Infrared Microspectroscopy A training guide for using light microscopy and infrared spectroscopy to analyze materials

Table of Contents Lectures I. Introduction II. Basic Principles of Microspectroscopy III. Transmission Theory IV. Reflection Theory V. Solving Problems with Infrared Microspectroscopy Laboratories I. Microscope Performance Validation a. Performance Validation of the IR-Plan Research Microscope b. Performance Validation of the IR-Plan Analytical Microscope c. Performance Validation of the Nic-Plan Microscope II. Transmission Experiments III. Reflection Experiments

Infrared Microspectroscopy A new technology formed by combining light microscopy and FT-IR spectroscopy Seeing the sample assures analysis of the correct area of interest Understanding microscopy principles is the foundation of infrared microspectroscopy

Microscopy Applications Small samples Plastics Packaging materials Pharmaceuticals Fibers Trace evidence Contaminants Multi-phase mixtures Failure analysis Coatings & inks Electronic materials Migration, diffusion and aging studies Reverse engineering Art conservation And much more

Rules of Microscopy The ruler in the microscopic world is marked in micrometers  m = mm or inches A human hair is 75 to 150  m; the resolving power of the eye is 75 to 100  m

Rules of Microscopy Spatial Resolution is the ability to separate objects into distinct parts Resolution limit or resolving power is the smallest distance two objects must be separated so they are seen as separate parts Detection is the ability to sense the presence of a feature with a reasonable degree of certainty

Infrared Microspectroscopy Observe SampleDefine Area of InterestCollect Spectrum Basic concept is a simple 1, 2, 3 operation 123

Infrared Microspectroscopy Transmission Theory Microscope Training Course

Transmission FT-IR Microscopy Fundamental Optical Factors Affecting FT-IR Microscopy –1. Diffraction –2. Refraction –3. Reflection –4. Scatter

Diffraction * Aperture * Sample Area * Shaded Area Indicates Diffraction

IR Transmission Path Diagram

Spurious-Energy Diagrams Frequency (cm-1) Spurious Energy for Various Width Specimens % of Total Signal 10um 20um 50um Numerical Aperture = 0.50 Single Aperturing

Spurious-Energy Diagrams Frequency (cm-1) Spurious Energy for Various Width Specimens % of Total Signal Numerical Aperture = 0.50 Dual Remote Image Masking 50um 20um 10um

Effects of Stray Light on Absorbance Value No Stray Light 5% Stray Light 10 % Stray Light Measured Absorbance Predicted Absorbance

Hair Sample Using Three Different Aperturing Techniques %Transmittance Wavenumbers (cm-1) Lower Mask Only 2 - Upper Mask Only 3 - Both Upper & Lower Masks

Experiment LAYER 1LAYER um 25 um Cross-Section of a Laminated Film

Outer Layer - Polypropylene Absorbance Wavenumbers 220 um wide layer

Middle Layer - PVA (top) 25 MICRON LAYER #2 - single aperture (bottom) 25 MICRON LAYER #2 - dual apertures Absorbance Wavenumbers

Refraction Normal n1n1 n2n2 n 1 < n 2

Windows and Their Effects Spherical Aberration is produced when the sample is placed on, or between, infrared transparent windows Spectra-Tech objectives and condensers correct for these aberrations, resulting in improved resolution window Spherical aberration is caused by the rays at high incident angles not coming to the same focal point as the low angle rays

Spherical Aberration Correction Images of Multi Layered Polymer Before Reflachromat CorrectionAfter Reflachromat Correction

Ray Trace of Reflachromat Objective

Internal Reflection in Transmission Measurements 123

Nylon 6.6 Fiber, flattened, in air %Transmittance Wavenumbers (cm-1) Interference fringes t(mm) = N/2 x [10/(W h - W l )] t(mm) = 2/2 x (10/540) = mm

Interference in a Thin Film AIR (n =1.0) SAMPLE (n =1.5) A B T  REFLECTANCE = (n-1)2 ( ) = (n+1)2 ( )2 = 0.4

Transmission Spectroscopy The ideal sample for transmission measurement –(1 to 15 um) thickness –large and uniform surface –low reflectivity to avoid thin film interference fringe patterns in the spectra Mounting in a micro compression cell between two infrared windows makes an ideal transmission sample

Micro Compression Cells Standard Micro Compression Cell with standard 13x2 mm Windows Diamond Micro Compression Cell

Micro Compression Cell Sample KBr AB1 B2

Sample Preparation Tools Roller KnifeTungsten Probe Kit

Preparing a Sample in a Micro Compression Cell

Nylon 6.6 Fiber Mounted in a Micro Compression Cell %Transmittance Wavenumbers (cm-1) 4000

Scatter

Sample Preparation onto an Infrared Window Step #1 Step #2 Step #3

Scatter Experiment Photocopier Toner Prepared in a Micro Compression Cell Photocopier Toner % Transmittance Wavenumbers (cm-1)

Scatter Experiment Photocopier Toner (baseline corrected) %Transmittance Wavenumbers (cm-1) Spectrum after Data Manipulation (Baseline Correction)

Transmission Experiments Practical Hands-On Sample Preparation