Flashcards
Some Questions you should know
Cards in this deck (14)
- 1What is a MTT assay? How does it work?The MTT assay is a colorimetric biochemical assay for measuring the activity of cellular enzymes that reduce the tetrazolium dye, MTT, to its insoluble formazan, giving a purple colour. The assays measure cellular metabolic activity via NADPH dehydrogenase to reduce the yellow water-soluble tetrazolium substrate into a dark blue/purple water-insoluble formazan product in viable cells.
- 2What is ethidium bromide (EB)/acridine orange (AO) fluorescent assay?EB and AO are fluorescent dyes. EB stains dead cells red to orange, and is excluded by viable cells, while AO is a cell-permeable dye that traverses the cell membrane and stains the nucleus of the viable cells bright green. They allow for rapid discrimination of viable and dead cells.
- 3What is CellRox assay?CellRod is a cell permeable fluorogenic probe used to detect ROS, te cellular marker of oxidative stress. The dye is non-fluorescent while in the reduced state and exhibits bright fluorescent upon oxidationi by ROS.
- 4What is endoderm?the primitive tissue from which the pancreas [also stomach, colon, liver, uniary bladder, epithlelial pars of trachea, lungs, pharynx, thyroid, intestine] develops
- 5What are pancreas progenitors?Pancreas progenitors are the cells that will differentiate to become alpha cells (glucagon secreting cells) and beta cells (insulin secreting cells)
- 6What is ELISA?ELISA involves the use of an antibody to quantify the amount of protein (antigen) in a complex mixture with many other proteins. The antibody binds selectively to the antigen (ignoring everything else in the original sample mixture) and this amount of bound antibody is quantified, often through a colorimetric assay and then compared to a calibration curve form which is inferred the concentration of test protein in the mixture.
- 7How is ELISA carried out?-the sample with an unknown amount of antigen is immobilized on a solid support either non-specifically (via adsorption to the surface) or specifically (via capture by another antibody to the same antigen, in a "sandwich" ELISA)
-the detection antibody is then added, forming a complex with the antigen. The detection antibody can be covalently linked to an enzyme, or can itself be detected by a secondary antibody that is linked to an enzyme through bioconjugation
-between each step, the plate is washed with a mild detergent soln to remove any proteins or antibodies that are non-specifically bound
-after the final wash, the plate is developed by adding an enzymatic substrate to produce a visible signal, indicating the quantity of antigen in the sample - 8What is the difference between monocytes and macrophages? What is a chemokine?A monocyte is one type of circulating leukocytes (white blood cells). At a site of inflammation it leaves the circulation and becomes a macrophage.
The macrophage is the active phagocytic cell associated with microbial cell destruction, among other functions. - 9What is flow cytometry? How does it work?Flow cytometry is a means of separating and analyzing cells on the basis of size (forward scatter) granularity (i.e. number/size of internal organelles, side scatter) and fluorescence, the last after labeling the cells with fluorescent antibodies or other markers. Cells are dispersed in a carrier fluid, passed through a nozzle where each cell is "interrogated" by one or more lasers. Detectors placed at 180, 90 or at other angles (relative to the incident beam) provide information about each cell passing at the incident beam. Cells can also be collected based on some or all of the detected characteristics and this is called "cell sorting"
- 10What is a dot plot and what do abscissa and ordinate represent? What do the lines represent?Each doe is a single cell (or an "event", sometimes cell debris can be recorded). Typicall 1000 or 10 000 cells are examined in each experiment. Each axis is a different measure of the cell. E.g. side scattered light intensity (@ 90) vs. forward scatted light intensity (@ 180). Dots in the upper right quadrant represent big cells (high forward scatter) and high granularity (high side scatter). Dots in the lower left quadrant are the opposite. A typical analysis might determine the number of dots/cells in each quadrant as the percentage of the total.
- 11What does "gating" in flow cytometry mean?Gating is setting thresholds such that only cells with a parameter bigger (or smaller) than the threshold are counted or analysed further.
- 12How does PTH control calcium levels?The net effect of PTH is to cause bone resorption by activating osteo*C*lasts to dissolve bone and release calcium into the blood. It can also activate osteo*B*lasts to produce bone.
- 13What is sandwich ELISA? Outline how it works.-an antibody specific to the protein of interest is used to quantify the amount of the protein.
-two or three antibodies are used: one predeposited on a plastic dish captures the antigen, a detection antibody also binds to the antigen, and a third antibody (the secondary antibody) which is linked to an enzyme binds the detection antibody.
-the capture antibody captures only the molecule of interest (everything else is rinsed away).
-the detection antibody is also specific to the protein of interest, but contains a site for binding of the secondary antibody.
-the capture antibody is designed to attach well to the plastic dish. The enzyme linked secondary antibody is directed against any antibody of the species used as the source of the detection antibody, so that it will bind only to the detection antibody.
-the amount of colour is directly related to the amount of enzyme, the amount of enzyme linked to the antibody, the amount of detection antibody and hence amount of antigen
-a calibration curve with known concentration standards is required to relate colour to protein conc - 14What is pharmacokinetics?Pharmacokinetics refers to the changes in blood or tissue concentration with time after the drug (the pharmacological agent) is injected, injested or otherwise administered.
Information
- Description
- final Eng Bio Flashcards on Some Questions you should know, created by Reena Cabanilla on 21/12/2015.
- Created by
- Reena Cabanilla
- Views
- 0
- Rating
More on Laboratory Equipment and Analytical Techniques