille. He specializes in Raman spectroscopy and instrumentation. D. O. London (not shown) is manager of research and development at Instruments S.A., Metuchen, N.J. His interests include optical spectroscopy and instrument design.
( 1 ) M. Delhaye and P. Dhamelincourt, J. Raman Spectrosc, 3, 33 (1975). (2) G. J. Rosasco, E. S. Etz, and W. A. Cas satt, Appl. Spectrosc, 29, 396 (1975); in Proc. of the Fifth Int. Conf. on Raman Spectroscopy, E. 0 . Schmid et al., Freiburg, Germany, Sept. 2-8, 1976. (3) M. Delhaye, P. Dhamelincourt, and Y. Moschetto, U.S. P a t e n t 4,030,827 (1977). (4) M. Delhaye, P. Dhamelincourt, and E. Da Silva, French Patent ANVAR 762 1539(1976). (5) P. Dhamelincourt and P. Bisson, Microsc. Acta, 79, 267 (1977). (6) P. Dhamelincourt, "Lasers in Chemis try," Elsevier, Amsterdam, T h e Nether lands, 1977. (7) G. J. Rosasco, E. Roedder, and J. H. Simmons, Science, 190, 557 (1975). (8) P. Dhamelincourt and H. J. Schubnel, Rev. Gemnol., 52, 11 (1977). (9) E. S. Etz and G. J. Rosasco, "Environ mental Analysis," Academic Press, New York, N.Y., 1977. (10) C. Ballan-Du Français, doctoral thesis, Paris, France, 1975. (11) C. Ballan-Du Français, Cellule, 70, 317(1974).
This study financially supported by C.N.R.S., University of Lille, the Direction Générale de la Recherche Scientifique, and the Agence Nationale pour la valorisation de la Recherche of France.
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