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CELL MOL LAB Report 10
organic- bio chemistry (CHEM153L)
Xavier University - Ateneo de Cagayan
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CELL AND MOLECULAR BIOLOGY
LABORATORY
Enzyme-linked
Immunosorbent Assay
(ELISA)
GROUP 2:
ERMINO, Andrea Alexa Joan V.
MORALES, Kaye S.
Sanchez, Shellou Grace G.
Experiment date: August 29, 2019
Submission date: September 5, 2019
Submitted to:
Mr. Paolo Antonio S. Fudalan
INTRODUCTION
In the course of advancement in science and technology, scientists introduced more profound findings and explorations in terms of diagnosis. They have applied the basic principles of anti-body mediated immunity to an assay for detecting infection by specific organisms. This assay is called an ELISA. ELISA stands for Enzyme-linked Immunosorbent Assay. It is based on the principle that antibodies produced in response to pathogens attached to their antigen targets with great specificity to form antigen- antibody complexes. The enzyme-linked assay is a commonly used analytical biochemistry assay, first described by Engvall as one antibody with great particularity for an antigen. The sample with an unknown amount of antigen is immobilized on a solid support either non-specifically or specifically.
There are two types of ELISA tests - direct ELISA and indirect ELISA. Indirect ELISA is used to detect infection or diseases through the testing of a patient's blood for the presence or absence of antibodies against a particular antigen. The presence of such antibodies indicates that the individual has been infected and that the body has launched an immune response against the disease-causing agent.
This experiment on ELISA helps in the detection of antibodies present in the simulated samples of random patients from A to F. This antibodies are antibodies that aids in the diagnosis if that certain patient is diagnosed with a human immuno deficiency virus or what is common known as HIV. If the sample, containing the chemicals or reagents, is added with chromagen and will turn its color to violet then, it would indicate that the patient is positive of HIV. This simply states that there are antibodies for HIV acting upon the patient.
The objective of this activity is to have a simulation on the actual experience of utilizing ELISA. It also helps in the distinguishing of antibodies in a given sample that could be a way for one to detect diseases present in the body.
14. Antigen-antibody complexes formed in the initial steps of and ELISA are not visible to the unaided eye. Therefore, a colorimetric detection system involving a secondary antibody and chromagen is employed. The secondary antibody, which is conjugated to an enzyme, recognizes and binds to primary antibodies of antigen-antibody complexes if they are present. Chromagen substrates is then added; if present, the enzyme linked to the secondary antibody changes the color of the chromogen. A color change will then appear to signify if the sample provided was positive or negative.
15. Performing three identical trials ensure to get rid or rule out of any false negatives or positives. Another is to justify that there are still the presence of antigens in the samples.
17. The human immunodeficiency viruses (HIV) are two species of Lentivirus (a subgroup of retrovirus) that causes HIV infection and over time acquired immunodeficiency syndrome (AIDS). AIDS is a condition in humans in which progressive failure of the immune system allows life-threatening opportunistic infections and cancers to thrive. Without treatment, average survival time after infection with HIV is estimated to be 9 to 11 years, depending on the HIV subtype.
In most cases, HIV is a sexually transmitted infection and occurs by contact with or transfer of blood, pre-ejaculate, semen, and vaginal fluids. Research has shown (for both same-sex and opposite-sex couples) that HIV is untransmissible through condomless sexual intercourse if the HIV-positive partner has a consistently undetectable viral load. Non-sexual transmission can occur from an infected mother to her infant during pregnancy, during childbirth by exposure to her blood or vaginal fluid, and through breast milk. Within these bodily fluids, HIV is present as both free virus particles and virus within infected immune cells.
18. According to the Department of Health and Human Services, to reduce the risk of HIV infection is to use condoms correctly, don’t inject drugs and if so, use only sterile injection equipment and water and never share your equipment with others. If not an HIV positive but are at high risk of becoming infected with HIV, talk to your health care provider about pre-exposure prophylaxis (PrEP). PrEP involves taking a specific HIV medicine every day to reduce the risk of HIV infection. For those who are not yet
infected, scholars and experts of the study highly encourage the people to follow the ABCD steps. First, abstain from casual sex, Second, Be Mutually-Faithful to your partner. Third, Condom Usage and Fourth, Detect Early. People who are infected already are undergoing treatment and taking medicines as prescribed by the doctors. They should keep their body healthy as they are prone to different diseases. Moreover, they should also stay in a place where they are safe from any harm or transmission of diseases either due to place or people.
Results
Before we started the experiment, we, the analysts inferred that Patients A, D and F are positive patients while Patients B, C, and E are negative for the infection. And here are the results of the experiments.
SAMPLE COLOR TEST RESULT
Postive Control Purple Positive Negative Control No Color Negative Patient A Purple Positive Patient B No Color Negative Patient C No Color Negative Patient D Purple Positive Patient E No Color Negative Patient F Purple Positive
directed against the protein to be measured. ELISA has been used as a diagnostic tool in medicine, plant pathology, and biotechnology, as well as a quality control check in various industries.
In the simplest form of an ELISA, antigens from the sample are attached to a surface. Then, a matching antibody is applied over the surface so it can bind to the antigen. This antibody is linked to an enzyme, and in the final step, a substance containing the enzyme's substrate is added. The subsequent reaction produces a detectable signal, most commonly a color change.
GENERALIZATION
ELISA (Enzyme-linked assay) is a helpful tool especially in the field of medicine. Since it can detect infectious diseases. With that, scientists and doctors will be given an idea on how to improve their works and studies. But the test has limitations, it can be true positive, false negative and false positive results. Commonly, the results it produces are seconded by other tests to confirm its validity. ELISA is often used as a screening tool before more in-depth tests are ordered.
HIV is a virus that targets and alters the immune system, increasing the risk and impact of other infections and diseases. Without treatment, the infection might progress to an advanced disease stage called AIDS. However, modern advances in treatment mean that people living with HIV in countries with good access to healthcare very rarely develop AIDS once they are receiving treatment.
AIDS can open the door to a range of infections known as opportunistic infections that pose a severe risk to health. Some are extreme or prolonged presentations of infections that would normally resolve quickly in a person with healthy immune function. And here's ELISA that helps a lot in determining this infectious disease that can harm many people.
REFERENCES
- Sinco, A. Cell and Molecular Biology Lab Manual
ELISA. 2019 Sep 4.
Overview of ELISA.
Prevent HIV with ABCD.
The Basics of HIV Prevention Understanding HIV/AIDS. 2019 Apr 29.
Picture 1. A Untested samples provided
Picture 1 Results of the experiment proper
CELL MOL LAB Report 10
Course: organic- bio chemistry (CHEM153L)
University: Xavier University - Ateneo de Cagayan
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