Premium
A Simple Approach for Multicolor Immunofluorescence Staining in Different Drosophila Cell Types
Author(s) -
Cipressa Francesca,
Di Giorgio Maria Laura,
Cenci Giovanni
Publication year - 2014
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.24506
Subject(s) - immunostaining , biology , dapi , primary and secondary antibodies , staining , computational biology , fixation (population genetics) , immunofluorescence , antigen , microbiology and biotechnology , antibody , computer science , pathology , immunohistochemistry , immunology , genetics , medicine , gene
Multicolor immunostaining analysis is often a desirable tool in cell biology for most researchers. Nonetheless, this is not an easy task and often not affordable by many laboratories as it might require expensive instrumentation and sophisticated analysis software. Here, we describe a simple protocol for performing sequential immunostainings on two different Drosophila specimens. Our strategy relies on an efficient and reproducible method for removal primary antibodies and/or fluorophore‐conjugated secondary antibodies that does not affect antigene integrity. We show that alternation of multiple rounds of antibody incubation and removal on the same slide, followed by registration of the same DAPI‐stained image, provides a simple framework for the sequential detection of several antigens in the same cell. Given that the sample fixation procedures used for Drosophila tissues are compatible with most specimen processing protocols, we can envisage that the multicolor immunostaining strategy presented here can be also adapted to different samples including mammalian tissues and/or cells. J. Cell. Physiol. 229: 683–687, 2014. © 2013 Wiley Periodicals, Inc.