Immunocytochemistry: Difference between revisions
Content deleted Content added
m Reverting possible vandalism by Roguehawkx30 to version by Skittleys. False positive? Report it. Thanks, ClueBot. (771859) (Bot) |
Roguehawkx30 (talk | contribs) ←Blanked the page |
||
| Line 1: | Line 1: | ||
{{unreferenced|date=September 2009}} |
|||
[[Image:immunohistochemistry.png|thumb|Immunocytochemistry labels individual proteins within cells, such as [[Tyrosine hydroxylase|TH]] (green) in the [[axons]] of sympathetic [[Autonomic Nervous System|autonomic]] neurons.]] |
|||
{{Expert-subject|Molecular and Cellular Biology|date=February 2009}} |
|||
'''Immunocytochemistry''' ('''ICC''') is a common [[laboratory technique]] that uses [[antibodies]] that target specific [[peptide]]s or [[protein]] [[antigen]]s in the [[cell (biology)|cell]] via specific [[epitope]]s. These [[bound antibodies]] can then be detected using several different [[Protein methods|methods]]. ICC allows researchers to evaluate whether or not cells in a particular sample [[protein expression|express]] the antigen in question. In cases where an [[wikt:immunopositive|immunopositive]] signal is found, ICC also allows researchers to determine which [[sub-cellular compartment]]s are expressing the antigen. |
|||
==Immuno''cyto''chemistry vs. immuno''histo''chemistry== |
|||
Immunocytochemistry differs from [[immunohistochemistry]] in that the former is performed on [[sample (material)|samples]] of intact cells that have had most, if not all, of their surrounding [[extracellular matrix]] removed. This includes cells grown within a [[cell culture|culture]], deposited from [[suspension]], or taken from a [[smear]]. In contrast, immunohistochemical samples are sections of [[tissue (biology)]], where each cell is surrounded by tissue architecture and other cells normally found in the intact tissue. |
|||
==Methods== |
|||
{{Details|immunohistochemistry|immunochemical methods}} |
|||
There are many methods to obtain immunological detection on tissues, including those tied directly to primary antibodies or antisera. A direct method involves the use of a detectable tag (e.g., fluorescent molecule, gold particles, etc., ) directly to the antibody that is then allowed to bind to the antigen (e.g., protein) in a cell. |
|||
Alternatively, there are many '''indirect methods'''. In one such method, the antigen is bound by a primary antibody which is then amplified by use of a [[secondary antibody]] which binds to the primary antibody. Next, a tertiary reagent containing an enzymatic moiety is applied and binds to the secondary antibody. When the quarternary reagent, or substrate, is applied, the enzymatic end of the tertiary reagent converts the substrate into a pigment reaction product, which produces a color (many colors are possible; brown, black, red, etc.,) in the same location that the original primary antibody recognized that antigen of interest. |
|||
Some examples of '''substrates''' used (also known as chromogens) are AEC (3-Amino-9-EthylCarbazole), or DAB ([[3,3'-Diaminobenzidine tetrahydrochloride|3,3'-Diaminobenzidine]]). Use of one of these reagents after exposure to the necessary enzyme (e.g., horseradish peroxidase conjugated to an antibody reagent) produces a positive immunoreaction product. Immunocytochemical visualization of specific antigens of interest can be used when a less specific stain like H&E (Hematoxylin and Eosin) cannot be used for a diagnosis to be made or to provide additional predictive information regarding treatment (in some cancers, for example). |
|||
Alternatively the secondary antibody may be covalently linked to a [[fluorophore]] ([[Fluorescein isothiocyanate|FITC]] and [[Rhodamine]] are the most common) which is detected in a fluorescence or confocal microscope. The location of fluorescence will vary according to the target molecule, external for membrane proteins, and internal for cytoplasmic proteins.In this way [[immunofluorescence]] is a powerful technique when combined with [[confocal microscopy]] for studying the location of proteins and dynamic processes ([[exocytosis]], [[endocytosis]], etc). |
|||
==References== |
|||
{{reflist}} |
|||
==External links== |
|||
* [http://www.prosci-inc.com/Immunocytochemistry-Protocol.html Immunocytochemistry Staining Protocol] |
|||
*[http://www.cshprotocols.org/cgi/content/full/2008/2/pdb.prot4820 Immunohistochemistry of Whole-Mount Mouse Embryos] |
|||
{{Immunologic techniques and tests}} |
|||
[[Category:Histology]] |
|||
[[Category:Immunology]] |
|||
[[Category:Protein methods]] |
|||
[[Category:Staining]] |
|||
[[Category:Laboratory techniques]] |
|||