Immunostaining of Drosophila Gut Tissue

Dissection, fixation and immunostaining were performed as described by Micchelli, 2014 (link). Dilutions of the various antibodies were: mouse anti-Armadillo N27A1 at 1:50 (DSHB), mouse anti-Connectin-C1-427 at 1/200 (DSHB), mouse anti-Prospero MR1A at 1:200 (DSHB), rabbit anti-PH3 at 1:1000 (Millipore, 06–570), Rabbit anti-Cleaved Caspase-3 at 1/600 (Cell Signalling Asp175 #9661), Goat anti-mouse AlexaFluor-647 at 1/500 (Molecular Probes Cat# A-21235), Goat anti-mouse AlexaFluor-546 at 1/500 (Molecular Probes Cat# A-11003), Goat anti-rabbit AlexaFluor-647 at 1/500 (Thermo Fisher Scientific Cat# A32733), Goat anti-rabbit AlexaFluor-546 at 1/500 (Thermo Fisher Scientific Cat# A-11010). For microscopy, guts were mounted in Fluoroshield DAPI medium (Sigma, # F6057) and immediately observed on a Zeiss Axioplan Z1 with Apotome 2 microscope. Images were analyzed using ZEN (Zeiss), ImageJ and Photoshop software. Image acquisition was performed at the microscopy platform of the Sophia Agrobiotech Institute (INRAE1355-UCA-CNRS7254 – Sophia Antipolis).

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Jneid R., Loudhaief R., Zucchini-Pascal N., Nawrot-Esposito M.P., Fichant A., Rousset R., Bonis M., Osman D, & Gallet A. (2023). Bacillus thuringiensis toxins divert progenitor cells toward enteroendocrine fate by decreasing cell adhesion with intestinal stem cells in Drosophila. eLife, 12, e80179.

Publication 2023

rabbit anti-PH3 Rabbit anti-Cleaved Caspase-3 Goat anti-mouse AlexaFluor-647 Goat anti-mouse AlexaFluor-546 Goat anti-rabbit AlexaFluor-647 Goat anti-rabbit AlexaFluor-546 Fluoroshield DAPI medium Axioplan Z1 ZEN

Alexafluor 647 Antibodies Armadillo Caspase 1 Connectin Dapi Dilutions Dissection Fluoroshield Goat Guts Microscopy Molecular probes Mouse Rabbit

Corresponding Organization :

Other organizations : Institut National de Recherche pour l'Agriculture, l'Alimentation et l'Environnement, Université Côte d'Azur, Lebanese University, Agence Nationale de Sécurité Sanitaire de l’Alimentation, de l’Environnement et du Travail

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Variable analysis

independent variables

Dissection, fixation and immunostaining procedures

dependent variables

Localization and expression of Armadillo, Connectin, Prospero, PH3, and Cleaved Caspase-3 proteins

control variables

Antibody dilutions
Mounting medium
Microscope and imaging software used

controls

Positive control: Not mentioned
Negative control: Not mentioned

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