Established cell culture.pptx

Established cell culture Shashi Singh Cells that can be passaged indefinitely , are transformed inherently. Such line scan be obtained from tumors/cancers. They arise from primary cell cultures. Primary cultures are initiated directly from the cells, tissues, or organs of animals and are...

Established cell culture Shashi Singh Cells that can be passaged indefinitely , are transformed inherently. Such line scan be obtained from tumors/cancers. They arise from primary cell cultures. Primary cultures are initiated directly from the cells, tissues, or organs of animals and are typically used in experiments within a few days. By convention, the passaging or subcultivation of a primary culture begins a cell line. So cell lines that start showing senescence are primary derived transient cell lines The ones that continue to grow without showing signs of ageing are the transformed cell lines. The first cell line that was established was by G Gey from JH Medical college from a middle age women suffering from cervical cancer. Henrietta Lacks, H Lacks did not survive but her cell lines continues to be passaged till today. The advantage of working on established cell lines is uniformity of samples. Unlimited supply no ethical issues No heterogeneity A cell line is a permanently established cell culture that will proliferate indefinitely given appropriate fresh medium and space. Cell lines differ from cell strains in that they become immortalized. Once established as a continuous cell line, the culture continue to proliferate indefinitely and becomes an inexhaustible source of material due to their immortal nature. Cell lines are often used for research, for diagnostic assays, therapeutic screens, etc due to relatively homogenous nature and character. It has been difficult initially to obtain cell lines with the first cell line established in 1950, now Methods of establishing continuous cell lines have made it relatively easy The chances of getting a cell line are higher, obtained from cancers/tumorigenic origins Spontaneous mutations – once in eternity one may spot a colony in primary cultures that appears un-aged And faster growing that colony can be picked up by localized trypsinization and replated in fresh dish. By cloning- cells are trypsinized and diluted to be plated singly in a 96 well plate After a few days the well which show up a colony can be transferred to bigger dishes Induced established cell lines Cellosaurus, which acts as a cell line knowledge resource, lists more than 107,000 cell lines, with approximately 80,000 of human origin AtCC that is source from where one can obtain cell lines for a price also lists a number of mammalian cell lines Although cell lines are widely used, they are subject to quality concerns. Misidentified cell lines fail to correspond to the correct donor origin and instead correspond to a cell line from a different donor. Misidentified cell lines can arise through mislabeling of cultures or through cross-contamination, where cells from another culture are accidentally introduced without the User’s knowledge. Cross-contamination initially results in a mixture of cells from different sources. However, a faster growing contaminant can replace the original, authentic material after only a few passages. One common source of contamination demonstrated that HeLa cells (the commonest cause of misidentification) are readily spread via droplets during handling procedures. So characterization of cell lines is essential and we shall tak eup various topics in this field Morphology so microscopy Fluorescence various fluorescence techniques used in cell culture characterization Gene expression based techniques for characterization STR analysis for cell lines identification