Lung Cancer Cell Lab Report

Next, the biuret test for protein turned solutions in test tube 9 and 12 purple, the biuret test was positive, which proves that protein was present in the solutions, whereas test tubes 1,2,3,4,5,6,7,8,10, and 11 had negative results, this is because those solutions do not have peptide bonds and a peptide bond is necessary for the biuret test to be positive. An unknown was also tested in this experiment and we have concluded that the unknown #267 is a protein because it

Non-small cell is the most common form of lung cancer. In fact 9 out of 10 cases of lung cancer are non-small cell. The other main type of lung cancer is small cell lung cancer, and together, they are the leading cause of cancer deaths in the United States, surpassing women’s breast cancer in 1987. In 2015, 221,200 people are expected to be diagnosed with some form of lung cancer. After a patient is diagnosed, their life expectancy drastically drops, as patients die on average within one year of being diagnosed. Non-small cell lung cancer accounts for 85% of the fatality rate.

Assays are used as an indicator of whether or not the purification process is working correctly, or if the proteins have been destroyed in the process (1). The lab consisted of purifying wheat germ by utilizing the enzyme that is already present in the substance itself, acid phosphatase. Acid phosphatase was being separated from the wheat germ using a centrifuge (a machine that spins at high speeds in order to separate substances with different densities in a solution) during each step of the experiment to continuously separate and purify the sample (3). There were three assays in which were used during the experiment. One assay used during the experiment was a Bradford dye reagent, which was a blue dye that was added to samples of the supernatant to display the different concentrations of proteins present in each. Another assay that was used during the lab was the KOH. This reagent was added to different samples of the supernatants than what were used for the Bradford dye. The KOH was used to help stop the reaction that had originally taken place from the addition of the acid phosphatase reagent in step three, but its main use was to distinct the different concentrations of the proteins present in each step of the purification

Second, for the western blotting, the primary antibodies that used to recognize the protein of interest (FLAG-PTEN) are rabbit anti-PTEN antibodies and the secondary antibodies that bind to the primary antibodies and convalently linked substrate to produce visible signal are goat anti-rabbit conjugated HRP (horseradish peroxidase) antibodies; for immunoprecipitation, the antibodies used to immobilize protein of interest are anti-FLAG monoclonal antibodies from morine cell. As introduction above, those antibodies come from different species. In order to be successful in western blotting, the primary antibodies recognize the protein of interest as well as bind to it. Consequently, rabbit anti-PTEN antibodies were used to bind to FLAG-PTEN. Meanwhile, goat anti-rabbit conjugated HRP antibodies, the secondary antibodies,

The size was also determined by this method. The buffer chambers were filled with SDS-PAGE (Sodium Dodecyl Sulfate polyacrylamide gel electrophoresis), which helped add negative charges to the proteins and ran at 120 v for 30-50 minutes and then 100v for an hour. The samples were loaded in the order of ladder, S1, P1, P1 med salt, P1 high salt, S2, P2, P2 med salt, and P2 high salt. Western Bolt

The prepared samples were heated with 12.5% (v/v) β-mercaptoethanol for 5 minutes in a boiling water bath to denature the proteins. The SDS-polyacrylamide gel (Biorad, Any kD, Mini-PROTEAN TGX Gel, 10 well, 30 µl) was loaded with heated protein samples of crude extract, flow through, pre-dialysis, post dialysis, and molecular weight standard (Biorad). The gel was run at 24 mA until the tracking dye reached the bottom of the gel. The gel was removed and placed within a stain (0.4 mg/mL Coomassie Blue R250, 40% (v/v) methanol, 5% (v/v) acetic acid) until bands were

The respiratory system is made up of several important parts, including the trachea or “windpipe” where the cancer is most likely to originate (see fig.2 for location). Other parts include the pleura; the thin coating separated from the lungs by a small amount of liquid. This is in place to allow for easier breathing and to not corrode the chest walls when individual is inhaling. Lungs are heavily impacted by the formation of cancer because of the fact that they keep blood pumping to the heart and without that, the heart stops and breathing of the individual ceases resulting in

The cancers that are most common in men's are lung cancer, prostate cancer, and colon cancer. Lung cancer kills more over 50,000 men a year. More men are likely to get lung cancer because in general men smoke more than women. Non-small cell lung cancer referred as NSCLC. This is a dangerous disease that can immediately affect your quality of life by causing breathing difficulties. Two types of lung cancer have different subtypes that can affect your overall prognosis. The main subtypes of NSCLC are adenocarcinoma which starts in the outer part of lungs. Squamous cell carcinoma where it starts in the middle portion of lungs, and undifferentiated carcinoma where its fast-growing cells that can start in any portion of the lungs. The expectations of survival rate of having NSCLC is on a five year survival rate; which lung cancer can not be reversed. It was suggested that loss of control of cell cycle checkpoints is a common occurrence in lung cancers, and support of the idea that functional cooperation between different cell cycle inhibitor the proteins constitutes to another level of regulation in cell growth control and tumors suppression. Present review of literature mainly describes the cell cycle control, the lung cancer molecular pathogenesis, the catalog of known genetic alterations and the recent advances in the global expression profiles of lung tumors. As lung cancer cells are not

