8 well chamber slide

Manufactured by Ibidi

Sourced in Germany, United States

The 8-well chamber slides are a lab equipment product designed for cell culture applications. The slides feature eight individual wells, allowing for the simultaneous culturing and observation of multiple samples. The product provides a convenient and standardized platform for various cell-based experiments and analyses.

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151 protocols using 8 well chamber slide

GBM Cell Irradiation and Viability

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After 24 h of treatment, GBM cells were irradiated with 2 Gy using an IBL 637 (CIS Bio-International, Codolet, France). Following the irradiation, the medium was replaced by fresh medium, followed by 72 h incubation at 37 °C and 5% CO₂. This treatment cycle was repeated once. Thereafter, cell viability was measured using the Calcein-AM assay. Double-strand breaks (DSB) were assessed with γ-H2AX staining in 8 Well chamber µ-slides (ibidi, munich, Germany) as described before [32 (link)].

Linke C., Freitag T., Riess C., Scheffler J.V., del Moral K., Schoenwaelder N., Fiedler T., Fiebig A., Kaps P., Dubinski D., Schneider B., Bergmann W., Classen C.F, & Maletzki C. (2023). The addition of arginine deiminase potentiates Mithramycin A-induced cell death in patient-derived glioblastoma cells via ATF4 and cytochrome C. Cancer Cell International, 23, 38.

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Intracellular Nanoparticle Trafficking and ROS Analysis

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Cells seeded in 8-well chamber µ-slides (ibidi, Martinsried, Germany) were treated with free TB and DNA/TB NG (8 μM TB) for 20 h. Then, the cells were incubated with Hoechst dye for 2 min to stain the nucleus. The cells were observed on an EVOS FL Auto microscope (Thermo Fisher Scientific, Waltham, United States) with a Cy5 channel for TB and a DAPI channel for Hoechst imaging. The intracellular TB fluorescence was quantified using Fiji/ImageJ software. For intracellular ROS analysis, cells were first treated with free TB and DNA/TB with the TB concentration set at 2 μM. Then, the cells were stained using the DCFDA probe following the manufacturer’s instructions. The cells were further irradiated by an LED light source (660 nm, 25 mW/cm²) for 10 min. Immediately after the light irradiation, DCFDA fluorescence from the cells was analyzed on a Gallios flow cytometer (Beckman Coulter, Brea, CA, United States).

Guo H., Wang H., Deng H., Zhang Y., Yang X, & Zhang W. (2023). Facile preparation of toluidine blue-loaded DNA nanogels for anticancer photodynamic therapy. Frontiers in Bioengineering and Biotechnology, 11, 1180448.

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Biofilm Thickness and Volume Quantification

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The treated biofilms in 8-well chamber slides (ibidi GmbH, Grafelfing, Germany) were stained using BacLight Bacterial Viability Kit^TM (L7007, Thermo Fisher Scientific, Waltham, MA, USA). The slides were then incubated in dark for 30 min following the manufacturer’s instructions. The stained biofilms were visualized using an oil-immersion objective lens (×60) of a confocal laser-scanning microscope (CLSM; Fluoview FV 1000, Olympus, Tokyo, Japan) at three randomly selected points. The biofilm z-stacks were reconstructed into 3D images, which were analyzed using IMARIS software (Bitplane, St. Paul, MN, USA) to quantify the biofilm thickness and volume.

Hu M., Kalimuthu S., Zhang C., Ali I.A, & Neelakantan P. (2022). Trans-cinnamaldehyde–Biosurfactant Complex as a Potent Agent against Enterococcus faecalis Biofilms. Pharmaceutics, 14(11), 2355.

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HPV16 Colocalization with EEA1 in HeLa Cells

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For HPV16 colocalization with EEA1, HeLa cells were seeded into 8-well chamber slides (ibidi) at 1 × 10⁴ cell/well and grown overnight. HPV16 PsV were diluted in PBS or treated with indicated concentration of RTD-1 or HD5 for 1 h at 37° prior to addition to cells. Cells were treated with 0.5 μg PsV/1 × 10⁶ cells for 2 h at 37 °C. After treatment, cells were gently washed with PBS to remove unbound excess virions and then fixed with 4% paraformaldehyde. Cells were permeabilized with triton X-100 and stained overnight in a humidified chamber at 4°C for HPV16 (H16.56E, 1:200) and EEA1 (1:250). After secondary antibody incubation for 30 min at RT, excess antibody was removed, and slides were coverslip mounted with ProLong Gold Antifade Mountant with DAPI (Thermo Fisher). Immunofluorescent (IF) images were captured on a Nikon Eclipse Ti-E laser scanning confocal microscope running Nikon Elements software (version 4.0). For colocalization analysis, 15 images from each treatment group with >20 cells/image were analyzed with Fiji (a distribution of ImageJ, NIH) (27 (link)). Using the JACoP plugin thresholds were automatically set, and the extent of colocalization was measured and reported as Mander's colocalization coefficient (28 (link)).

