Phenotypic Screening

Phenotypic Screening is used to characterise the effect of test molecules by quantifying morphological/phenotypic changes in cell-based models.

Tight Junction Proteins (TJP)

Tight Junction (TJ) proteins are major components of cell–cell adhesion complexes that maintain cell polarity, barrier integrity and regulate diffusion of certain molecules, cell proliferation and differentiation.

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Caco-2 cells were differentiated for 21 days and stained for tight junction proteins such as ZO-1 (purple) and Occludin (orange) along with DAPI (blue- nucleus). The images were captured at 20X magnification using high content imager and the intensities of each fluorescence analysed.

Wound Healing

Wound healing is a complex and dynamic physiological process of replacing devitalised and missing cellular structures and tissue layers. Dysregulated wound healing is a sign of underlying disease pathology.

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A scratch was formed on a A549 cell monolayer and treated with Olaparib (Ola), 5-Fluorouracil (5-FU); a combination of 5-fluorouracil and Olaparib (5-FU + Ola). Images were acquired using live imaging platform at the indicated time points (hrs). The wound size/scratch was measured at all time points. The wound closure percentage was analysed assuming 0% closure at 0 hours, and calculating all the following time points as ratio to 0 hours [ *p<0.05; **p<0.01;***p<0.001; ± SEM].

Senescence

Senescence is the process by which cells irreversibly stop dividing and enter a state of permanent growth arrest without undergoing cell death. Senescence can be induced by unrepaired DNA damage or other cellular stresses and is often lined with ageing, cancer and fibrosis.

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Primary human chondrocytes were treated with H2O2 for 24 hours followed by 72 hours recovery period and then treatment with ABT 263. Cells were fixed followed by immunofluorescence staining for DAPI and β-galactosidase was performed.

Phagocytosis

Phagocytosis is a process mediated by a specialised group of innate immune cells called phagocytes, including neutrophils, monocytes and macrophages. This is an essential process to maintain tissue homeostasis.

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Monocytes (CD14+) were isolated from PBMCs of healthy donors followed by treatment with M-CSF for 6 days. On day 6, the naïve macrophages were exposed to GM-CSF + IFNγ and M-CSF to generate M1 and M2 macrophages (MΦ) respectively. The cells were then exposed to fluorescent zymosan particles (green) and incubated for 3 hours. After 3 hours, the cells were washed, stained with fluorescent membrane marker (red) and DAPI (blue) for nucleus.

The images above showed M2 macrophages engulfed higher numbers of zymosan particles when compared to M1 macrophages (Objective= 20X).

Collagen Gel Contraction

Gel contraction assay is used to study the cell-mediated reorganisation of the extracellular matrix. Hyperreactivity of smooth muscle cells/fibroblasts can be investigated using this principle.

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Percentage change in gel contraction upon pre-treatment with 2,3 BDM followed by incubation for 4 hours with Carbachol. For each treatment the percentage change in the gel contraction was calculated based on the values obtained at time 0. Each condition is representative of 3 biological replicates (n=3); mean+SEM. Data analysis was performed using the 2-WAY ANOVA analysis, *p<0.05; **p<0.01, ***p<0.0001; Carbachol vs. test ingredients groups. 

Antibody Dependent Cellular Phagocytosis (ADCP)

ADCP is a mechanism by which macrophages engulf and eliminate the tumour cells targeted with monoclonal antibodies.

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The target Raji cells were pre-treated with or without Rituximab at different concentrations and stained with pHrodo Red Cell Labelling (red). The THP-1 derived macrophages were stained with calcein staining (green). To assess antibody-dependent cellular phagocytosis of target Raji cells by THP-1 derived macrophages the two cell types were co-cultured for 6 hours. Data is represented as the red fluorescence intensity (RFI)  at 0 hours vs. 6 hours in all treatment groups.

Histology and IHC

This is an imaging technique to aid microscopic anatomy of biological tissues.

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Normal Human Bronchial Epithelial Cells were grown on Air Liquid Interface (ALI) and allowed to differentiate for 28 days. At the end of the culture the cells were fixed and embedded into paraffin. The paraffin blocks were sectioned at 5µm thickness using Microtome and fixed onto slides. Haematoxylin and Eosin (H & E) staining were performed and slides were imaged at 4, 20 and 40X magnification. Distinct goblet and ciliated cells were observed on Day 28 of the ALI differentiation

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To request a consultation where we can discuss your exact requirements, please contact Cellomatics Biosciences.