(II) Our Mission ~ Part 2

- Histochemical Staining

1) What are the colours of the stained tissue?

Pale pink and Purplish-pink.

2) What is the pattern of colour distribution in the stained tissue?

There are about 25% Purplish-pink and the rest are Pale pink.

3) Can you identify which are the nuclei of the cells? What colour are they?

Purplish-pink round spots are the nuclei of the cells.

4) Can you identify the cytoplasm of the cells? What colour are they?

The pale pink Y-shapes are the cytoplasm of the cells.

5) What are the relative sizes and shapes of the cells in the tissue?

6) How many different types of cells do you think there are in the tissue?

1 type of cell.

7) Are there any other observations you identified? If so, describe them.

The dye was unevenly spread thus could contribute to an unreliable result.

- Immunohistochemical Staining

1) What are the colours in the tissue?

Light brown and a tinge of green.

2) What is the pattern of colour distribution in the stained tissue?

10% brown surrounding the 90% green.

3) Are the colours and pattern of colour distribution similar to that in the first slide?

No.

4) The purpose was to determine the presence and location/distribution of a protein called vimentin. Which colour do you think represents vimentin? Why?

Light brown. Cells produce vimentin thus the surface area of the cell should be surrounded by vimentin and light brown was observed on the surface around the cells.

5) Describe the location/distribution of vimentin in the tissue.

It is distributed unevenly and found in the greatest concentration around the cells.

6) What does the other colour(s) represent?

It represents the cytoplasm of the cells.

7) Are there any other observations you have identified? If so, describe them.

None observed.

- Fluorescence Staining

1) Can you see the tissue using the fluorescence microscope? Does the tissue apear similar to the first and second slide?

The tissue is not visible under the fluorescence microscope and the organelles observed are nothing similar to the first and second slides.

2) Describe what you observe. How many colours appear when using the fluorescence microscope and what are the colours?

Blue and Black colours were observed.

3) What do you think the colours represent? Why?

Blue represents the nuclei of the liver cells while black represents the background and the rest of the organelles of the cells that cannot be stained by the Hoescht dye. The hoescht dye becomes visible as a blue colour under the UV lights at a certain wavelength and since it only attaches to the DNA in the nuclei of the liver cells, the nuclei appears to be blue while the unstained portions of the tissue appears black as UV lights are not visible naturally.

4) Can you determine what is/are the pattern of location/distribution of the colours in the tissue? If no, why not?

No, we do not know how the cells in the liver tissue looks like under the fluorescence microscope as it only shows us the nuclei of the cells.

5) Are there any other observations you have identified? If so, describe them.

Yes, all the nuclei of the liver tissue appears to be of the same shape, This can attribute to identifying different substances quickly.

With reference from Manual for applied challenge project, Dr. Kenneth Yu, Division of Chemical Biology and Biotechnology, School of Biological Sciences, Nanyang Technological University.