By Awtar Krishan
A much-needed primer at the use of laser circulate cytometry for stem telephone analysis
Laser circulation cytometry is a robust instrument for quick research of cells for marker expression, telephone cycle place, proliferation, and apoptosis. besides the fact that, no assets in particular deal with using this system for the examine of stem cells; this can be specially vital as stem phone research consists of really expert equipment and marking techniques in keeping with particular features reminiscent of marker expression, phone measurement, drug delivery, and efflux of the stem cells.
Now, this e-book reports those strategies, discusses the technology in the back of them, and gives real-world examples to demonstrate the usefulness of the equipment. It brings jointly world-class specialists in pathology, biophysics, immunology, and stem cellphone examine, who draw upon their broad event with the tools and exhibit examples of excellent information to aid consultant researchers within the correct path.
bankruptcy insurance contains:
- Stem phone research and sorting utilizing part inhabitants
- Flow cytometry within the examine of proliferation and apoptosis
- Stem mobilephone biology and alertness
- Identification and isolation of very small embryonic-like stem cells from murine and human specimens
- Hematopoietic stem cells—issues in enumeration
- Human embryonic stem cells: long term tradition and cardiovascular differentiation
- Limbal stem cells and corneal regeneration
- Flow cytometric sorting of spermatogonial stem cells
- Breast melanoma stem cells
- Stem cellphone marker expression in cells from physique hollow space fluids
This publication is an important source for all graduate scholars, practitioners in constructing international locations, libraries and booklet repositories of universities and learn associations, and person researchers. it's also of curiosity to laboratories engaged in stem telephone examine and use of stem cells for tissue regeneration, and to any association dealing in stem telephone and tissue regeneration research.Content:
Chapter 1 fundamentals of circulation Cytometry (pages 1–12): H. Krishnamurthy and L. Scott Cram
Chapter 2 sensible concerns for circulate Cytometric Sorting of Stem Cells (pages 13–23): Geoffrey W. Osborne
Chapter three Stem telephone research and Sorting utilizing facet inhabitants research (pages 25–43): William Telford
Chapter four move Cytometry within the research of Proliferation and Apoptosis (pages 45–60): Michael G. Ormerod and Ronald M. Hamelik
Chapter five stream Cytometric research of Drug delivery and Efflux in Stem Cells (pages 61–74): Awtar Krishan and Ronald M. Hamelik
Chapter 6 Stem cellphone Biology and alertness (pages 75–89): Swapnil Totey, Rajarshi buddy, Murali Krishna Mamidi, Vijayendran Govindasamy and Satish Totey
Chapter 7 identity and Isolation of Very SmaU Embryonic?like Stem Cells from Murine and Human Specimens (pages 91–101): Ewa ok. Zuba?Surma, Dong?Myung Shin, Habella Klich, Janina Ratajczak, Magda Kucia and Mariusz Z. Ratajczak
Chapter eight digital quantity of Hematopoietic Stem Cells (pages 103–114): Siddharth Sharma and Awtar Krishan
Chapter nine Hematopoietic Stem Cells: concerns in Enumeration (pages 115–134): Michael Keeney and D. Robert Sutherland
Chapter 10 Embryonic Stem Cells: improvement and Characterization (pages 135–159): Vijay Bhaskar R. Konala, Villoo Morawala?Patell and Aparna Khanna
Chapter eleven Human Embryonic Stem Cells: Long?Term tradition and Cardiovascular Differentiation (pages 161–173): Maneesha Inamdar
Chapter 12 Mesenchymal Stromal Cells and Their scientific functions (pages 175–188): Jyoti Kode and Vivek Tanavde
Chapter thirteen Circulating grownup Stem Cells of Hematopoietic beginning for Vascular and Neural Regeneration (pages 189–209): Lissy okay. Krishnan
Chapter 14 move Cytometric Characterization of Neural Progenitors Derived from Human Pluripotent Stem Cells (pages 211–222): Raj R. Rao, Sujoy okay. Dhara, Shilpa Iyer and David W. Machacek
Chapter 15 Limbal Stem Cells and Corneal Regeneration (pages 223–240): Geeta ok. Vemuganti, Murali Mohan Sagar Baila and Shubha Tiwari
Chapter sixteen move Cytometric Sorting of Spermatogonial Stem Cells (pages 241–250): B. S. Srinag, J. M. Kalappurakkal, G.H. Mohan and H. Krishnamurthy
Chapter 17 Breast melanoma Stem Cells (pages 251–270): Devaveena Dey and Annapoorni Rangarajan
Chapter 18 Tumor Stem mobilephone Marker Expression in Cells from physique hollow space Fluids (pages 271–277): Awtar Krishan, Deepti Sharma and Ronald M. Hamelik
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Additional resources for Applications of Flow Cytometry in Stem Cell Research and Tissue Regeneration
Triel C, Vestergaard ME, Bolund L, Jensen TG, Jensen UB. 2004. Side population cells in human and mouse epidermis lack stem cell characteristics. Exp Cell Res 295:79-90. Umemoto T, Yamato M, Nishida K, Kohno C, Yang J, Tano Y, Okano T. 2005. Rat limbal epithelial side population cells exhibit a distinct expression of stem cell markers that are lacking in side population cells from the central cornea. FEBS Lett 579:6569-6574. Umemoto T, Yamato M, Nishida K, Yang J, Tano Y, Okano T. 2006. Limbal epithelial side-population cells have stem cell-like properties, including quiescent state.
However, it still retains some of its blue fluorescence emission when not bound to chromatin, but loses almost all of its red fluorescence in the loosely bound or unbound state. The decrease we observe in Hoechst 33342 fluorescence in the SP (unbound) population is due largely to the shift in red fluorescence; the SP population is therefore not a straight line down to the minimal axis, but is curved and arched, maintaining some blue fluorescence but losing almost all red emission. This bichromatic shift allows the SP population to be distinguished from other sub-Gi populations when displayed in a bivariate dot plot, including dead cells, debris, and some mature cell populations that also display dye efflux.
2. Practical Aspects of Hoechst SP Analysis The cell-permeable DNA binding dye Hoechst 33342 (not to be confused with Hoechst 33258, which is spectrally similar but is not cell permeable) has an excitation maxima in the ultraviolet (Xex = 351 nm). An ultraviolet laser source is therefore required for good Hoechst SP resolution. Unfortunately, ultraviolet lasers have traditionally been uncommon fixtures on flow cytometers [Shapiro and Telford, 2009]. Until recently, a large-frame water-cooled gas laser, such as a Coherent or Spectra-Physics laser, was usually needed to produce ultraviolet laser excitation for flow cytometry.