TEER Explained: Measuring Cellular Barrier Integrity

TEER (Trans-Epithelial Electrical Resistance) is a widely accepted method for quantifying epithelial and endothelial barrier integrity by measuring electrical resistance across a confluent cell monolayer; impedance technology detects how much current is blocked by tight junctions and cell membranes, with low frequencies sensitive to barrier properties and high frequencies detecting cell confluence, enabling real-time, non-invasive monitoring of barrier function for applications in disease modeling and drug development.

The blood-brain barrier (BBB) consists of endothelial cells, pericytes, and astrocytes working together to protect the brain. It exhibits two primary barrier functions: physical barrier (high electrical resistance of 2000-5000 ohm·cm² limiting paracellular transport) and metabolic barrier (efflux pumps like P-glycoprotein actively transporting drugs back to bloodstream). Despite extensive drug development, the amount of drugs crossing the BBB remains extremely limited. iPSC-derived brain endothelial cells offer a promising approach for creating more accurate BBB models, but traditional methods using iPSC-derived cells alone often fail to achieve barrier functions comparable to in vivo conditions. Co-culturing with primary pericytes and astrocytes significantly increases barrier function. During brain development, the BBB forms in a hypoxic environment but requires re-oxygenation for maturation. Applying hyperoxic stimulation (approximately 5% oxygen tension) during the maturation phase significantly enhances barrier function by increasing tight junction protein expression. Microfluidic chips for BBB research consist of two interconnected channels separated by a porous membrane, with the lower channel containing brain endothelial cells and the upper channel containing co-cultured pericytes and astrocytes. This design allows simulation of blood flow conditions and easy sampling for quantitative analysis. Validation involves measuring electrical resistance (TEER), with well-established BBB models achieving values of 2000-5000 ohm·cm². Research has shown that efflux pumps are expressed in static culture conditions but their active transport function is significantly enhanced under dynamic flow conditions. Transcytosis assessment reveals that different antibodies targeting the same receptor can show dramatically different transcytosis efficiencies depending on their ability to undergo exocytosis. Osmotic opening is a strategy for temporarily increasing BBB permeability by administering hyperosmotic agents (such as mannitol) that cause brain endothelial cells to shrink, creating gaps that allow larger molecules to pass through. This process is reversible when isotonic media is reintroduced.

Organ-on-a-chip technology uses microfluidic engineering to create microengineered models of human tissues and organs by mimicking their structural features (such as the three-layer barrier between lung tissue and capillaries) and environmental conditions (like mechanical deformation during breathing), enabling researchers to study complex physiological processes, model diseases, and test drug responses in ways that traditional petri dish cultures cannot achieve.

Microfluidic organ-on-a-chip technology creates physiologically relevant 3D tissue models addressing limitations of traditional cell cultures. Platforms use 384-well formats with integrated microfluidic channels enabling high-throughput screening using standard laboratory equipment. Different chip designs accommodate various applications: basic designs enable simple tissue models, intermediate designs enable gradient formation and multiple tissue types, and advanced designs enable organoid integration. Endothelial co-culture significantly enhances physiological differentiation across organ systems, with tissue-matched microvascular endothelium enhancing barrier function, brush border formation, and proper polarization. For inflammatory disease modeling, four principal components must be captured: priming stimulus, biochemical changes inducing chemokine/adhesion molecule expression, immune cell recruitment resisting blood flow, and effector functions driving barrier dysfunction. Microfluidic models demonstrate inflammation-selective recruitment of immune cell subsets matching human patient observations. Validated therapeutics including dexamethasone, vedolizumab, and anti-TNF therapies show expected effects, enabling support of new therapeutic pipelines.

The intestinal epithelium maintains barrier function through tight junctions (ZO-1) and adherens junctions (E-cadherin), preventing paracellular passage of large molecules. FITC-dextran assays demonstrate functional barrier integrity in organoids, with disruption causing fluorescent tracer entry into the lumen. Clostridium difficile toxins (A and B) disrupt barrier function through differential mechanisms: both are glucosyltransferases targeting Rho GTPases, but Toxin A is significantly more potent than Toxin B in inducing barrier breakdown. This differential effect correlates with reduced myosin light chain phosphorylation in Toxin A-treated samples. The mechanism explaining this potency difference remains under investigation.

