Histological and Functional Testing

Histological and functional results provide crucial, multi-dimensional information on tissue morphology, subcellular structure, and mitochondrial function. Protheragen offers a wide range of the most detailed histological and functional analyses needed for mitochondrial disease research or therapy development. Adaptable assay design and validation are available to satisfy particular investigation needs from novel biomarker identification efforts to in-depth mechanism-of-action studies and extensive preclinical efficacy analysis.

Overview of Histological and Functional Testing

Histological and functional testing is the backbone of phenotyping mitochondrial integrity and health. This multi-faceted approach combines high-resolution microscopy and biochemical assays to study the morphology as well as the intrinsic function of mitochondria in tissue and cells. Histology includes the visualization of mitochondrial morphology, distribution, or associated damage by specific staining and advanced microscopy. Functional testing the work capacity of the organelle down to fine detail, and centers on the operation of the Electron Transport Chain (ETC), cellular respiration rates, ATP production, and calcium handling functions. Together, these analyses provide quantitative information for the development of disease biomarkers and testing candidates for therapeutic intervention.

The pS129 lesion is associated with abnormal mitochondrial morphology in neurons. Fig.1 Neurons harboring pS129 lesions exhibit altered mitochondrial morphology. (Ciceri, D., et al., 2023)

Application of Histological and Functional Testing

Preclinical Efficacy Evaluation
Assesses therapeutic candidate effects at the tissue level, observing restoration of mitochondrial morphology, reduction in oxidative damage, and improvement in disease model histopathology endpoints.
Biomarker Discovery & Validation
Discovers and spatially localizes protein and metabolite markers for monitoring disease progression or response to therapy in affected tissue. Facilitates diagnostic and pharmacodynamic applications.

Disease Mechanism Elucidation
Correlating certain genetic defects or biochemical defects to structural changes (for example, mitochondrial ultrastructure and tissue remodeling) and functional deficits in the target organs (e.g., ‏muscle, brain, liver).
Disease Model Characterization & Validation
Fully phenotyping cellular and animal models of mitochondrial disease to verify that they recapitulate major human pathological features, thereby aiding in drug translational research.

Our Services

When combined with our dedicated team of histopathologists and cell biologists, steadfast imaging and analytical platforms, we offer accurate, reliable, and biologically meaningful data. This expertise provides tailored study designs, both from standard staining protocols and multiplexed assays that are fully customized, ensuring clients obtain the specific insights required to advance their research and development programs.

Workflow for Histological and Functional Testing Services

Consultation & Protocol Design: Starting with a comprehensive consultation to learn the client's research objectives, desired sample type, and readouts, followed by a bespoke study plan.
Sample Receipt and Processing: Samples (e.g., tissue biopsies, isolated mitochondria, cultured cells) are received, followed by quality control checks and appropriate preparation for either histology (fixation, embedding) or functional assays (isolation, plating).
Integrated Assay Execution: Rigorously performing both histological and functional analyses. This involves executing specialized tissue staining and microscopy imaging, followed by quantitative morphological analysis by experienced pathologists, or running established functional assays using appropriate controls and replicates.
Reporting & Data Delivery: Provision of detailed reports containing high-quality images, quantitative datasets, and expert interpretation of findings.

Methodologies of Histological and Functional Testing

Advanced Imaging and Morphometric Analysis

Integrating high-resolution light microscopy, TEM, and multiplex immunofluorescence for comprehensive ultrastructural and spatial evaluation of mitochondrial pathology in tissues/cells, including quantification of mitochondrial morphology, cristae integrity, and protein localization.

Histochemistry and Enzyme Activity Profiling

Applying direct enzyme histochemistry (e.g., COX, SDH) on tissue sections to visualize localized respiratory chain deficiencies, complemented by quantitative spectrophotometric assays of individual mitochondrial complex activities in tissue homogenates or cell lysates.

Bioenergetics and Metabolic Flux Analysis

Using real-time metabolic flux analyzers to determine oxidative phosphorylation and glycolysis, together with stable isotope-resolved metabolomics for dynamic tracing of pathway usage and detection of metabolic bottlenecks, supporting therapy candidate identification.

Functional Imaging & Live-Cell Assays

Employing fluorescence-based multiparameter assays in live cells to simultaneously and dynamically evaluate key functional metrics, including mitochondrial membrane potential, reactive oxygen species production, network dynamics, and calcium homeostasis.

Advanced Platforms for Histological and Functional Testing

Muscle Biopsy Analysis Platform
Muscle biopsies are used in the evaluation of both the structure and function of muscles affected by mitochondrial disease. This assay provides a foundational entry point into the diagnostic evaluation and can provide valuable information about the integrity and function of the mitochondria.

Neuroimaging Service Platform
Multi-modal imaging approach incorporating structural MRI, functional MRI, and metabolic imaging to evaluate central nervous system involvement, with particular focus on basal ganglia, brainstem, and white matter abnormalities characteristic of mitochondrial disorders.

Cellular Bio energetics Testing Platform
Systematic assessment of mitochondrial function in cultured cells or tissue samples using integrated respirometry, fluorometric assays, and enzymatic activity measurements. It is useful in detecting imbalances of the Krebs cycle, electron transport chain, or cytochrome enzyme systems.

Contact Us

Protheragen's end-to-end histological and functional testing capabilities offer the critical link between molecular dysfunction and tissue-level pathology, informing mitochondrial disease research. Through the combination of technical precision and a deep biological understanding, clients are supported to make informed decisions about their diagnostic and therapeutic pipeline development. Contact our scientific team to discuss your project in more detail and explore how our capabilities can support your specific objectives.

Reference

Ciceri, Dalila et al. "Histological Examination of Mitochondrial Morphology in a Parkinson's Disease Model." Journal of visualized experiments: JoVE ,196 (2023).

For research use only, not for clinical use.