Figure 1: SDS PAGE Gel of Murine Fibroblast Cell Lysate samples. Lane 1, 5 µL pr

Question

Figure 1: SDS PAGE Gel of Murine Fibroblast Cell Lysate samples. Lane 1, 5 µL protein marker added. Lane 2, sample 3 containing 5 µL of a 16 µL of cell lysate solution. Lane 3, containing of 5 µL of a 9.6 µL cell lysate and 6.4 µL distilled water dilution solution. Lane 5, containing 5 µL of an 8 µL Bovine Serum and 32 µL distilled water solution. Lane 6, containing 5 µL of a 4 µL Bovine Serum and 16 µL of distilled water solution. Lane 7, containing 5 µL of a 2 µL Bovine Serum and 8 µL of distilled water solution. Lane 8, containing 5 µL of an 8 µL Horse Serum and 32 µL distilled water solution. Lane 9, containing 5 µL of a 4 µL Horse Serum and 16 µL of distilled water solution. Lane 10, containing 5 µL of a 2 µL Horse Serum and 8 µL of distilled water solution.

Fro left to right: Lane 10, Lane 9, Lane 8, Lane 7, Lane 6, Lane 5, Lane 4, Lane 3, Lane 2. Lane 1

Lab: SDS PAGE Gel of Murine Fibroblast Cell Lysate samples.
1. Discuss how one could improve there SDS PAGE Gel technique?
2. give some possible errors that could affect results when using SDS PAGE Gel to seperate and figure out what proteins are shown?
3. what would cause SDS PAGE Gel bands to be unclear? too thick? too fuzzy?

Explanation / Answer

According to the information, it can be judged that lanes 1,2 and 3 contain different amount of cell lysates in them. It can be seen that although the lane 1 gives some bands but no bands are visible in lane 2 or 3. This denotes that there is extremely low or degraded protein. Also, different concentrations of serum proteins give different intensities of bands. Higher serum concentration gives stronger bands and reverse is true for low concentrations of serum.Thus, the answers can be found as below:

Part 1: In order to improve the quality of SDS PAGE, following trouble shoots can be performed:

Thus, by performing these troubleshoots, a better SDS PAGE can be run.

Part 2: Some of the possible errors can be discussed as ahead:

Part 3: The possible reasons that the bands are unclear can be either protein degradation or poor sample preparation. The reason for too thick bands is presence of too much protein in some of the samples, especially low molecular weight proteins. Similarly, fuzzy bands appear when some of the proteins degrade. Also, poor preparation of SDS PAGE gel can lead to poor resolution of proteins, hence fuzzy bands appear.

Thus, following these troubleshoots, a better SDS PAGE can be performed.

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