The textbook “Analytical Techniques in Life Sciences: A Textbook for UG and PG Students in Indian Universities” is designed to provide a comprehensive and accessible guide to the essential analytical methods used in modern biological, chemical, and physical sciences. As scientific research grows increasingly interdisciplinary, a firm understanding of analytical techniques becomes vital for students and researchers alike. This book aims to bridge the gap between theory and practical application, catering to undergraduate and postgraduate students across Indian universities.
The content begins with the fundamental principles of microscopy, exploring traditional light microscopy as well as advanced techniques like phase contrast, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). These techniques are crucial for the visualisation and structural analysis of biological and material samples at both surface and internal levels.
The chapter on biochemical buffers and pH measurement emphasises the role of buffer systems in maintaining pH stability, especially in enzymology and metabolic studies. Detailed explanations of common buffers and the use of pH meters provide students with foundational knowledge relevant to various life science experiments.
Centrifugation, a pivotal technique in cell and molecular biology, is explained with clarity ranging from basic principles to advanced ultracentrifugation used for subcellular fractionation and macromolecular analysis.
The book presents chromatography as a central tool for separating and identifying biomolecules. It covers techniques such as thin-layer chromatography (TLC), gas-liquid chromatography (GLC), and high-performance liquid chromatography (HPLC), illustrating their applications in pharmaceuticals, environmental analysis, and biochemical research.
Electrophoresis is introduced as a powerful method for analysing nucleic acids and proteins. The book details the methodologies and applications of both agarose and polyacrylamide gel electrophoresis, including DNA fingerprinting and RNA quantification.
The section on spectroscopy offers thorough insights into techniques such as UV-Visible spectrophotometry and fluorescence spectroscopy, as well as advanced methods like mass spectrometry (MS) and nuclear magnetic resonance (NMR), which are critical in molecular characterisation and structural elucidation.
Further, the book discusses advanced analytical tools such as Fourier Transform Infrared (FTIR) spectroscopy, atomic absorption spectroscopy (AAS), X-ray diffraction (XRD), and flow cytometry, highlighting their importance in biomedical, pharmaceutical, and environmental studies.
Throughout the book, theoretical principles are integrated with real-world applications, laboratory relevance, and instrumental insights. Each chapter has been carefully crafted to meet the curriculum requirements of Indian universities while also encouraging deeper exploration and hands-on learning.