Cancer affects everyone at some point in their life, whether it be themselves or a relative. For myself, it was niece that was diagnosed with leukemia. While scientists have identified countless carcinogens, there still isn’t an exact science on how to avoid cancer. “Leukemia is the most common type of childhood malignancy under the age of 15” 1, and it makes me wonder what these kids at such a young age are predisposed to, that makes them more susceptible. When I think about cancer I associate it with radiation whether it be treatment or cause. Ionizing radiation is a known carcinogen and can increase the risk of developing Leukemia. What about radon? The radioactive gas we are subject to every day. My niece lives in an older home, has a

The method used in this experiment is the non- organic extraction where cell lysis solutions, proteinase K, isopropanol and ethanol was used. The functions of these solutions will be further discussed in the discussion.

A cell suspension containing 0.5 -1.0 x 10^6 cells/ml in media 10% FBS (Fetal Bovine Serum) was placed on a 24- well plate so a monolayer can be created at the bottom of the wells. The first step will consist of the lab student aspirating the media from the wells making sure to disrupt the monolayer of cells at the bottom of the wells. Next the wells will be washed with PBS to remove dead cells and debris. After the wells are washed 1ml of media (DMEM) in a 5% concentration will be added to keep the cells from dehydrating. 5% Media is used because the goal here is to allow migration and minimize proliferation. Next the lab student will create a horizontal “scratch” along the bottom of the well using a P200 pipette. The lab student should use enough force to crate the scratch but not too much because too much force will create uneven scratches. Finally the treatments will be added to the wells. The plate will then be placed under the light microscope and pictures of the scratches will be taken which will be labeled as (T0) Time point zero. Other pictures will be taken 24 hrs. later at (T24) time point 24. The photos will then be analyzed and compared the lab group to determine if the is any cell

Cancer is a name given to many diseases that are similar. In all kinds of cancer, blood cells divide uncontrollably(NCI). Science has had a huge affect on the way cancer is being diagnosed and treated. When doctors start to see symptoms like new moles/skin changes and quick weight gain or loss, they will do a number of tests, to see if you have cancer. Some of the tests include, Lab tests, biopsy, MRI, X-ray, and CT scan, though there are many additional tests. In lab tests, they check and see if the ratio of substances needed for your body are normal, or not. Normally, for these tests they use urine or blood. In a biopsy, a doctor will take a tissue sample and examine it to see if anything looks different. To remove tissue, doctors

type of tumor (eg. small cell or non-small cell): therapy changes depending on the tumor type.

The first thing we had to do in this experiment was label the 12 test tubes as C1- C4, G1- G4, and P1-P4. We filled one cylinder with water, another with glucose, and another with protein. Then we took the cylinder containing water and using a pipet we added 5 ml of water in each test tube marked C1-C4. We then took the cylinder containing glucose and with using a pipet we added 5 ml of glucose in each test tube labeled G1-G4. In the other tubes, we took the cylinders that contained protein and using a pipet, we put 5 ml in the test tubes that were labeled P1-P4. After

Figure 1 shows a standard curve developed from the cytoplasmic protein standards of lung cancer cells. A gradual increase in corrected absorbance can be notes, which more noticeably increases after 125 μg/mL. After this point the values nearly double. The absorbance readings for the samples were recorded along with the standard concentrations [500 μg/mL, 250 μg/mL, 125 μg/mL, 62.5 μg/mL, and 0 μg/mL (PBS only) 
The corrected absorbance for the protein standards were found by subtracting the average absorbance of the PBS blanks (0 μg of protein) from all other averages. 
Excel was then used to create a standard curve for the above protein standards, where protein concentration was plotted on the X-axis and corrected absorbances on the y-axis. The linear trend line was used to determine the protein concentration of the diluted sample according to the average corrected absorbance values. The dilution factor was accounted for by multiplying the calculated concentrations of all protein samples by 20. The volume (in μL) needed to load an equal amount of the volume (in μg) of the second protein sample was calculated, followed by the calculation of the RIPA volume needed to fill the protein sample to 10 μL. Four times the volume was then calculated for each volume and these values were used for the SDS-PAGE procedure the following week. The sample with the lowest concentration was used to calculate the amount of protein (in μg) used in the 10 μL solution, indicating the maximum