Antigen retrieval - Revision history

Jakob Suckale: /* External links */

Jakob Suckale — Mon, 02 Nov 2009 19:13:53 GMT

External links

←Older revision		Revision as of 19:13, 2 November 2009
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	* [[Image:3stars.png]] [http://www.ihcworld.com/_technical_tips/antigen_retrieval_tips.htm detailed description of antigen retrieval methods at IHCWorld]		* [[Image:3stars.png]] [http://www.ihcworld.com/_technical_tips/antigen_retrieval_tips.htm detailed description of antigen retrieval methods at IHCWorld]
	* [[Image:3stars.png]] [http://www.ncbi.nlm.nih.gov/pubmed/17197287 Yamashita 2007] - 60 page detailed review article including antibody generation, fixation, and details on several antigen retrieval techniques		* [[Image:3stars.png]] [http://www.ncbi.nlm.nih.gov/pubmed/17197287 Yamashita 2007] - 60 page detailed review article including antibody generation, fixation, and details on several antigen retrieval techniques
		+	* [http://www.nordiqc.org/Techniques/Epitope_retrieval.htm review of retrieval methods at NordiQC and examples of which antibodies work with which method]
	* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]		* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]
	* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]		* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]

Jakob Suckale: /* Types of retrieval methods */

Jakob Suckale — Mon, 19 Oct 2009 15:40:11 GMT

Types of retrieval methods

←Older revision		Revision as of 15:40, 19 October 2009
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	== Types of retrieval methods ==		== Types of retrieval methods ==
-	# Heat treatment ~~- mechanism~~: heat cleaves cross-links, exposes buried epitopes of proteins	+	=== Heat treatment ===
-	# Protease treatment	+	* Mechanism: heat cleaves cross-links, exposes buried epitopes of proteins, protein unfolding/refolding
-	# Detergent treatment	+	* Specific protocol 1: citrate buffer [http://openwetware.org/wiki/Livesey:_Antigen_retrival][http://www.ihcworld.com/_protocols/epitope_retrieval/citrate_buffer.htm]
		+	* Specific protocol 2: Tris-EDTA buffer [http://www.ihcworld.com/_protocols/epitope_retrieval/tris_edta.htm]
		+
		+	=== Protease treatment ===
		+	* Specific protocol 1: proteinase K [http://www.ihcworld.com/_protocols/epitope_retrieval/proteinase-k.htm]
		+	* Specific protocol 2: trypsin [http://www.ihcworld.com/_protocols/epitope_retrieval/trypsin.htm]
		+
		+	=== Detergent treatment ===
		+	* Specific protocol: SDS [http://openwetware.org/wiki/Griffin:Immunofluorescence_Cell_Staining#Antigen_retrieval_for_cryostat_tissue_sections_or_cultured_cells]

	== Personal and lab-specific protocols ==		== Personal and lab-specific protocols ==

Jakob Suckale: removed detailed protocol from this more general page (protocol is still linked)

Jakob Suckale — Mon, 19 Oct 2009 15:32:49 GMT

removed detailed protocol from this more general page (protocol is still linked)

←Older revision		Revision as of 15:32, 19 October 2009
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	# Protease treatment		# Protease treatment
	# Detergent treatment		# Detergent treatment
-
-	~~== Sample heat-treatment protocol using a citrate buffer ==~~
-	~~# Choose desired slides (paraformaldyhyde-fixed, 10 micron thick crysections).~~
-	~~# Leave the chosen slides at room temperature just until the frost melts.~~
-	~~# Hydrate slides in 1x PBS for 10 seconds.~~
-	~~# Put slides into '''0.01 M Citric acid (pH 6.0), microwave''' just to a boil, and let cool for 5 minutes.~~
-	~~# Repeat Step 4 two more times (three times total).~~
-	~~# Rinse the slides 6 times in 1x TBST for 8 minutes each.~~
-	~~# Block with 5% milk powder in 1x TBST for 30 minutes at room temperature. Optional: 1-2% BSA can be added to TBST.~~
-	~~# Incubate overnight at 4oC with primary antibodies in TBST plus 5% milk.~~
-	~~# Rinse for 10 seconds, 1x TBST. Then, rinse 3 times in 1x TBST for 8 minutes each.~~
-	~~# Add secondary antibodies in TBST for 1 hour at room temperature in the dark.~~
-	~~# Rinse 3 times in 1x TBS for 8 minutes each.~~
-	~~# Add Fluorescent Mounting Medium, coverslip, and examine under a fluorescent microscope.~~
-	~~# Lastly, seal the edges of covered-slip slides with a clear nail polish. This helps slides retain fluorescence longer during storage.~~
-	~~TBST = TBS + 0.1% Triton~~

	== Personal and lab-specific protocols ==		== Personal and lab-specific protocols ==

