Question

In this experiment, the samples were fixed using a solution of 4% formaldehyde. This solution was dissolved in a solution that contained 50 mM PIPES, 2 mM MgSO4 and 2 mM EGTA, and also contained 0.1% (v/v) of DMSO and a detergent called Triton X-100.

Q1. If a blocking solution was not used in the experiment, what might happen in the labelling reaction, and how might this affect the immunolocalisation of the microtubules that you would view with the microscope?

Q2. In this experiment, the microtubules were labeled with a secondary antibody tagged with the dye Cy3. What are the maximum excitation and emission wavelengths for this dye

Q3. The dividing plant cell shows two microtubule arrays (patterns) that are associated with cell division that do not occur in animal cells. What are these arrays and at what stages during cell division do they occur?

Answer 1

1. If a blocking solution is not used, there can be nonspecific interactions of the antibodies used to label the microtubule proteins and they might bind to regions that are not associated with the microtubules or where they are not intended to bind.This will lead to incorrect visualization of regions that might not be associated with microtubules.

2. Cy3 has an excitation maxima of 550nm and an emission maxima of 570 nm.

3. The microtubules patterns seen in plant cells are

pre prophase band(PPB) and phragmoplast.

the preprophase band is found in plant cells that are about to undergo cell division and enters the preprophase of mitosis. ( This phase regulates the control of cell division plate and placement of the cell wall )

phragmoplast- forms during late cytokinesis. it helps in formation of the cell plate and subsequently a new cell wall that separates the two new cells.