Breast Cancer - Extensive Glycosylation of the Cells - Lab Report Example

For the sixth cell line, 5ug/ml street texas-red and 10 ug/ml VVA-Biotin was used. Then the blocking was done with the 5% Carbo-Free Blocking Solution.
Hypothesis
The key difference observed based on the way the two cell lines were treated includes: For the fifth cell line, was treated with 20 ug/ml VVA-Biotin and blocked with a 5%BSA Blocking solution. On the other hand, the sixth cell line was treated with 10 ug/ml VVA-Biotin and blocked with a 5% Carbo-Free Blocking Solution.
Aim of the experiment
Based on the treatment differences, the experiment aimed to ascertain the level of staining that could be observed between these two cell lines.

Results
The materials that were compared included pictures of breast cancer cell lines (fifth and sixth cell lines). The ImageJ software was used to analyze the data that belonged to the two breast cancer cell lines. In this case, the two cell lines were compared after staining using street Texas-red with the treatment of BSA-blocking solutions and Carbon-free blocking solutions. To determine the glycosylation’s intensity, in each cell line, histograms were used (see images below). This shows the distribution of gray values. Despite, they were RGB images, it is seen that each pixel had to be converted to grayscale with an inbuilt formula. The total pixel count has also been indicated together with the standard deviation, maximum, minimum, and mean as well as the gray value mode. The results are presented in the next section.

In Figure 1 above it can be observed that the total pixel count is equal. Hence the results can be compared. The mean in the fifth cell line is 8.26 while for the sixth cell line is 29.628. Therefore, it is evident that the color intensity is higher in the sixth cell line than in the fifth cell line

The same pattern is observed in the surface plot below whereby the sixth cell line has a higher intensity than the fifth cell line
According to studies, it is known that glycosylation is more pronounced in breast cancer cells (Hakomori 1985). The reason for this is because of the increase in the oligosaccharide structures (Hudis 2007). Such alterations might be identified through lectins which are carbohydrate-binding proteins that have very high specificity for the saccharides(Brooks and Harris 2006). The lectins normally function as data mediators in biological systems. In this case, they interact with the glycoproteins glycolipids as well as the oligosaccharides(Nilsson 2007). They also bind specifically to the carbohydrate epitopes(Staffieri et al. 2012).

The immunofluorescence method as well as the direct lectins cytochemistry with the use of the FITC-labeled lectins enables the visualization of the glycosylation’s impact on the therapies which target the surface of the cells that bind to theHER2 receptors (Gabius 2009. The results from the histograms above (fig 1) indicate the reason why the glycosylation’s intensity is higher in the sixth cell line when compared to that of the fifth cell line because of the different blocking agents used. In summary, further study needs to be done to clearly understand why there was a difference in the intensity of these two cell lines. It could be due to the above three mentioned reasons. Unfortunately, this study could not ascertain this.

Conclusion
In conclusion, based on the difference in the treatment of the two cell lines, the sixth cell line which was blocked with a carbon-free blocking solution is seen to have intense staining as compared to the fifth cell line which was blocked with a BSA blocking solution. The reasons that could have resulted in the difference in the intensity could be due to; firstly, the BSA could be a better-blocking agent than carbon-free because as indicated in the figures above, there is less nonspecific binding when the cells were blocked with BSA as compared to when they were blocked with carbon-free.In this case,e the high intensity could be due to the nonspecific binding of VVA-Biotin. Therefore, BSA could be considered the best blocking agent over carbon-free. Secondly, there could be less glycosylation in the fifth cell line than in the sixth cell line. As seen in the results, the sixth cell line staining is more intense than the fifth cell line. Thirdly, since there was a difference in the amount of the VVA-Biotin used between these two cell lines whereby 20 ug/ml VVA-Biotin was used for the fifth cell line and 10 ug/ml VVA-Biotin for the sixth cell line, Read More