ELISA Lab

Introduction: (A) Elisa, which stands for Enzyme-Linked Immunosorbent Assay, is a test that can detect diseases in blood and bodily fluids using antigens that bind to the foreign antigens. When antigens trigger an immune system response, millions of antibodies find the antigens and bind to it. The secondary antigens that are enzyme linked are the next ones to bind to the antigens. Next, Chromogenic enzyme substrate is added which will color the wells. If they remain colorless, then they are negative, and if colored, the results are positive. Today, ELISA is used for many different purposes which include pregnancy tests, detecting drug use and disease in people, reassuring that food is labeled correctly and testing indoor air quality. With its rapid test results, ELISA has been very helpful for many diverse purposes. A very important aspect of the ELISA test is the antibodies. They prove to be useful because they can bind really well (with antigens). Some different uses of binding are immunostaining, immunoblotting or western blotting. Different kinds of cells (such as cancer cells) may be identified by the process of immunostaining. A protein's size and quantity may be determined by immunoblotting or western blotting. Antibodies can also be manufactured. Polyclonal antibodies are made by immunizing an animal. An animal (most likely a goat, rabbit, or sheep) would be injected with a purified protein that would allow the animal to make different antibodies to develop within the animal. Serum can then be collected by drawing blood from the animal and removing the blood cells from it. The serum that is isolated from the blood sample of the animal is called antiserum. The final step is to separate the antibodies from the antiserum. These antibodies are called polyclonal. The only down side to polyclonal antibodies is that no two batches will ever be the same. (B) The purpose of the experiment is to find out which two people in the class were originally diseased. This will be done by having everyone share fluids with three other people. After the ELISA test, the results will help to determine who the two disease spreaders were. In reality, the sharing of bodily fluids can be quite common. Someone may cough or sneeze on you with out you knowing and before you know it, you have become a victim to their cold they have shared with you. One in four teenage girls have an STD (from Mr. Chugh's stat of the day) and ELISA would be a way for them to find out. (C) On the first day of the lab, we will obtain a tube of some "bodily fluids". Then, with three other people, we will share our bodily fluids we got and record their tube number and the time at which the sharing occurred. On the second day, we will receive  twelve wells which we will add positive/negative controls, and our shared fluids to. It will then be cleaned out with wash buffer. The next step will be to add the primary antibodies then it will be cleaned out with wash buffer. The secondary antibodies are added next and then also cleaned out again. After it is rinsed with the wash buffer, substrate will be added and show the results. (D) I'm not really able to predict if i will be infected or not because the two diseased people could be anybody. It depends on who I share my fluids with. There are two controls in this experiment: a positive and a negative. The positive wells will be added with fluids that are diseased and should show up as blue. The negative wells will be added with fluids that are diseased and should be clear. For the results in our wells to be sufficient, the positive and negative controls must come out as expected when the substrate is added to it.

Results/Observations: After adding substrate, all twelve of the wells appeared to remain the same. This might mean a source of error occurred while the lab took place. Therefore, it remains a mystery if my lab group and I have been infected.

Discussion: With missing data from table four and five, it is almost impossible to determine who is the original diseased people are. The originally diseased people did share very quickly though because everyone is infected except for a few. The source of error in our lab was washing out the wells before adding the positive/negative controls and our fluids. The primary antigens did not have a chance to bind which made it impossible for the secondary antibody to bind and so on.

Proteomics

Introduction: (a) Genomics is the study of genomes and DNA. Proteomics is the study of proteins. The protein has shown us how the information that has been translated by DNA is used to make an organism. It can be studied to learn how genes that are made of DNA are transcribed by messenger RNA and then transformed into proteins.With the significant help of genomics and proteomics, there have been many discoveries that were once never considered abled to discover or explore. The Human Genome Project sparked the realization of the importance of the necessary variety of proteins. This was what led to a new field of study termed proteomics. Proteomics is more complicated than genomics because the proteome differs from cell to cell and is always changing. Specifically, proteomics studies the functions of proteins, interactions between proteins, their locations in the cell, their expression, and modifications of all proteins within development. The Human Proteome Organization is an international collaboration project which aims to achieve all the above studies of a protein. (b) In this lab, we will will use gel electrophoresis to look at muscle proteins from related fish species. Whether they are closely related or not will be determined at the end of the lab depending on the samples brought in and supplied. We will be able to identify similarities and differences based on the fish's proteins. (c) We will use gel electrophoresis to determine relations of fish.With this, we will need to collect samples from the fish (muscle). Adding buffer (the ingredients needed for the gel to work) to the samples will be the next step before adding the samples into the gel. The gel will run and hopefully the results will turn out to show how closely related each fish is to one another. (d) To insure that the results from gel are accurate, there will be a marker to act as a control for the gel electrophoresis.

GMO:yes or no???

Intoduction: (a) Genetically Modified Organisms or GMO, argued by proponents, are believed to solve the problem of the overuse of pesticides and herbicides on crops that may have long term effects on the environment and human health. GMO's are used to genetically manipulate crop plants to have specific traits. This technique is not new, but what is new is the option to take the genes out of another organism (doesn't have to be from a plant species) for a specific trait and directly place it into crop plants. Some examples include GMO's that are resistant to pests or drought, improved appearance, and increased crop yield. GMO's can be identified by and antibody based test (enayme-linked immucosorbent assay) which can spot the proteins that are made by the GM crops. Another way to detect GMO foods is by a process called the polymerase chain reaction that searches for the specific DNA sequence of a GM food. Today, there is much controversy over GM crops and the absence of the requirement to label GM foods in the US by the government. People who oppose GMO's say that there is the potential for "superweeds" that would occur by the cross pollination of natural weeds and herbicide resistent crops. Also "superbugs"could arise due to the toxins in pest-resistent crops. There is no diversity in the crops (monoculture) which makes them suseptable to a massive wipeout. People could have allergic reactions to certain proteins of the GM crops. Many say there isn't even enough reasearch done to forsee and understand the tinkering of plant diversity and manipulation. (b) The purpose of this lab is to test and see if the food item brought in is genetically modified. Since a label for GM foods is not required in the US, we are unable to identify them. With this experiment, we are able to find out if a favorite food we have been eating is genetically modified or not. (c) DAY 1-The first step will be to extract DNA from food samples. The sample which would be turned into a slurry will then be combined with instagene, the same will be done with the certified non-GMO food product(this will act as a control to test if the lab worked properly). The samples will then be put into a waterbath for five munutes and then refridgerated. DAY 2-Master mix is added to each pcr tube which will be iced. Then DNA template will be added to the pcr tubes but avoiding the pellet from the instagene. The pcr tubes will then be set in a thermal cycler which rapidly heats and cools. This allows the template strands to seperate or denature and the primers to anneal to the denaturated template strands. While in the cycler the DNA polymerase also extends the primers and makes complete copies of each DNA strand. DAY 3-After the pcr tubes have been in the microtube adaptor, loading dye will be added to each sample. The samples will be loaded into the agarose gel. Results are seen after stained in fast blast DNA stain.