Contents –
1. ANALYTICAL TECHNIQUES
1.1 Principles and Applications of Microscopy
1.1.1 Types of Microscopies
1.1.2 Types of Optical Microscopes
1.1.3 Electron Microscopy
1.1.4 Importance of Microscopy in Science
1.1.5 Microscopy: A Key Tool in Biological Research
1.1.6 Measurement Units in Microscopy
1.2 Light Microscopy
1.2.1 Principles of Light Microscopy
1.2.2 Types of Light Microscopes
1.2.3 Applications for Light Microscopy
1.2.4 What is a Simple Microscope?
1.2.5 Light Microscopy: Principle and Key Components
1.2.6 Sample Preparation for Microscopy
1.2.7 Variations to Bright Field (Transmission) Microscopy
1.3 Phase Contrast Microscopy
1.3.1 Working of Phase-Contrast Microscopy
1.3.2 Parts of a Phase-Contrast Microscope
1.3.3 Applications, Advantages, and Limitations of Phase-Contrast Microscopy
1.3.4 Advantages of Phase-Contrast Microscopy
1.3.5 Working Mechanism of Electron Microscopy
1.4 Scanning Electron Microscopy (SEM)
1.4.1 Definition of Scanning Electron Microscope (SEM)
1.4.2 Principle of Scanning Electron Microscope (SEM)
1.4.3 Working Principle of Scanning Electron Microscope (SEM)
1.4.4 Electron Source and Beam Generation
1.4.5 Electron-Specimen Interaction and Image Formation
1.4.6 Scanning Process and Image Production
1.4.7 Contrast and Image Clarity in Scanning Electron Microscopy
1.4.8 Parts of a Scanning Electron Microscope (SEM)
1.4.9 Applications of the Scanning Electron Microscope (SEM)
1.4.10 Advantages of the Scanning Electron Microscope (SEM)
1.5 Transmission Electron Microscope (TEM)
1.5.1 Historical Background
1.5.2 Parts of the Transmission Electron Microscope (TEM)
1.5.3 How a Transmission Electron Microscope (TEM) Works
1.5.4 Preparation of Specimen for Visualisation by TEM
1.5.5 Freeze-Etching Treatment
1.5.6 Applications of Transmission Electron Microscope (TEM)
1.5.7 Advantages of Transmission Electron Microscope (TEM)
1.5.8 Limitations of Transmission Electron Microscope (TEM)
1.6 Biochemical Buffers and pH Measurement
1.6.1 pH Measurement
1.6.2 Preparing Buffers Correctly
1.6.3 Guidelines for Buffer Preparation
1.7 pH Measurement
2. BASIC PRINCIPLES OF SEDIMENTATION
2.1 Sedimentation Process Through an Experiment
2.1.1 Applications of Sedimentation
2.1.2 Types of Sedimentation
2.1.3 Types of Sedimentation Tanks
2.1.4 What is Water?
2.1.5 Uses of Sedimentation
2.2 Centrifugation
2.2.1 Principle of Centrifugation
2.2.2 Types of Centrifugations
2.2.3 Principle of Analytical Centrifugation
2.2.4 Steps of Analytical Centrifugation
2.2.5 Uses of Analytical Centrifugation
2.2.6 Density Gradient Centrifugation
2.2.7 Steps of Density Gradient Centrifugation
2.2.8 Uses of Density Gradient Centrifugation
2.2.9 Differential Centrifugation
2.2.10 Steps and Uses of Differential Centrifugation
2.2.11 Principles and Techniques of Centrifugation
2.2.12 Isopycnic Centrifugation
2.2.13 Rate-Zonal Density Gradient Centrifugation (Moving Zone Centrifugation)
2.2.14 Differential Velocity (Moving Boundary) Centrifugation Principle
2.2.15 Equilibrium Density Gradient Centrifugation
2.2.16 Sucrose Gradient Centrifugation
2.3 Ultracentrifugation
2.3.1 Applications of Ultracentrifugation
2.3.2 Principle of Ultracentrifuge
2.3.3 Types of Ultracentrifuges
2.3.4 Parts of Ultracentrifuge
2.3.5 Types of Optical Systems
2.3.6 Procedure/Steps of Operating an Ultracentrifuge
2.3.7 Procedure for Preparative Ultracentrifuge
2.3.8 Precautions for Operating Ultracentrifuge
3. CHROMATOGRAPHY
3.1 History of Chromatography
3.1.1 Key Milestones in Chromatography
3.1.2 The Role of Chromatography in Biomolecular Separation
3.1.3 Importance of Chromatography in Everyday Life
3.1.4 General Principles of Chromatography
3.2 Phases in Chromatography
3.2.1 Column Chromatography: A Common Technique
3.2.2 A Key Technique for Molecular Separation
3.2.3 Types of Chromatography and Their Applications
3.3 Thin-layer Chromatography (TLC)
3.3.1 Principle of Thin Layer Chromatography (TLC)
3.3.2 Components, Procedure, and Applications of Thin Layer Chromatography (TLC)
3.3.3 Procedure of Thin Layer Chromatography (TLC)
3.3.4 Applications of Thin Layer Chromatography (TLC)
3.4 Gas Chromatography (GC)
3.4.1 Main Components of a Gas Chromatograph
3.4.2 Procedure of Gas Chromatography
3.4.3 Applications of Gas Chromatography
3.4.4 Advantages and limitations of Gas Chromatography
3.5 Liquid Chromatography (LC)
3.5.1 Mobile Phase
3.5.2 Principle of HPLC
3.5.3 Types of HPLC
3.5.4 Instrumentation of HPLC
3.5.5 Applications, Advantages, and Disadvantages of HPLC
3.6 Ion-Exchange Chromatography
3.6.1 Ion Exchange Process and Forms of Ion Exchange Chromatography
3.6.2 Instrumentation of Ion Exchange Chromatography (IC)
3.6.3 Ion-Exchange Chromatography: Process and Applications
3.6.4 Applications of Ion-Exchange Chromatography
3.6.5 Advantages of Ion-Exchange Chromatography
3.7 High-Performance Liquid Chromatography (HPLC)
3.7.1 Components of an HPLC System
3.7.2 Instrumentation
3.7.3 Normal Phase Chromatography
