Immunological Methods Used in Biology Labs

Immunological Methods Used in Biology Labs
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  • 0:04 A Fishing Example
  • 0:42 Antigen-Antibody Interactions
  • 1:39 First Method: Western Blotting
  • 2:58 Second Method:…
  • 3:57 Third Method:…
  • 4:50 Lesson Summary
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Lesson Transcript
Instructor: Angela Hartsock

Angela has taught college Microbiology and has a doctoral degree in Microbiology.

In this lesson, we will discuss immunological methods used in cellular and molecular biology labs. These methods are based on antigen-antibody interactions and include western blots, immunofluorescence, and immunohistochemistry.

A Fishing Expedition

Imagine you're planning a little fishing outing to your favorite pond. But this time you're not taking the usual laid back approach; you are on a mission. You want to catch a specific fish. You don't just want to catch a largemouth bass; you want to catch a 20-inch largemouth bass named Fred with a nick in his left fin. You do not want your hook to land in the mouth of any other fish out there. You dream of a bait that only Fred would bite on, saving you a lot of time and frustration.

Of course, this is only a dream. In the biology lab, though, that's exactly what we do when we are fishing for proteins. We have a specific bait for a specific target.

Antigen-Antibody Interactions

Let's add some details to this idea of a specific bait for a specific target. Antigens are molecules, usually proteins or parts of proteins, that trigger an immune response. The immune system responds by producing antibodies, or proteins that will stick to the antigens. So antigens trigger the production of antibodies, and antibodies attack or attach to the antigens.

When you want to keep track of a protein, it's like that fishing expedition. There are tons of proteins, but if you have an antibody that goes to your protein of interest, you have that dream specific bait. You can now exploit antigen-antibody interactions as an experimental tool.

Here we will discuss three common techniques using antigen-antibody fishing. These methods are all used in the lab for scientific research, but they are also used to diagnose or detect many diseases and infections, such as HIV, rabies, influenza, Lyme disease, autoimmune disorders, and cancer.

First Method: Western Blotting

Western blotting is a method that extracts proteins from a cell, separates them, adds an antibody that binds to the protein of interest, and then triggers a visible chemical reaction that highlights the bound antibody.

Let's walk through the technique. In the lab, you start with a cell or tissue sample in which you want to confirm the presence of a specific protein. First, extract all the proteins from your sample. At this point, you have a little tube containing fluid and all the proteins mixed together. Next, get them all spread out using gel electrophoresis, which separates the proteins based on size. Now you have to figure out if your protein is there. Now transfer the proteins from the gel where they are embedded inside onto the surface of a special membrane where the antibodies can get access to them. Then, take the antibody that fits your protein and apply it to the membrane. If the antibody finds the protein, it will bind to it.

Now, you might think this is the last step, but even after the antibody attaches to the protein, it's still completely invisible to your eye. Not very useful, right? Now you just need a trick to let you see what's happening. So we add a second antibody that will grab that first antibody. And, lastly, we can add a chemical to the whole membrane, and wherever the second antibody is attached, it will chop up the chemical, making it turn a bright color.

Second Method: Immunofluorescence

Immunofluorescence uses a microscope to detect fluorescently tagged antibodies that have bound to antigen. This is a great method if you don't want to extract the proteins from your cells, but rather want to see where the proteins are in your cell.

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