Vibrational Spectroscopic Imaging for Biomedical Applications

Contents......Page 7
Contributors......Page 15
Preface......Page 17
1 Toward Automated Breast Histopathology Using Mid-IR Spectroscopic Imaging......Page 23
1.1 Introduction......Page 24
1.1.1 FT-IR Imaging......Page 26
1.1.2 FT-IR Spectroscopic Characterization of Cells and Tissues......Page 27
1.1.3 FT-IR Imaging for Pathology......Page 28
1.1.5 Modified Bayesian Classification and Automated Tissue Histopathology......Page 30
1.2 Materials and Methods......Page 32
1.2.1 Models for Spectral Recognition and Analysis of Class Data......Page 33
1.2.2 Automated Metric Selection and Classification Protocol Optimization......Page 34
1.2.3 Spectral Metrics and Biochemical Basis......Page 36
1.2.4 Validation and Dependence on Experimental Parameters......Page 37
1.2.5 Application for Cancer Pixel Segmentation......Page 40
1.2.6 Application for Patient Cancer Segmentation......Page 43
1.3 Conclusions......Page 47
References......Page 48
2 Synchrotron-Based FTIR Spectromicroscopy and Imaging of Single Algal Cells and Cartilage......Page 51
2.1 Introduction......Page 52
2.2 IR Environmental Imaging......Page 53
2.2.1 Beamline Design and Implementation......Page 54
2.2.2 Initial Measurements with IRENI......Page 57
2.3 Flow Cell for In Vivo IR Microspectroscopy of Biological Samples......Page 58
2.3.1 Flow Chamber Design......Page 61
2.3.2 Mid-IR and Vis Measurements......Page 64
2.3.3 Viability Tests: PAM Fluorescence Measurements......Page 66
2.3.4 Initial Flow Cell Measurements with IRENI......Page 69
2.4.1 Calcium-Containing Crystals and Arthritis......Page 70
2.4.3 Biologic Models of Calcium- Containing Crystal Formation......Page 71
2.4.4 Synchrotron-Based FTIR Microspectroscopy Spectral Analysis of Calcium-Containing Crystals......Page 72
Acknowledgments......Page 77
References......Page 78
3.1 Introduction......Page 81
3.2.1 Archived Tissue: Paraffin Embedded and Frozen Specimens......Page 83
3.2.2 Preparation of Tissues for Diagnostic Assessment Using FTIR and Raman Microspectroscopy......Page 85
3.2.3 The Effects of Xylene on Fixed Tissue and Deparaffinization of Paraffin-Embedded Tissue......Page 90
3.3.1 Chemical Fixation for FTIR and Raman Imaging......Page 93
3.3.2 Sample Preparation for Biomechanistic Studies......Page 100
3.3.3 Growth Medium and Substrate Effects on Spectroscopic Examination of Cells......Page 102
3.3.4 Preparation of Living Cells for FTIR and Raman Studies......Page 107
3.4 Summary......Page 114
References......Page 116
4.1 Introduction......Page 121
4.2 Theoretical Considerations......Page 122
4.3 Historical Development......Page 124
4.4 Experimental Implementation......Page 129
4.5 Benefits of ATR Microspectroscopic Imaging for Biological Sections......Page 133
4.6 Macro ATR Imaging......Page 139
4.7 ATR Microspectroscopic Raman Imaging......Page 141
References......Page 143
5.1 Introduction......Page 147
5.2 Introduction to Fungi......Page 149
5.2.1 Specimen Preparation......Page 151
5.3 Vibrational Spectroscopy......Page 152
5.3.1 Spectral Resolution......Page 154
5.4 sFTIR Spectra of Fungi......Page 155
5.4.1 Physical Considerations and Spectral Anomalies in sFTIR Spectra......Page 156
5.5 Raman Spectroscopy of Fungi......Page 159
5.5.2 Raman Map of Spore Branch......Page 161
5.5.3 Detection of Crystalline Materials by IR and Raman......Page 162
5.6 SERS Discovery and Development......Page 164
5.6.2 SERS: Applications for Fungi......Page 167
5.7 Conclusions: Lessons Learned, Caveats, Challenges, Promise......Page 172
Acknowledgments......Page 173
References......Page 174
6.1 Introduction......Page 179
6.2.2 Line Mapping......Page 180
