By L. Scott Cram (auth.), R. C. Sobti, Awtar Krishan (eds.)
Flow cytometry has speedily advanced right into a approach for fast research of DNA content material, mobile marker expression and digital sorting of cells of curiosity for extra investigations. movement cytometers are being commonly used for tracking of mobile DNA content material, phenotype expression, drug delivery, calcium flux, proliferation and apoptosis. Phenotypic research of marker expression in leukemic cells has develop into an immense software for diagnostic and healing tracking of sufferers. fresh stories have explored using movement cytometry for tracking hormone receptor expression in human reliable tumors and for reports in human genomics. Contributions within the present quantity are in accordance with shows made on the First Indo-US workshop on circulation Cytometry within which specialists from united states, united kingdom and India mentioned functions of circulation cytometry in organic and clinical study. This publication could be of curiosity to publish graduates and researchers within the fields of pathology, cytology, cellphone biology and molecular biology.
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Extra info for Advanced Flow Cytometry: Applications in Biological Research
33, pp. 377-383 (1990) . • L. S. Cram, F. A. Ray, and M. F. Bartholdi, Univariate Analysis of Metaphase Chromosomes Using the Hypotonic Potassium ChloridePropidium Iodide Protocol. In: Methods in Cell Biology, Vol. 33, pp. 369-377 (1990). Acknowledgements Collaborators at the National Flow Cytometry Resource, Los Alamos National Laboratory, who have made outstanding contributions to our development of flow cytogenetics include James Jett, Carolyn Bell, Joe Fawcett, John Martin, and Larry Deaven.
Sorted human chromosomes, as well as other species, have been cloned using several vectors as diagramed in Figure 8. The availability of human libraries constructed from a signal human chromosome simplified the project by being able to assign and map DNA sequences known to have come from a single chromosome type. Cosmid vectors can accept a 40 kb fragment of DNA. DNA of this size and molecular weight can be achieved in a flow cytometer by taking appropriate steps. Successful sorting is accomplished using the polyamine buffer as the sheath (see Appendix), accumulating a packed pellet of material, and maintaining the sorted material at wet ice temperatures.
Syringe 3x with a 22 gauge needle (1-1/2" long) using 3 ml size for Iml volume; 5 ml syringe for 3 ml sample. 12. Pool tubes if necessary. 13. Incubate 37° for 30 min in water bath. 14. Stain with DNA stain: use 20 Jlg/ml PI or 2 Jlg/ml Hoechst 33258 or 20 Jlg/ml CA3. Appendix 3. Polyamine Buffer for Bivariate Human Flow Cytogenetic Analysis and Sorting I. Materials A. Stock solutions: Stock solution I: Prepare a lOx solution (stock solution I) containing Tris, EDTA, KCI, and NaC!. 150 mM Tris-HCI 20 mM EDTA (Trizma Hydrochloride, Sigma cat.
Advanced Flow Cytometry: Applications in Biological Research by L. Scott Cram (auth.), R. C. Sobti, Awtar Krishan (eds.)