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a-Satellite probes to chromosomes 1, 15 and 17 were labeled by nick translation with biotin-11-dUTP, ChromaTide Texas Red-12-dUTP (C7631) and ChromaTide Oregon Green 488-5-dUTP, respectively. Following simultaneous hybridization of all three probes, the biotinylated chromosome 1 probe was detected with HRP–streptavidin conjugate and Alexa Fluor 546 tyramide (TSA Kit ##23, Cat. No. T20933). HRP activity from this first TSA detection step was then quenched by treatment with 1% hydrogen peroxide for 30 minutes. Lastly, the Oregon Green 488 dye–labeled chromosome 17 probe was detected with anti–fluorescein/Oregon Green antibody (Cat. No. A6421) followed by HRP-conjugated goat anti–mouse IgG antibody and Alexa Fluor 594 tyramide (TSA Kit ##5, Cat. No. T20915). HRP activity from this second TSA detection step was then quenched by treatment with 1% hydrogen peroxide for 30 minutes. The Texas Red dye–labeled chromosome 15 probe was then detected with rabbit anti–Texas Red antibody (Cat. No. A6399) followed by HRP-conjugated goat anti–rabbit IgG antibody and Alexa Fluor 488 tyramide (TSA Kit ##12, Cat. No. T20922). After counterstaining with Hoechst 33258 (Cat. No. H1398, H3569, H21491), the images were acquired using filters appropriate for DAPI, FITC, TRITC and Texas Red dyes.
|Tested species reactivity||Chem|
|Published species reactivity||Not Applicable|
|Host / Isotype||Rabbit / IgG|
|Storage buffer||PBS, pH 7.2|
|Contains||5mM sodium azide|
|Storage Conditions||4° C|
|Tested Applications||Dilution *|
|Immunocytochemistry (ICC)||Assay Dependent|
|Immunohistochemistry (IHC)||Assay Dependent|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
|Immunohistochemistry (Paraffin) (IHC (P))||See 1 publications below|
Unlike biotin, which is an endogenous ligand in mitochondria, dye-based haptens permit background-free staining of cells and tissues and a sensitive alternative to biotin-streptavidin methods.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Implementation of TMA and digitalization in routine diagnostics of breast pathology.
A-6399 was used in immunohistochemistry - paraffin section to study the implementation of tissue microarrays and digitalization in routine diagnosis of breast pathology.
|Rossing HH,Talman ML,Laenkholm AV,Wielenga VT||APMIS : acta pathologica, microbiologica, et immunologica Scandinavica (120:341)||2012|