5. Reductionist approach – isolate cells from animal and we can reduce the environment to very simple questions. Allows us to look at very specific questions but can be a disadvantage, taking cell out of normal environment so there is possibility that the cells wont behave the same as in its natural environments
9. Chemical – chemical enzymes that break up tissue gently. Trypsin/protinase K/collagenase. These cleave the tight adhesions between cells as well as the proteins that make adhesions with the exc matrix. Break up without damaging cell. Chemical treatments reduces the amont of mechanical dissociation needed. Ca2+ and Mg2+ ions are required for the proteins that hold the proteins that are held to the exc matrix to function. You can remove all the ions from the tissue and that will cause the proteins holding the cells to be released and the cells will release. This Is non-enzymatic.
10. Mechanical – put in buffer and take a homgenizer and break up tissue into small components.
12. Centrifuge – get rid of suspended cell pieces. Membrane bits etc. and take centrifuge material and plate on a primary culture. Will divide until reach confluency – cells have touched one another in a single cell layer. At this stage, cells stop growing b/c receptors required for cells to divide are bound to another saying theyre interacting with the cells and not enough growth factor to continue growing. This can also lead to cell death. But some will live and a secondary culture is created.
18. senescence and many die. (human cells die almost always.) mouse cells enter crisis, and some continue to grow (secondary culture) – immortal, cell lines.
19. There are human cell lines, but sometimes we must infect with a virus to generate a population of transformed cells. Or take directly from human tumor.
20. Embryonic stem cells can be used to ask specific questions and without destroying the embryo.
22. Prepare sample for imaging. If you have thick samples, you must reduce thickness of sample. A common way is to section it on a microtome. Take fine blade and slice in order to visualize.
28. Magnification – lets you see how large you can view an object while keeping the components as distinct objects. How big can the object be while maintaining objects as distinct?
29. Contrast – formed by property of cells themselves. Light passes through cells. The cells and organelles are transparent. No definition with organelles and outside environment. Light microscopes. Use dyes to stain to provide contrast and most dyes require you fix and kill cells. To be living you need to manipulate the technology. Add phase contrast or use DIC – interference contrast technology.
![1.20.10<br />cysteine – creates disulfide bonds. They occur when the proteins are processed in the golgi apparatus and are either secreted or transported to the plasma membrane. <br />Consider it hydrophilic than hydrophobic. <br />C and H bonds make amino acids hydrophobic.<br />Cell culture:<br />Epithelial – skin, intestines – cells held together and held on exc matric very tightly. Hard to break apart, but can be done. <br />Mesenchyal – blood/bone/smooth muscle. An exc matrix that is widespread and is connective is a mesenchyal. <br />Primary tissue has yielded cell lines – characteristics of the primary tissue but are continually dividing without reaching senescence. <br />,[object Object]](https://image.slidesharecdn.com/1-20-2010lecture-100503153930-phpapp01/85/1-20-2010-lecture-1-320.jpg)
