A practical strategy for immunofluorescent detecting multiple targets in mouse tissues without restrictions on the host specious resources of the primary antibodies

Shangyi Yu, Xi He, Aleksandra Drelich, Barbara Judy, Qing Chang, Shuhui Cao, Thomas Ksiazek, Zhiyun Xu, Bin Gong

Research output: Contribution to journalArticle

Abstract

Commercial deficiency of practical system to label multiple targets in experimental mouse tissues significantly hinders the feasibility to study the potential association between/among multiple targets using tissue-based immunofluorescence (IF) staining. We have developed a new protocol to do dual – labeling immunofluorescences on mouse tissues by combining direct and indirect immunofluorescence, making it possible to use commercial antibodies from the same specious (rabbit) to detect multiple targets in formalin-fixed paraffin-embedded (FFPE) archival mouse tissues simultaneously. This method applies indirect immunofluorescence to assess the first antigen in mouse tissues by using a rabbit anti-mouse polyclonal antibody and goat anti-rabbit antibody. After that, normal rabbit serum was employed to blocking the free binding sites of the previous antibodies. Direct immunofluorescence was used to assess the second antigen by a commercial kit-labeled rabbit anti–human (mouse) antibody at different emission wavelength. At last, cell nuclei were co-stained by DAPI. The outcomes demonstrated that this protocol obtain promising signals of both antigens and the nuclei. Moreover, this method also works on infection disease models in which samples are often over fixed due to biosafety rules.

Original languageEnglish (US)
JournalPathology Research and Practice
DOIs
StatePublished - Jan 1 2019

Fingerprint

Antibodies
Rabbits
Direct Fluorescent Antibody Technique
Indirect Fluorescent Antibody Technique
Antigens
Fluorescent Antibody Technique
Antibody Binding Sites
Feasibility Studies
Cell Nucleus
Goats
Paraffin
Formaldehyde
Anti-Idiotypic Antibodies
Staining and Labeling
Infection
Serum

Keywords

  • Antibody
  • Immunofluorescent staining
  • Strategy

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Cell Biology

Cite this

A practical strategy for immunofluorescent detecting multiple targets in mouse tissues without restrictions on the host specious resources of the primary antibodies. / Yu, Shangyi; He, Xi; Drelich, Aleksandra; Judy, Barbara; Chang, Qing; Cao, Shuhui; Ksiazek, Thomas; Xu, Zhiyun; Gong, Bin.

In: Pathology Research and Practice, 01.01.2019.

Research output: Contribution to journalArticle

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AU - Chang, Qing

AU - Cao, Shuhui

AU - Ksiazek, Thomas

AU - Xu, Zhiyun

AU - Gong, Bin

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