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content for id "main" goes here procedures home pretreatment ihc process antibodies breast estrogen receptor herceptin receptor progesterone receptor endothelial factor 13a cd 31 cd 34 epithelial light heavy pancytokeratin lymphoid t cell markers b cell markers cd 45 cd 68 non-hodgkins muscle desmin myogenin smooth muscle actin vimentin melanoma hmb 45 melan a s100 mesothelial calretinin cd 141 carcinoembryonic antigen ck 5/6 ep4 neuroendocrine chromogranin ck20 neurone specific elastase synaptophysin urological/prostatic ckhmw prostatic specific antigen prostatic specific acid phosphatase developements dualstain triplestain lung alk extra useful information suppliers ihc terms troubleshooting - what is ihc or icc? what is ? icc protocol contents or ihc refers to the process of detecting antigens in cells of a tissue section via antigen retrieval. this is carried out by exploiting the principle of antibodies binding specifically to antigens in biological tissues. immunohistochemical staining is widely used in the diagnosis of cancerous tumors from hodgkins lymphoma to differentiation of different tumours eg ductal carcinoma in the breast based on specific antigen staining. specific molecular markers are characteristic of particular cellular events such as proliferation or cell death apoptosis. ihc is also widely used in basic research to understand the distribution and localization of biomarkers and differentially expressed proteins in different parts of a biological tissue. visualising an antibody-antigen interaction can be accomplished in a number of ways. in the most common instance, an antibody is conjugated to an enzyme, such as peroxidase , that can catalyse and make the area visible. the discipline of or immunocytochemistry was known since the 1930s but it was really made possible only in 1942 when the first ihc study was reported by coons. here he used fitc-labeled antibodies to identify antigens in infected tissue. since then, improvements have been made in protein conjugation, tissue fixation methods, detection labels and microscopy, making a routine and essential tool in diagnostic and research laboratories. to carry out the procedure the general protocl used below are followed, sections cut at 3 m m and floated onto superfrost glass slides. sections are drained and then left in the 56c incubator for one hour or at room temp overnight. 1 deparaffinise paraffin sections in xylene – 5 mins. (for resin sections, de-resin in xylene overnight) 2 rinse through graded alcohols to water – 5 mins in each. 3 wash in water - 5 mins 4 block with 3% aqueous hydrogen peroxide - 10 mins 5 wash in water - 5 mins 6 apply antigen retrieval as applicable: a) microwave at full power in cc1 – 2 x 10 mins remember to top up fluid level after the first 10 minutes b) or apply pronase 1 reagent for the appropriate time in the humidity chamber 7 wash in running cold water and allow to cool – 5mins 8 wash in reaction buffer and apply dako pen around sections 9 place sections into the humidity box 10 apply sufficient prediluted primary antibody to cover the sections for the appropriate time. 11 rinse with reaction buffer – 2 x 5min 12 apply the polymer link (universal hrp multimer)- 30 mins 13 rinse with reaction buffer – 2 x 5min 14 apply the universal dab chromogen, (2 – 3 drops) 15 add the same amount of universal dab h2o2, (2 – 3 drops) to activate the chromogen 16 visualise microscopically to determine optimal staining. then wash well in water. 18 counterstain lightly in haematoxylin for about 15 seconds, then wash well in water to “blue-up” sections 19 enhance staining by placing slides in 0.5% aqueous cuso4 for 10 minutes 20 wash in water. dehydrate through graded alcohols, clear in xylene & mount in cv mount . procedures home pretreatment ihc process antibodies breast estrogen receptor herceptin receptor progesterone receptor endothelial factor 13a cd 31 cd 34 epithelial light heavy pancytokeratin lymphoid t cell markers b cell markers cd 45 cd 68 non-hodgkins muscle desmin myogenin smooth muscle actin vimentin melanoma hmb 45 melan a s100 mesothelial calretinin cd 141 carcinoembryonic antigen ck 5/6 ep4 neuroendocrine chromogranin ck20 neurone specific elastase synaptophysin urological/prostatic ckhmw prostatic specific antigen prostatic specific acid phosphatase developements dualstain triplestain lung alk extra useful information suppliers ihc terms troubleshooting home feedback histological techniques spanish

URL analysis for immunohistochemistry.org.uk


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