3.8 Reverse-Phase Chromatography (RP-HPLC)
3.9 Size Exclusion Chromatography (SEC)
4. ELECTROPHORESIS
4.1 Principles and Mechanism of Electrophoresis
4.1.1 Types of Supporting Gels
4.1.2 Applications and Advantages of Electrophoresis
4.1.3 Principle of Electrophoresis
4.1.4 Types of Electrophoretic Tanks
4.1.5 Procedure
4.1.6 Detection and Quantitative Assay
4.1.7 Quantitative Analysis
4.2 Gel Electrophoresis
4.2.1 Starch Gel Electrophoresis
4.2.2 Staining Solution (For Single Use Only)
4.2.3 Starch Gel Electrophoresis Protocol
4.2.4 Staining Procedure for Starch Gel Electrophoresis
4.2.5 Types of Electrophoresis
4.2.6 Variants of PAGE for Protein Analysis
4.3 Native PAGE Gel Electrophoresis
4.4 SDS-PAGE Electrophoresis
4.5 Isoelectric Focusing (IEF)
4.6 Capillary Electrophoresis (CE)
4.6.1 Instrumentation
4.6.2 Sensitivity and Sample Stacking
4.6.3 Detection Methods
4.6.4 Applications of Capillary Electrophoresis
4.7 Two-dimensional (2D) Electrophoresis (2D-PAGE)
4.8 Microchip Capillary Electrophoresis
4.8.1 Theory and Mechanisms of Action in Microchip CE: Injection
4.8.2 Injection in Conventional vs Microchip CE
4.8.3 Types of Injection in Microchip CE
4.8.4 Detection in Microchip CE
4.8.5 Applications of Microchip Capillary Electrophoresis (Microchip CE)
4.9 Agarose Gel Electrophoresis: Ideal for DNA and RNA Analysis
4.9.1 Principle of Gel Electrophoresis
4.9.2 Requirements for Gel Electrophoresis
4.9.3 Procedure for Agarose Gel Electrophoresis
5. SPECTROSCOPY
5.1 Fundamentals of Spectroscopy
5.1.1 Types of Spectroscopies
5.1.2 Applications of Spectroscopy
5.2 UV-Visible Spectrophotometry
5.2.1 Principles of UV-Visible Spectroscopy
5.2.2 Instrumentation of UV-Visible Spectroscopy
5.2.3 Instrumentation Components of UV-Visible Spectroscopy
5.3 Single-Beam Spectrophotometer
5.4 Double-Beam Spectrophotometer
5.4.1 Differences Between Single-Beam and Double-Beam Spectrophotometers
5.4.2 Applications of UV-Visible Spectroscopy
5.4.3 Additional Application
5.5 IR Spectroscopy
5.5.1 Basics of Infrared Spectroscopy
5.6 Instrumentation of Infrared (IR) Spectroscopy
5.6.1 Applications of IR Spectroscopy in Chemical Analysis
5.6.2 Applications of Infrared (IR) Spectroscopy
5.7 Fluorescence Spectroscopy
5.7.1 Fluorescence and Luminescence
5.7.2 Historical Background
5.7.3 Applications of Fluorescence Spectroscopy
5.8 Steady-State Fluorescence Spectroscopy
5.8.1 Principles and Theory of Fluorescence Spectroscopy
5.8.2 Key components of the Jablonski diagram
5.8.3 Important Characteristics
5.8.4 What is a Fluorescence Measurement?
5.8.5 Fluorescence Lifetime
5.8.6 Advantages of Lifetime Measurements
5.8.7 What is a Fluorophore?
5.9 Mass Spectrometry (MS)
5.9.1 Modern Era and Technological Breakthroughs
5.9.2 Basic Principle
5.9.3 Ionisation by Electron Bombardment
5.9.4 Applications of Mass Spectrometry
5.9.5 Principle and Working of Mass Spectrometry (MS)
5.9.6 Instrumentation and Steps of Mass Spectrometry (MS)
5.9.7 Common Types of Mass Analysers (Detectors)
5.9.8 Applications of Mass Spectrometry (MS)
5.10 Fourier Transform Infrared (FTIR) Spectroscopy
5.10.1 Introduction of Fourier Transform Techniques
5.10.2 Commercialisation and Advancement
5.10.3 Modern Era of FTIR Spectroscopy
5.10.4 Advanced FTIR Capabilities
5.10.5 Application of Fourier Transform Infrared Spectrophotometry in Pharmaceutical Drug Analysis
5.10.6 Principle of FTIR (Fourier Transform Infrared) Spectroscopy
5.10.7 Quantitative and Qualitative Analysis
5.10.8 FTIR Instrumentation and Interferometry
5.10.9 Fourier Transform and Spectrum Generation
5.10.10 Principles of FTIR Spectroscopy
5.11 Atomic Absorption Spectroscopy (AAS)
5.11.1 Principle and Analytical Significance of Atomic Absorption Spectroscopy (AAS)
5.11.2 Principle of Atomic Absorption Spectroscopy (AAS)
5.11.3 Beer-Lambert Law
5.11.4 Types of Atomic Absorption Spectroscopy (AAS)
5.11.5 Process Steps in GFAAS
5.11.6 Instrumentation of Atomic Absorption Spectroscopy (AAS)
5.11.7 Applications of Atomic Absorption Spectroscopy (AAS)
5.11.8 Working Principle of an Atomic Absorption Spectrometer (AAS)
5.11.9 Atomic Absorption Spectrophotometry (Flame Method)
6. X-RAY DIFFRACTION (XRD)
6.1.1 X-Ray Diffraction (XRD): An Overview
6.1.2 Instrumentation
6.1.3 Sample Mounting System
6.1.4 Basics of Crystallography
6.1.5 Basic Concepts in Crystallography
6.1.6 Principles of X-ray Diffraction
6.1.7 Crystal Thickness and Resolution
6.1.8 X-ray Diffraction Techniques
6.2 Single-Crystal X-ray Diffraction (SCXRD)
6.3 High-Resolution X-ray Diffraction (HRXRD)
6.4 Grazing-Incidence X-ray Diffraction (GIXRD)
6.4.1 XRD in Geology
6.4.2 The Diffraction of X-rays
7. FLOW CYTOMETRY
7.1 Advanced Flow Cytometry: An Overview
7.2 Fundamental Principle
7.3 Key Applications of Flow Cytometry
7.4 Applications of Flow Cytometry
7.5 Research Applications of Flow Cytometry
7.6 Principle of Flow Cytometry
7.7 Hydrodynamic Focusing in Flow Cytometry
7.8 Adjusting Flow Rate and Resolution
7.9 Optics System in Flow Cytometry