6.2.4 Widefield Imaging......Page 181
6.3 Raman Imaging of Cells and Tissues......Page 183
6.4.1 Fluorescence......Page 186
6.4.4 Instrument Response Correction......Page 188
6.4.5 Flatfielding......Page 189
6.4.6 Baseline Correction......Page 190
6.5 Chemometric Analysis of Widefield Raman Images......Page 192
6.5.1 Principal Component Analysis......Page 193
6.5.2 Mahalanobis and Euclidean Distance......Page 195
6.5.3 Spectral Mixture Resolution......Page 200
6.5.4 Derivatives......Page 201
6.6 Chemometrics in the Analysis of Non-Widefield Raman Images......Page 202
6.6.1 PCA......Page 203
6.6.2 Linear Discriminant Analysis......Page 206
References......Page 209
7.1 Introduction......Page 215
7.2 Optical Properties and Resonance Raman Scattering of Carotenoids......Page 218
7.3 Spatially Integrated Resonance Raman Measurements of Macular Pigment......Page 221
7.4 Spatially Resolved Resonance Raman Imaging of Macular Pigment—Methodology and Validation Experiments......Page 227
7.5 Spatially Resolved Resonance Raman Imaging of Macular Pigment in Human Subjects......Page 233
7.6 Raman Detection of Carotenoids in Living Human Skin......Page 237
7.7 Conclusions......Page 243
References......Page 244
8 Raman Microscopy for Biomedical Applications: Toward an Efficient Diagnosis of Tissues, Cells, and Bacteria......Page 247
8.1 Introduction......Page 248
8.2 Raman Imaging of Tissue......Page 249
8.2.1 Mouse Brains......Page 250
8.2.2 Human Brain Tumors......Page 253
8.2.3 Human Colon Tissue......Page 258
8.2.4 Human Lung Tissue......Page 261
8.3 Raman Imaging of Cells......Page 263
8.3.1 Lung Fibroblast Cells......Page 264
8.3.2 Red Blood Cells......Page 267
8.4.1 Species Classification......Page 270
8.4.2 Imaging Single Bacteria......Page 277
8.5 Conclusions......Page 281
References......Page 282
9.1 Introduction......Page 287
9.1.1 History......Page 288
9.1.2 Principles......Page 289
9.2 Instrumentation......Page 290
9.2.1 Laser......Page 291
9.2.3 Filters......Page 292
9.2.5 CCD......Page 293
9.3 Imaging Techniques......Page 294
9.4 Data Analysis: Spectra to Image(s)......Page 298
9.4.1 Classification Techniques......Page 299
9.5.1 Single Cells......Page 300
9.5.2 Tissues......Page 305
9.6 Limitations and Perspectives......Page 313
References......Page 315
10 Vibrational Spectroscopic Imaging of Microscopic Stress Patterns in Biomedical Materials......Page 321
10.1 Introduction......Page 322
10.2 Principles of Raman Spectroscopy......Page 324
10.3.1 Spectral Features......Page 327
10.3.2 PS Behavior......Page 329
10.4.1 Micromechanics of Fracture and Crack-Tip Stress Relaxation Mechanisms......Page 331
10.4.2 Residual Stress Patterns on Ceramic-Bearing Surfaces of Artificial Hip Joints......Page 334
References......Page 337
11.1 From Spontaneous to Coherent Raman Spectroscopy......Page 341
11.2.1 First Generation CARS Microscopes......Page 344
11.2.2 Second Generation CARS Microscopes......Page 345
11.3 CARS Basics......Page 346
11.3.1 Nonlinear Electron Motions......Page 347
11.3.2 Resonant and Nonresonant Contributions......Page 348
11.4.1 Signal Generation in Focus with Pulsed Excitation......Page 349
11.4.3 CARS Chemical Selectivity......Page 350
11.4.4 CARS Sensitivity......Page 352
11.5 CARS and the Multimodal Microscope......Page 354
11.6.1 Focusing in Tissues......Page 355
11.6.2 Backscattering in Tissues......Page 357
11.6.3 Typical Endogenous Tissue Components......Page 358
11.7.1 Ex Vivo Nonlinear Imaging......Page 359
11.7.2 In Vivo Nonlinear Imaging......Page 363
References......Page 364
Index......Page 371
B......Page 373
C......Page 374
F......Page 375
I......Page 376
M......Page 377
P......Page 378
R......Page 379
S......Page 380
Z......Page 381