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Reseptör İmmünohistokimyasında Mikrodalga Işınımlı “Antijen Retrieval” Yönteminin Kullanımı

Usage of Microwave Stimulated Antigen Retrieval Method for Receptor Immunohistochemistry

Journal Name:

  • Uludağ Üniversitesi Tıp Fakültesi Dergisi

Publication Year:

  • 2002

Key Words:

Keywords (Original Language):

Author NameUniversity of Author
Abstract (2. Language): 
To solve the insufficiency or masking of the antigen that is one of the important problem of the receptor immunohistochemistry, antigen retrieval (AR) techniques have been employed on the tissue blocks or sections. In this study, we assessed if the usage of the stimulation of AR technique by microwave irradiation (MW–AR) is necessary for the antibody which is specific to the kainate receptor subunit GluR5, and if so, which AR solution was appropriate. For this aim, rat brain tissue was fixed via perfusion and 50 µm –thick vibrotome sections were collected. Following the MW– AR procedures in either of two different freshly prepared solutions (50 mM trisodium citrate solution, pH 4.5 and 6.0 or 10 mM EDTA solution, pH 8.0), floating section immunohistochemistry was employed on the sections. Light microscopic analyses revealed that the sections which were treated with 50 mM trisodium citrate solution (pH 6.0) possessed the best immunolabeling and no background staining. In conclusion, it was suggested that the AR technique is necessary for the anti–GluR5 antibody used in this study and that the pH and the final temperature of the solution is important for a specific immunoreaction.
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Abstract (Original Language): 
Reseptör immünohistokimyasında önemli problemlerden biri olan antijen miktarının azlığı veya maskelenmesi sorununun giderilmesi amacıyla, doku ya da kesitlere; antijen retrieval (AR) yöntemleri uygulanmaktadır. Çalışmamızda, pek çok antijen için başarılı sonuçlar verdiği bilinen mikrodalga ışınımlı AR (MW–AR) yönteminin kainat reseptör altünitesi GluR5’e özgün antikor (anti-GluR5) için gerekli olup olmadığı, gerekli ise hangi AR solüsyonunun uygun olabileceği araştırıldı. Bu amaçla, perfüzyon yoluyla fikse edilen sıçan beyin dokusundan 50 µm kalınlığında vibratom kesitleri alındı. Kesitlere, taze hazırlanmış iki farklı solusyonda (50 mM trisodyum sitrat solüsyonu, pH 4.5 ve 6.0 ya da 10 mM EDTA solüsyonu, pH 8.0) MW–AR işlemini izleyen, yüzen kesit immünohistokimyası uygulandı. Işık mikroskobik incelemelerde, 50 mM trisodyum sitrat solüsyonu (pH 6.0) ile AR uygulanan kesitlerde özgün immün işaretlemenin olduğu ve bu kesitlerde zemin boyanmasının olmadığı gözlendi. Sonuç olarak, kullanılan antikor için AR’ın gerektiği ve özgün bir immün reaksiyon için AR solüsyonunun pH’nın ve erişilen son sıcaklığının önemli olduğu belirlendi.
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  • 2
21-26
  • Turkish

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