ADVANCED MAMMALIAN CELL CULTURE TECHNIQUES. PRINCIPLES AND PRACTICES

• SECTION 1. BASICS IN CELL CULTURE
1. Highlights: History of Cell Culture and Its Applications
2. Aseptic Technique and Essentials in Cell Culture
3. Laboratory Safety
4. Preparation of Media for Cell Culture
5. Primary Cell Culture
6. Passaging of Cell Lines
7. Cell or Tissue Culture Models and Cell Line Immortalization
8. Counting of Cells by Hemocytometer
9. Cryopreservation of Cells
10. Thawing of Cells
11. Detection of Cacterial and Fungal Contamination
12. Testing for Mycoplasma Contamination
13. Cellular Cytotoxicity
• SECTION 2. ISOLATION AND ANALYSIS OF NUCLEIC ACIDS AND PROTEINS
14. Isolation of Total RNA and Assessment of Purity and Concentration
15. Target Gene Amplification and Expression by qRT-PCR
16. Cell Lysis, Protein Extraction Using RIPA Buffer
17. Estimation of Protein by Bradford’s Assay
18. SDS-PAGE Analysis
19. Immunoblotting
• SECTION 3. CELL STANING TECHNIQUES
20. Cell Nuclear Staining by DAPI
21. Immunofluorescence Staining
22. Estimation of Intracellular ROS using H2DCFDA Molecular Probes
23. Analysis of Cell Cycle Regulation and Apoptosis Using Annexin-V and Propidium Iodide (PI) Staining
24. Estimation of Chemokines and Cytokines Using Bio-Plex Assay
• SECTION 4. CELL TRANSFECTION AND SINGLE-CELL ANALYSIS
25. Reporter Gene Construction and Assay
26. Vector Transfection and Validation
27. Single-Cell Analysis

This up-to-date book compiles both basic and advanced laboratory techniques of mammalian cell culture. It is divided into four major sections encompassing the basics of cell culture, nucleic acid and protein isolation, cell-staining techniques, and cell transfection and single-cell analysis. The topics include aseptic handling, media preparation, and passaging of cells. The book also outlines downstream assays such as nucleic acid and protein isolation from in-vitro cell cultures.

Key Features:
• Covers cellular staining using fluorescent dyes, genetic manipulation of cells via transfection, and an introduction to single-cell analyses
• Discusses basics in cell culture and downstream applications including gene and protein expression analysis
• Includes the principles underlying each of the techniques and provides a detailed methodology to practice
• Explores the whole range of techniques – from basic to downstream applications and advanced methods

The book is essential for students and researchers in the field of life sciences, biotechnology, genetics, and molecular biology.