Disadvantages Of Immunoassay

Affinity chromatography involves the property of bio-recognition for the separation of proteins based on the reversible interaction between the protein and the specific ligand bound to a chromatography matrix. It enables us to purify bio-molecule on the basis of its biological function and its chemical structure. The goal of the experiment was to gain hands-on experience in protein purification by using AKTA FPLC system. The IgG antibody contained in the mouse antiserum was purified by protein G affinity chromatography using a HiTrap protein G HP column. The chromatogram was then obtained and analyzed to ensure the separation of IgG antibody. Such purification of the protein before sample preparation is usually done to avoid any undesired interactions.

ELISA works on the principle of an antigen binding to specific antibody (lock and key), which can be used as a way to identify quantities of proteins in a small sample of fluid. The specific proteins used in an ELISA are estimated quantitatively. The ELISA test is carried out by incubating the serum that contains the antigen of interest with antibody’s within a well, in order for the antibody’s to bind with the specific antigens. The plate is then washed with a mild detergent in order to remove any proteins that have not been bound. The washing of the plates is carried out between every step in order

Western blotting - In Western blotting first, the macromolecules have to be separated via gel electrophoresis. The molecules now separated by electrophoresis are blotted onto either a nitrocellulose or a polyvinylidene difluoride (PVDF) membrane (a second matrix). To inhibit the binding of nonspecific antibodies to the membrane surface it is subsequently blocked. Then a complex is formed (a probe) from the protein that was transferred and an enzyme linked with an antibody. The enzyme is supplied a substrate then the 2 together should create a product e.g. chromogenic precipitate that can be detected. Detection methods with most sensitivity use chemiluminescent substrate because light is a by-product of the reaction between the substrate and the enzyme. The output of the light can be measured using a CCD camera or on the other hand, antibodies that have been tagged with fluorescents that are detected with a fluorescence imaging system can be used (Thermo-Fisher Scientific 2015).

The tray lid was discarded and the dish containing membrane was put on the rocking platform for 15 minutes, washing the blot three times with 10 mL of TBS Tween at 5-minute intervals. Following the removal of all liquid from the dish at the end of the wash 10 mL of secondary antibody (goat anti-rabbit IgG HRP) diluted in milk blocking solution were added. Afterwards, the blot was incubated on the rocking platform for 20 minutes. During this incubation period the coomassie blue stained gel was placed on white paper for clear visibility and a photo was taken to document its appearance. This gel was previously incubated in Coomassie blue for approximately 1 hour. The gel was then discarded in the biohazard bag. After removing the membrane from the rocking platform the secondary antibody solution was removed from the membrane and discarded. Then 10 mL of TBS-Tween were added to the blot and the tray was manually shaken back and forth to remove leftover milk blocking solution and secondary antibody, which was immediately poured off. The blot was washed three times with 10 mL of TBS-Tween, at 5-minute intervals using the rocking platform. Upon completion of the washes a picture of the membrane was taken. All liquid was removed

Traditional reagent strip testing for protein uses the principle of the protein error of indicators to produce a visible colorimetric reaction. The protein (primarily albumin) accepts hydrogen ions from the indicator causing change in color. (Susan King Strasinger, 2008)

The testing of various proteins was performed by comparing the molecular weight of proteins using SDS PAGE. The molecular focus in the lab was the testing of proteins, which are macromolecules consisting of amino acid monomers linked through chemical bonds. These proteins have a hierarchy of structure that consists of folding that determines the direct function of each protein.. The molecular weight of these proteins were measured using SDS PAGE. SDS PAGE stands for sodium dodecylsulfate polyacrylamide gel electrophoresis. SDS is an anion detergent that binds with the protein structures and causes them to separate due to the change in bonding charge. SDS and heat are how the proteins are denatured. The process of denaturing a protein is breaking

Many patients are dying of prostate cancer as standard treatments are not providing the necessary results. There are new types of immunotherapy drugs which are known to work miracles for several forms of cancer. The probability of this drug helping those with prostate cancer is extremely small. There has yet to be evidence collected about the benefits and pitfalls of the treatment. If doctors were to test it on patients outside of a clinical trial, then that could be up for debate within the medical community. The drugs can have potentially deadly side effects including liver failure and nerve damage, but most patients only experience minor problems. Doctors are able to determine from biomarkers if immunotherapy treatment will help patients, but that testing is not completely accurate. Some doctors believe that they should try every possible

A majority of the population believe that vaccinations have a chance of diagnosing them with other diseases or unwanted side effects that could hinder them. There are both issues with taking vaccinations and avoiding vaccinations.

The purpose of this lab was to purify and test a GFP protein via several laboratory methods for the purpose of purifying and testing the protein in SDS-PAGE. To purify the protein chromatography and gel electrophoresis were the methods used in the experiment. GFP in the samples were tested using an ultraviolet light. When GFP was found present the cell were transformed into a petri dish containing ampicillin and arabinose. The cells were then lysed and SDS-PAGE was used to test.

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

Chinchar, V. G., Wang, J., Murti, G., Carey, C., & Rollins-Smith, L. (2001). Inactivation of frog

One is not likely to remember being vaccinated as a child, but those who have been vaccinated are able to benefit from their parents’ choice. Prior to the measles, mumps, and rubella (MMR) vaccine there were hundreds of thousands of incidents of these diseases, but now there are barely any (Colorado Children’s Immunization Coalition [CCIC], n.d. para 3). It is necessary to vaccinate children against measles, mumps, and rubella (MMR) in order to protect against these serious diseases, and to prevent the possibility of an outbreak. Furthermore, the benefits of being vaccinated outweigh the risks.

Some T lymphocytes (T cells), unlike B cells are able to attack targeted antigens directly and neutralise their effect on the body. Some send chemical messages to the rest of the immune system which help produce effective defences against the bacteria or virus, and some T cells even help B cells produce antibodies.

concentration, record the absorbance readings at a fixed wavelength, and plot the absorbance vs. concentration data. The wavelength of 520 nm was selected for experiment Part

Western Blotting can be used to detect the Myosin actin light chain in different species of fish and is used to distinguish from different species based on variation, commonality, or evolutionary divergence. First, proteins are extracted from the tissue and loaded into a gel matrix. The matrix will separate the proteins according to size using an electric current. Proteins that are separated after are blotted from the gel and onto a paper membrane. An antibody is then added to the membrane paper and causes a colored reaction. Following the reaction, the results