Barrier integrity can be assessed using fluorescently labeled dextran molecules of different sizes. With no cells present, large dextrans (e.g., 70 kDa FITC-dextran) freely diffuse across the membrane. When brain microvascular endothelial cells are present, permeability is dramatically reduced, demonstrating successful barrier formation. This simple assay provides visual confirmation of BBB function and allows comparison between different experimental conditions or cell sources. The degree of permeability reduction correlates with the tightness of the barrier and the effectiveness of the BBB-inducing protocol.

Intestinal permeability is measured by gavaging mice with fluorescently-labeled dextran (FITC-dextran), which passes through damaged intestinal barriers into the bloodstream. Blood samples are then taken, and fluorescence intensity indicates how much of the marker has entered circulation, providing a quantitative measure of barrier integrity disruption.

Three validation methods are used to confirm that a cell monolayer represents a permeability barrier: (1) measuring trans-epithelial electrical resistance (TEER), which quantifies the integrity of the cell layer; (2) immunostaining cell junctions to visualize tight junction proteins; and (3) assessing passive permeability using a detectable tracer molecule.

Tight junctions are occluding junctions that maintain epithelial cell polarity by preventing apical and basolateral membrane proteins from diffusing between domains. They form a tight seal near the apical domain, creating a barrier to solute diffusion. Electron-dense tracer experiments demonstrate that solutes cannot pass through tight junctions from apical to basolateral side. Tight junctions are not completely impermeable; the paracellular pathway allows certain small molecules and ions to pass. The CFTR chloride channel is an example of a component that can cross tight junctions, and its mutation causes cystic fibrosis. Tight junctions are essential for transcellular transport, such as glucose absorption in the intestine, using sodium-glucose symporters and GLUT2 transporters. Tight junctions are composed of occludin, claudin (multipass transmembrane proteins), and JAM (single-pass transmembrane protein), forming tight bands that create the seal.

Research using Caco-2 intestinal epithelial cell monolayers demonstrated that prebiotic treatment protects against pathogen-induced barrier disruption. Cells were pretreated with inulin or scFOS before infection with a bacterial pathogen known to induce barrier disruption. Transepithelial electrical resistance (TEER) measurements showed that infected control cells experienced approximately 75% reduction in barrier integrity, while prebiotic-pretreated cells maintained approximately 50% of baseline resistance. Similarly, fluorescent dye flux assays showed significantly reduced leakage through prebiotic-treated monolayers. Immunostaining confirmed that prebiotics prevented the pathogen-induced disorganization of tight junction proteins.

The Tissue Engineering Laboratory at API Geneva has developed an integrated system that measures trans-epithelial/endothelial electrical resistance (TEER) in real-time and non-invasively to assess the quality of biological barriers, with applications in toxicology testing and pharmacological studies such as drug discovery, addressing the challenge that only one in nine drugs receives regulatory approval due to inability to reach targeted sites.

Advanced barrier models provide multiple readout parameters for comprehensive infection analysis: transepithelial electrical resistance measures epithelial integrity (higher resistance indicates better barrier function); live-cell imaging tracks mucociliary clearance dynamics; immunofluorescence detects viral localization and mucus production; ELISA quantifies pro-inflammatory cytokines; plaque assays measure infectious viral load released to apical or basolateral compartments. This multi-parameter approach enables detailed mechanistic studies of host-pathogen interactions.

Transepithelial Electrical Resistance (TEER) is a well-established technique used to assess the barrier function and integrity of epithelial and endothelial cell layers by applying a small AC current to the cell monolayer and measuring the resistance to ion flow, providing valuable insights into tight junction integrity and cell-cell interactions; WPI's EVOM family of meters, including manual and automated systems for 24-well and 96-well plates, serves as the gold standard in TEER technology, having been trusted for nearly 40 years and cited in thousands of peer-reviewed publications for applications in drug absorption studies, tissue engineering, disease modeling, and quality control.

Concentric electrodes integrated into the skin-on-a-chip device enable non-destructive real-time monitoring of trans-epithelial electrical resistance (TEER), reflecting overall barrier integrity to ions. During culture, TEER values increase progressively reflecting accumulation of stratum corneum and gradual formation of tight junctions. Exposure to benchmark irritants like SDS causes decreased TEER values, confirming disruption of barrier function through histological analysis and luciferin dye penetration studies.