4. (25 points total) You want to understand how the density of P-selectin on the
ID: 150913 • Letter: 4
Question
4. (25 points total) You want to understand how the density of P-selectin on the endothelial lining of blood vessels affects the rolling interactions of neutrophils (a type of white blood cell) when they are subjected to hydrodynamic drag forces in the blood. You introduce P-selectin into a synthetic lipid bilayer and attach it to a glass slide mounted on the floor of a flow chamber on a microscope stage. This arrangement allows you to measure neutrophil motion at different densities of P-selectin and at different flow rates. At high densities of P-selectin, from 40 to 400 molecules per um2, the cells stay attached to the membrane floor and roll slowly and irregularly with the flow of the medium. At P-selectin densities from 1 to 15 molecules per m2, the cells either move freely with the medium or become transiently tethered to the membrane floor before beginning moving again. When the system is treated with monoclonal antibodies against P-selectin, the tethering events do not occur, and the cells flow freely with the medium At the lower P-selectin densities, the number of tethering events per unit time is directly proportional to the density of P-selectin At a low flow rate, roughly equivalent to a cell being subjected to 40 piconewtons of shear, the time cells stay tethered follows a simple exponential decay curve with a cellular off rate, koff, of about 1.0 sec This off rate constant is unaffected by the density of P-selectirn a. What is the point of doing the experiment with the monoclonal antibody? (5 points) b. Is a single interaction between one P-selectin molecule and one copy its glycoprotein ligand on the -1 surface of the neutrophil sufficient to tether the cell to the floor of the flow chamber transiently? Explain your reasoning. (10 points) c. If you increase the flow rate by a factor of three, increasing the shear force felt by each cell to about 110 -1 piconewtons, kor increases to about 3.5 sec. Explain how this force can affect a dissociation rate constant. (10 points)Explanation / Answer
Answer of a- Monoclonal antibodies have monovalent affinity if monoclonal antibody does not binds with P- selectin ligand its result P- selectin have specific ligand on their surfaces.
Neutrophils bind with P-selectin resulted P-selectin have ligand that will bind with the receptor of neutrophil.
We can use monoclonal antibodies as a control for determinig the flow rate and roll of neutrophils against P-selectin.
We can determine the the teathering time of neutrophils against P-selectin using monoclonal antibodies as a controle.
We can determine the specificity of P-selectin specificity against neutrophils in the cancer and other deases.
Answer of b-
P-selecctin glycoprotein-1 is situated on neutrophil cell that will bind specifically with P-selectin.
Glycoprotein may have single functional group which will bind with P-selectin transientally.
Due to this binding the flow rate will be high.
When the single P-selectin it will have single glycoprotein so it will bind sufficientaly.
Due to single P- selectin shear force would be high so.
Their will be transient teathering as a result flow rate will be high because of single glycoprotein ligand.
although shear force is independent on the density of P-selectin
althoug both netrophile and P-selectin are having specific group for binding between them so only one iis enough.
Answer of C-
The flow rate depends on density of P-selectin
More the density of P- selectin flow rate will be slow.
If the density will be low then the interaction of neutrophil and P-selectin will be tranisient.
On increasing the shear force flow rate will be high because of high flow rat tension.
On increasing the flow rat by factor of three dessociation rate will be high because mentioned in question that 40 piconewton force increases 1 per second dissociation.
So flow rate by factor of three will increase three time more shear force as 40 into 3 = 120 near about and dissociation of neutrophil also increase three times as 3.5 netrophil per second.
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