20: Laboratory Analysis of the Immune Response
Many laboratory tests are designed to confirm a presumptive diagnosis by detecting antibodies specific to a suspected pathogen. Unfortunately, many such tests are time-consuming and expensive. That is now changing, however, with the development of new, miniaturized technologies that are fast and inexpensive. For example, researchers at Columbia University are developing a “lab-on-a-chip” technology that will test a single drop of blood for 15 different infectious diseases, including HIV and syphilis, in a matter of minutes. 1 The blood is pulled through tiny capillaries into reaction chambers where the patient’s antibodies mix with reagents. A chip reader that attaches to a cell phone analyzes the results and sends them to the patient’s healthcare provider. Currently the device is being field tested in Rwanda to check pregnant women for chronic diseases. Researchers estimate that the chip readers will sell for about $100 and individual chips for $1. 2
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- 20.1: Practical Applications of Monoclonal and Polyclonal Antibodies
- In addition to being crucial for our normal immune response, antibodies provide powerful tools for research and diagnostic purposes. The high specificity of antibodies makes them an excellent tool for detecting and quantifying a broad array of targets, from drugs to serum proteins to microorganisms. With in vitro assays, antibodies can be used to precipitate soluble antigens, agglutinate cells, and neutralize drugs, toxins, and viruses.
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- 20.2: Detecting Antigen-Antibody Complexes in vitro
- Laboratory tests to detect antibodies and antigens outside of the body (e.g., in a test tube) are called in vitro assays. When both antibodies and their corresponding antigens are present in a solution, we can often observe a precipitation reaction in which large complexes (lattices) form and settle out of solution. In the next several sections, we will discuss several common in vitro assays.
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- 20.3: Agglutination Assays
- In addition to causing precipitation of soluble molecules and flocculation of molecules in suspension, antibodies can also clump together cells or particles (e.g., antigen-coated latex beads) in a process called agglutination. Agglutination can be used as an indicator of the presence of antibodies against bacteria or red blood cells. Agglutination assays are usually quick and easy to perform on a glass slide or microtiter plate.
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- 20.4: Enzyme Immunoassays (EIA) and Enzyme-Linked Immunosorbent Assays (ELISA)
- Enzyme immunoassays (EIA) are used to visualize and quantify antigens. They use an antibody conjugated to an enzyme to bind the antigen, and the enzyme converts a substrate into an observable end product. The substrate may be either a chromogen or a fluorogen. Immunostaining is an EIA technique for visualizing cells in a tissue (immunohistochemistry) or examining intracellular structures (immunocytochemistry). Direct ELISA is used to quantify an antigen in solution.
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- 20.5: Fluorescent Auto-Antibody Techniques
- Rapid visualization of bacteria from a clinical sample such as a throat swab or sputum can be achieved through fluorescent antibody (FA) techniques that attach a fluorescent marker (fluorogen) to the constant region of an antibody, resulting in a reporter molecule that is quick to use, easy to see or measure, and able to bind to target markers with high specificity. We can also label cells, allowing us to precisely quantify particular subsets of cells or even purify them for further research.
Footnotes
- 1 Chin, Curtis D. et al., “Mobile Device for Disease Diagnosis and Data Tracking in Resource-Limited Settings,” Clinical Chemistry 59, no. 4 (2013): 629-40.
- 2 Evarts, H., “Fast, Low-Cost Device Uses the Cloud to Speed Up Testing for HIV and More,” January 24, 2013. Accessed July 14, 2016. http://engineering.columbia.edu/fast...g-hiv-and-more .
Thumbnail: Enzyme-linked antibodies against CD8 were used to stain the CD8 cells in this preparation of bone marrow using a chromogen. (credit: modification of work by Yamashita M, Fujii Y, Ozaki K, Urano Y, Iwasa M, Nakamura S, Fujii S, Abe M, Sato Y, Yoshino T).