Protocol for whole-mount immunofluorescence staining of ECM gel-embedded innervated pancreatic organoids
H. Erdinc Besikcioglu, Ümmügülsüm Yurteri, Linhan Ye, Fangfang Zhang, Alessandra Moretti, İbrahim Halil Gürçınar, Alper Doğruöz, Didem Karakaş, Helmut Frieß, Güralp O. Ceyhan, Rouzanna Istvánffy, İhsan Ekin Demir
Abstract
The mandatory usage of extracellular matrix (ECM) gels in 3D cultures limits antibody penetration and increases background, while the removal of ECM gel causes disruption of morphology and sample loss. These factors pose challenges to effective immune labeling-based staining. Here, we present a protocol for whole-mount immunofluorescence staining of gel-embedded pancreatic organoids. We describe steps for sample fixation, blocking, and antibody incubation. We detail procedures for washing antibodies and mounting.
Topics & Concepts
StainingExtracellular matrixImmunofluorescenceOrganoidAntibodyFixation (population genetics)PathologyBiologyChemistryCell biologyImmunologyMedicineBiochemistryGene3D Printing in Biomedical ResearchCancer Cells and MetastasisMonoclonal and Polyclonal Antibodies Research