Immunofluorescence Staining Using ibidi Labware

With ibidi labware, all steps of an immunofluorescence staining can be performed in the same microscopy chamber: cell can be cultured, fixed, stained, and imaged directly without any sample or coverslip transfer.

This guide compares the ibidi Chambered Coverslips, Channel Slides, and removable Chamber Slides for immunofluorescence workflows, helping you select the right format based on microscopy setup, reagent volume, handling requirements, and sample storage.

Rat dorsal root ganglionic cells and Schwann cells cultured in a µ-Slide 8 Well and stained for neurofilament (green), NGFR (magenta), and DAPI (white). The image was obtained with a LEICA SP8X laser scanning microscope. Data by Tamara Weiss, Division of Plastic and Reconstructive Surgery, Medical University of Vienna, Austria.

Benefits of ibidi µ-Chambers for Immunofluorescence

1. Fast and simple handling
   All-in-one chambers simplify your immunofluorescence workflow.
2. Cost-effective experiments
   Requires only a small number of cells and low reagent volumes.
3. High-resolution imaging
   Ideal for many basic and advanced microscopy methods, such as widefield fluorescence, confocal imaging, 2-photon microscopy, and super-resolution microscopy.

Immunofluorescence Protocol Comparison: Coverslips vs. ibidi Labware

Using ibidi labware significantly shortens the immunofluorescence workflow compared to traditional coverslip methods. Cells can be cultured and stained directly in µ-Slides, µ-Dishes, or µ-Plates, eliminating additional handling steps and reducing protocol time. Explore the step-by-step workflow for traditional immunofluorescence staining using coverslips (left) in comparison to the step-by-step workflow for staining directly in ibidi µ-Slides (right), minimizing handling and reducing overall assay time.

Traditional Immunofluorecence Protocol
With Cells on Coverslip

All-In-One Immunofluorescence Protocol
With ibidi µ-Slides

Comparison of ibidi Labware for Immunofluorescence Staining

The table below compares ibidi Chambered Coverslips, Channel Slides, removable Chamber Slides, and traditional 6 well plate workflows for immunofluorescence staining. It summarizes key differences in handling, reagent volume, coverslip requirements, microscopy compatibility, sample storage, and typical use.

The comparison above helps identify the most suitable ibidi labware format for immunofluorescence staining based on sample handling, reagent volume, microscopy method, and storage requirements. Chambered Coverslips and Channel Slides support direct staining and imaging in the same microscopy-ready vessel without handling any additional coverslips, while removable Chamber Slides are better suited for mounted samples and long-term storage. This makes the choice of labware an important factor for reproducible fluorescence staining, reduced sample handling, and efficient immunofluorescence workflows.

Immunofluorescence Application Examples in 2D and 3D Cell Culture

These application examples illustrate how ibidi labware supports immunofluorescence staining and fluorescence microscopy across different sample types and assay formats. The selected images show 2D and 3D cell samples, including endothelial junctions, neuronal and glial markers, cytoskeletal structures, mitochondria, and cells cultured under flow conditions.

Immunofluorescence staining of primary mouse brain microvascular endothelial cells (pMBMECs), cultured in the 12 Well Chamber, removable. Red: endothelial cell junctions labeled for zonula occludens (ZO)-1. Blue: nuclei stained with DAPI. The image was acquired with a 20× objective on a Nikon Eclipse microscope. Data by Sidar Aydin, Britta Engelhardt, Ruth Lyck, Theodor Kocher Institute, Bern, Switzerland.

Confocal maximum intensity projection (MIP) image of an 8-day breast tumor spheroid mounted on an ibidi µ-Slide 18 Well Glass Bottom. The image shows stainings for nuclei (blue), Ki67+ cells (green) and gap junction protein (red). Image by Marina Rodriguez-Candela Mateos, Institute of Biomedical Research of A Coruña (INIBIC), A Coruna, Spain.

Confocal microscopy of rat hippocampal neurons plated over astrocytes, cultured in a µ-Plate 24 Well. Red: Synapsin 1, a pre-synaptic marker. White: neuron-specific Tubulin Beta-III. Blue: DAPI nuclear counterstain. Data by Daniel Hoeppner, Lieber Institute for Brain Development, Baltimore, MD, USA.

Human umbilical vein endothelial cells (HUVECs) cultured under flow conditions in the µ-Slide I ^0.4 Luer. Red: actin cytoskeleton labeled with Cy5-conjugated antibody. Green: adherens junctions marked by VE-cadherin with Alexa 488-conjugated antibody. Blue: nuclear counterstaining with DAPI. Data by S. Zahler, Munich, Germany.

Fluorescence microscopy of immunostained MCF-7 cells in the 8 Well Chamber, removable. Green: alpha-Tubulin. Red: F-actin stained with phalloidin. Blue: nuclei stained with DAPI. Widefield fluorescence images were taken with a 20× objective.

COS7 cells were cultured in the µ-Slide 8 Well ^high Glass Bottom and immunolabeled for two mitochondrial proteins. Alexa Fluor 594 and Abberior STAR RED signals were imaged by STED microscopy using a 775 nm depletion laser and a 100× oil immersion objective. Image courtesy of Till Stephan, Goethe University, Frankfurt am Main, Germany.