Skeate J.G., Segerink W.H., Garcia M.D., Fernandez D.J., Prins R., Lühen K.P., Voss F.O., Da Silva D.M, & Kast W.M. (2020). Theta-Defensins Inhibit High-Risk Human Papillomavirus Infection Through Charge-Driven Capsid Clustering. Frontiers in Immunology, 11, 561843.

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Imaging FITC-labeled RFPs in MCF7 Cells

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MCF7 cells were grown in 8-well chamber slides (Ibidi) and treated with FITC-labeled RFPs (5 μM) in 10% FBS-containing RPMI for 8 h at 37 °C. Cells were washed 5× in PBS, fixed in 2% paraformaldehyde/PBS and stained with DAPI according to the manufacturer’s protocol (Sigma). A coverslip was mounted onto the slide and confocal fluorescence microscopy performed with an Olympus DSU spinning disk confocal microscope (Olympus Corporation of the Americas, Center Valley, PA) with a Hamamatsu model C9100 EM-CCD camera (Hamamatsu Photonics, Skokie, IL) run by SlideBook v5.0 software (Intelligent Imaging Innovations, Denver, CO). Post-acquisition processing (multi-channel overlay) was performed using ImageJ software (NIH). Results are representative of images taken from three fields across the same well in at least two biological replicates.

Mitra S., Montgomery J.E., Kolar M.J., Li G., Jeong K.J., Peng B., Verdine G.L., Mills G.B, & Moellering R.E. (2017). Stapled peptide inhibitors of RAB25 target context-specific phenotypes in cancer. Nature Communications, 8, 660.

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Visualizing Intracellular Parasite Replication

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In addition to light microscopy, CLSM (TCS-SP8, Leica, Bensheim, Germany) was applied to observe the morphology of the infected cell cultures, for localization of parasites within the host cells, and to estimate the extent of intracellular replication. Therefore, 2 × 10⁶ PBMC per well were cultivated in 8-well chamber slides (Ibidi, Martinsried, Germany) for 4 days to grow pure cultures of primary macrophages. Infection conditions were the same as mentioned above for all infection groups (TH, TL, EH, EL, CI and NC). For CLSM, cultures were fixed with methanol for 10 min before further processing. 4', 6-diamidino-2-phenylindole (DAPI, Sigma-Aldrich, USA) was used to stain cell nuclei. Cell imaging was carried out by Leica Application Suite X (LAS X, Leica Microsystems, Wetzlar, Germany).

Zhang R., Thabet A., Hiob L., Zheng W., Daugschies A, & Bangoura B. (2018). Mutual interactions of the apicomplexan parasites Toxoplasma gondii and Eimeria tenella with cultured poultry macrophages. Parasites & Vectors, 11, 453.

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Immunocytochemistry of Schwann Cells and Fibroblasts

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Schwann cells or fibroblasts were seeded on 8-well chamber slides (Ibidi) coated with poly-L-lysine and laminin, as described.⁹ (link) Cells were harvested 24 to 48 h later and fixed in 4% paraformaldehyde (EM Sciences) for 15 min at RT. Fixed cells were permeabilized in PBS containing 1% bovine serum albumin and 0.2% Triton X-100 for 30 min at RT, and further incubated with anti-p75 NGFR overnight at 4°C in the dark. The next day, cells were washed in PBS containing 1% BSA two times for 5 min at RT, incubated with anti-phalloidin for 1 h in the dark at RT, and washed again as described above. Cells were mounted with ProLong Gold Antifade Mountant with DAPI (Thermo) and coverslipped using Gold Seal Cover Glass (EM Sciences). Images were acquired using a Cytation 5 cell imaging reader (BioTek).