Jakob Suckale: /* External links */

Jakob Suckale — Mon, 19 Oct 2009 15:24:44 GMT

External links

←Older revision		Revision as of 15:24, 19 October 2009
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	== External links ==		== External links ==
	* [[Image:3stars.png]] [http://www.ihcworld.com/_technical_tips/antigen_retrieval_tips.htm detailed description of antigen retrieval methods at IHCWorld]		* [[Image:3stars.png]] [http://www.ihcworld.com/_technical_tips/antigen_retrieval_tips.htm detailed description of antigen retrieval methods at IHCWorld]
		+	* [[Image:3stars.png]] [http://www.ncbi.nlm.nih.gov/pubmed/17197287 Yamashita 2007] - 60 page detailed review article including antibody generation, fixation, and details on several antigen retrieval techniques
	* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]		* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]
	* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]		* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]

Jakob Suckale: /* Types of retrieval methods */

Jakob Suckale — Mon, 19 Oct 2009 15:21:19 GMT

Types of retrieval methods

←Older revision		Revision as of 15:21, 19 October 2009
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	== Types of retrieval methods ==		== Types of retrieval methods ==
-	# Heat treatment	+	# Heat treatment - mechanism: heat cleaves cross-links, exposes buried epitopes of proteins
	# Protease treatment		# Protease treatment
	# Detergent treatment		# Detergent treatment

Jakob Suckale: /* External links */

Jakob Suckale — Mon, 19 Oct 2009 15:02:14 GMT

External links

←Older revision		Revision as of 15:02, 19 October 2009
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	== External links ==		== External links ==
		+	* [[Image:3stars.png]] [http://www.ihcworld.com/_technical_tips/antigen_retrieval_tips.htm detailed description of antigen retrieval methods at IHCWorld]
	* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]		* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]
	* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]		* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]

Jakob Suckale: external links

Jakob Suckale — Mon, 19 Oct 2009 14:59:44 GMT

external links

←Older revision		Revision as of 14:59, 19 October 2009
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	* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]		* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]
	* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]		* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]
		+	Commercial antigen retrieval kits:
		+	* [http://www.bdbiosciences.ca/ptProduct.jsp?prodId=19647&catyId=76670 BD Pharmigen Retrievagen A pH 6] and [http://www.bdbiosciences.ca/ptProduct.jsp?prodId=19653&catyId=76670 BD Pharmigen Retrievagen B pH 9.5]
		+	* [http://www.genetex.com/WebPage/Product/ProductDetail.aspx?CatalogItemID=33939 GeneTex antigen retrieval solution (heat-based)] and [http://www.genetex.com/WebPage/Product/ProductDetail.aspx?CatalogItemID=66695 GeneTex antigen retrieval solution (protease-based)]
		+	* [http://www.abcam.com/10x-Heat-Mediated-Antigen-Retrieval-Solution-pH-6-0-ab973.html abcam heat-based antigen retrieval solution 10x]
		+	* [http://www.biocompare.com/ProductListings/17837/Antigen-Retrieval-Solutions.html antigen retrieval search at biocompare]

	[[Category:Protocol]]		[[Category:Protocol]]

Jakob Suckale at 14:46, 19 October 2009

Jakob Suckale — Mon, 19 Oct 2009 14:46:23 GMT

←Older revision		Revision as of 14:46, 19 October 2009
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	# Detergent treatment		# Detergent treatment

-	== ~~Protocol~~ ==	+	== Sample heat-treatment protocol using a citrate buffer ==
	# Choose desired slides (paraformaldyhyde-fixed, 10 micron thick crysections).		# Choose desired slides (paraformaldyhyde-fixed, 10 micron thick crysections).
	# Leave the chosen slides at room temperature just until the frost melts.		# Leave the chosen slides at room temperature just until the frost melts.

Jakob Suckale at 14:40, 19 October 2009

Jakob Suckale — Mon, 19 Oct 2009 14:40:57 GMT

←Older revision		Revision as of 14:40, 19 October 2009
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	TBST = TBS + 0.1% Triton		TBST = TBS + 0.1% Triton

-	== ~~Lab~~-specific protocols ==	+	== Personal and lab-specific protocols ==
	* Korey Griffin's notes:		* Korey Griffin's notes:
	:* [[Griffin:Antigen Retrieval Technique]] - overview and comparison of techniques		:* [[Griffin:Antigen Retrieval Technique]] - overview and comparison of techniques

Jakob Suckale: external links

Jakob Suckale — Mon, 19 Oct 2009 14:40:19 GMT

external links

←Older revision		Revision as of 14:40, 19 October 2009
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	* [[Antibodies]]		* [[Antibodies]]
	* [[Griffin:Antibody Basics]]		* [[Griffin:Antibody Basics]]
		+
		+	== External links ==
		+	* [http://www.bioreagents.com/pages/protocols.cfm#3\|Antigen retrieval protocols using heat at Thermo]
		+	* [http://www.rockland-inc.com/commerce/misc/Antigen_retrieval_methods.jsp\|Heat and protease antigen retrieval at Rockland, Inc.]

	[[Category:Protocol]]		[[Category:Protocol]]