Immune electron microscopy
Immune electron microscopy (more often called immunoelectron microscopy) is the equivalent of
Process
Antigens and their respective antibodies (usually two) interact in the section.[1] Transmission electron microscopy then detects the antibody and, therefore, the protein. The second antibody is typically bound to gold because gold has a high atomic number, making it very dense. Colloidal gold particles make the antibodies visible by conjugating with them, because their exact diameter is known.[3] When electrons pass through the microscope, they hit this gold particle. The dense gold atom reflects the electrons being emitted from the electron microscope and causes the appearance of the target particle within the specimen.[1]
Another possible process involves
While using immune electron microscopy, the specimen can either be in thin sections so the electrons can penetrate it or negatively stained.
Complications and Results
Potential Complications
The sections under the microscope must be very thin to allow the electrons to pass through. Some complications can arise during the preparation steps necessary to create the thin sections, including chemical fixation and embedding (usually in plastic). These harsh preparations can denature antigens, interrupting their necessary bond with the antibodies. Researchers have invented and utilized specific processes to circumvent these issues and preserve the interaction between the antigen and antibodies. These methods include light fixation rather than chemical fixation, freezing the specimen prior to sectioning it, and incubating it at room temperature rather than high temperatures.[1]
Bonds between antibodies and their respective antigen or between antibodies and their gold labels may be only partially secure due to the effects of low concentrations or
Results
Results from immune electron microscopy are typically quantified visually. The sample must have certain features for quantitative analysis to be effective, limiting its frequency of use. It is applicable in situations like seeing how many colloidal gold particles are attached to a particular antibody.[5] During successful experiments, immune electron microscopy can accurately locate proteins and strengthen comprehension of the relationship between structure and function. These processes in labeling and localization help researchers understand various cellular pathways and processes.[3]
History
In 1931, Ernst Ruska (1986 Nobel Prize award winner) and Max Knoll created the first electron microscope. This invention led to the scanning electron microscope and transmission electron microscope, which later contributed to immunoelectron microscopy. At first, technology only allowed for two-dimensional images, but now with modern technology, three-dimensional images are also available.[3]
Immunoelectron microscopy came about when two independent groups in the 1940s combined the tobacco mosaic virus and its antiserum. They then examined it under an electron microscope. At this time, resolution was much poorer due to a lack of additional contrast and poor quality microscopes of the day. The particles used in the experiment were known to be rod-shaped, and both groups of researchers found these rods clumping together in a group about twice their original size. More than a decade and a half later, researchers began to use singular antibodies attached to viruses. Finally, in 1962, negatively stained antibodies came out.[4]
Applications
Viruses
Transmission electron microscopy successfully provides general information about structure but struggles to differentiate more detailed parts of a
Renal Biopsies
Initially,
References
- ^ ISBN 978-1464183393.
- ^ "Immuno-Electron Microscopy Services at the Core Electron Microscopy Facility - UMASS Medical School". UMass Chan Medical School. 2 November 2013. Retrieved 5 December 2022.
- ^ a b c "Immunoelectron microscopy". The Human Protein Atlas.
- ^ S2CID 80868758. Retrieved 6 December 2022.
- ISBN 978-0-7020-6896-6. Retrieved 6 December 2022.
- S2CID 26970189. Retrieved 6 December 2022.