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Optimering av enzym-baserad immunohistokemisk metod i jämförelse mot immunofluorescens med fryssnittade hudbiopsier
Jönköping University, School of Health and Welfare, HHJ, Dept. of Natural Science and Biomedicine.
2022 (Swedish)Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesisAlternative title
Optimization of enzyme-based immunohistochemical method in comparison with immunofluorescence with frozen-cut skin biopsies (English)
Abstract [sv]

Med hjälp av immunohistokemi kan antigen och antikroppar som är bundna till vävnaden detekteras. Autoimmuna hudsjukdomar är exempel på sjukdomar som diagnosticeras med immunohistokemi. På Falu lasarett användes immunofluorescens för diagnostik av autoimmuna hudsjukdomar. Syftet med denna studie var att optimera enzym-baserad immunohistokemi för epitoperna IgA, IgG, IgM och C3 och jämföra med immunofluorescens vad gäller specificitet, signalstyrka och upplösning. Vävnader som analyserades var tonsill, lever, tarmslemhinna och hudbiopsier. Fixering gjordes i 4% formaldehyd av vävnaderna som infärgades med ultraView DAB och ultraView DAB med FITC. Vävnad som infärgades med DIF med FITC sköljdes enbart i Reaction Buffer. Vävnaderna färgades enligt protokollen ultraView DAB, ultraView DAB med FITC och DIF med FITC. En manuell infärgning med aktiverat DAB utfördes. Resultatet visade bakgrundsinfärgning för samtliga infärgningar. DIF med FITC var tydligare infärgad och lättare att skilja mellan specifik och ospecifik infärgning. Det är svårt att optimera enzym-baserad IHC och epitopen IgA hade generellt starkare infärgning jämfört med epitopen IgM. För att erhålla tillförlitliga resultat krävs det att flera vävnadsprover analyseras.

Abstract [en]

Using immunohistochemistry, antigens and antibodies bound to the tissue can be detected. Autoimmune skin diseases are examples of diseases that are diagnosed with immunohistochemistry. At Falu Hospital, immunofluorescence was used to diagnose autoimmune skin diseases. The aim of this study was to optimize enzyme-based immunohistochemistry for the epitopes IgA, IgG, IgM and C3 and to compare with immunofluorescence in terms of specificity, signal strength and resolution. Tissues analyzed were tonsil, liver, intestinal mucosa and skin biopsies. Fixation was done in 4% formaldehyde of the tissues stained with ultraView DAB and ultraView DAB with FITC. Tissue stained with DIF with FITC was rinsed in Reaction Buffer only. A manual staining with activated DAB was performed. The result showed background staining for all stainings. DIF with FITC was more clearly stained and easier to distinguish between specific and nonspecific staining. It is difficult to optimize enzyme-based IHC and the epitope IgA generally had stronger staining compared to the epitope IgM. To obtain reliable results, several tissue samples must be analyzed.

Place, publisher, year, edition, pages
2022. , p. 42
Keywords [en]
Ventana BenchMark ULTRA System, Fluorescein Isothiocyanate, Diaminobenzidine
Keywords [sv]
Ventana BenchMark ULTRA System, Fluorescein Isothiocyanat, Diaminobenzidin
National Category
Biomedical Laboratory Science/Technology
Identifiers
URN: urn:nbn:se:hj:diva-57143ISRN: JU-HHJ-BLA-1-20220126OAI: oai:DiVA.org:hj-57143DiVA, id: diva2:1668946
Subject / course
HHJ, Biomedical Laboratory Science
Available from: 2022-06-15 Created: 2022-06-13 Last updated: 2025-10-13Bibliographically approved

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