Osum S.H., Oribamise E.I., Corbière S.M., Taisto M., Jubenville T., Coutts A., Kirstein M.N., Fisher J., Moertel C., Du M., Bedwell D., Largaespada D.A, & Watson A.L. (2023). Combining nonsense mutation suppression therapy with nonsense-mediated decay inhibition in neurofibromatosis type 1. Molecular Therapy. Nucleic Acids, 33, 227-239.

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Live-cell Imaging of Cell-Cell Interactions

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LifeA-ctGFP-HaCa4 and LifeAct-RFP-mLECs were co-cultured or single-cultured in 8-well chamber slides (ibidi, Gräfelfing, Germany) with 10 µM Hepes. Time-lapse experiments were performed with a Nikon Eclipse Ti2 microscope and the NIS-elements acquisition software (Nikon Instruments, Nikon Europe, Amstelveen, The Netherlands) at 20× magnification, with controlled temperature and CO₂ levels. Images were captured every 3 min over 13 h.

Cazzola A., Calzón Lozano D., Menne D.H., Dávila Pedrera R., Liu J., Peña-Jiménez D., Fontenete S., Halin C, & Perez-Moreno M. (2023). Lymph Vessels Associate with Cancer Stem Cells from Initiation to Malignant Stages of Squamous Cell Carcinoma. International Journal of Molecular Sciences, 24(17), 13615.

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Patient-Derived Ex Vivo Tumor Culture

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Tumor specimens for patient-derived ex vivo cultures were collected under the IRB protocol 93-085 with written informed consent from chemoresistant patients on the day of surgery. They were then digested mechanically and enzymatically using human tumor dissociation kit (Miltenyi Biotec). Cultures were initially grown in EpiCult medium (STEMCELL Technologies) with 10% FBS. Once the cultures began to establish, serum-free medium was used to discourage proliferation of fibroblasts. During passaging, fibroblasts were further separated from the samples using trypsin-EDTA. To assess efficacy of drugs, the cultures were seeded in 8-well chamber slides (ibidi) at 70% confluency and exposed to 72-hour drug treatment. Cell viability was assessed using trypan blue. Immunostaining of these cultures was also performed, as described above.

Koh S.B., Ross K., Isakoff S.J., Melkonjan N., He L., Matissek K.J., Schultz A., Mayer E.L., Traina T.A., Carey L.A., Rugo H.S., Liu M.C., Stearns V., Langenbucher A., Saladi S.V., Ramaswamy S., Lawrence M.S, & Ellisen L.W. (2021). RASAL2 Confers Collateral MEK/EGFR Dependency in Chemoresistant Triple-Negative Breast Cancer. Clinical Cancer Research, 27(17), 4883-4897.

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Fabrication of Alginate-Fibrin Interpenetrating Hydrogels

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UltraPure MVG sodium alginate (molecular weight >200,000 g/mol, Pronova, Lysaker, Norway) was oxidized to a degree of oxidation of 1% with sodium periodate (MilliporeSigma, St. Louis, MO) as previously described [14 (link)] and dissolved in PBS at 3% w/v. Fibrinogen (MilliporeSigma) was mixed with alginate solution at 30 mg/mL and allowed to dissolve at 37°C in PBS overnight. This solution remained sterile and stored at 4°C until use for subsequent experiments.
3.5 kDa MWCO dialysis membranes (Spectrum Labs, Rancho Dominguez, CA) were hydrated in sterile ultrapure water for at least 10 min. We prepared solutions of calcium chloride (CaCl₂, MilliporeSigma) in water at 10 mM or 40 mM. To obtain a 2% w/v alginate and 20 mg/mL Fibrinogen IPN, 66 μL of Fibrinogen-alginate solution was mixed with 33 μL of thrombin solution with a concentration range of 0–75 U/mL, and 80 μL was immediately pipetted into 8 mm circular, silicon molds. Fibrin networks were allowed to form for 45 min at 37°C. A hydrated dialysis membrane was then placed gently on top of molds, and CaCl₂ solution was pipetted on top until fully covered. Alginate was allowed to crosslink for 30 min at 37°C. Resulting constructs were gently transferred from molds to non-coated 8-well chamber slides (ibidi, GmbH, Planegg, Germany).

Vorwald C.E., Gonzalez-Fernandez T., Joshee S., Sikorski P, & Leach J.K. (2020). Tunable Fibrin-Alginate Interpenetrating Network Hydrogels to Support Cell Spreading and Network Formation. Acta biomaterialia, 108, 142-152.

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