Biology subjects

Challa, P.

Publications and source records attributed to Challa, P..

3 recordsLinked to original sources

bioRxiv2020

Machine learning aided top-down protein identification on a microfluidic platform

The ability to determine the identity of specific proteins is a critical challenge in many areas of cellular and molecular biology, and in medical diagnostics. Here, we present a microfluidic protein characterisation strategy that within a few minutes generates a three-dimensional fingerprint of a protein sample indicative of its amino acid composition and size and, thereby, creates a unique signature for the protein. By acquiring such multidimensional fingerprints for a set of ten proteins and using machine learning approaches to classify the fingerprints, we demonstrate that this strategy allows proteins to be classified at a high accuracy, even though classification using a single dimension is not possible. Moreover, we show that the acquired fingerprints correlate with the amino acid content of the samples, which makes it is possible to identify proteins directly from their sequence without requiring any prior knowledge about the fingerprints. These findings suggest that such a multidimensional profiling strategy can lead to the development of novel method for protein identification in a microfluidic format.

biophysics↗

bioRxiv2020

Antifibrotic activity of a rho-kinase inhibitor restores outflow function and intraocular pressure homeostasis

Glucocorticoids are widely used as an ophthalmic medication. A common, sight-threatening adverse event of glucocorticoid usage is ocular hypertension, caused by dysfunction of the conventional outflow pathway. We report that netarsudil, a rho-kinase inhibitor, rapidly reversed glucocorticoid-induced ocular hypertension in patients whose intraocular pressures were uncontrolled by standard medications. Mechanistic studies in our established mouse model of glucocorticoid-induced ocular hypertension show that netarsudil both prevented and reversed intraocular pressure elevation. Further, netarsudil reversed characteristic steroid-induced pathologies as assessed by quantification of outflow function and tissue stiffness, and morphological and immunohistochemical indicators of tissue fibrosis. Thus, rho-kinase inhibitors act directly on conventional outflow cells to efficaciously prevent or reverse fibrotic disease processes in glucocorticoid-induced ocular hypertension. These data motivate a novel indication for these agents to prevent or treat ocular hypertension secondary to glucocorticoid administration, and demonstrate the antifibrotic effects of rho-kinase inhibitors in an immune-privileged environment.

physiology↗

bioRxiv2020

Direct digital sensing of proteins through single-molecule optofluidics

The detection of proteins is of central importance to biomolecular analysis and diagnostics, yet fundamental limitations due to the surface-based nature of most sensing approaches persist, and limited improvements have been designed to integrate multimodal information beyond concentration measurements. Here we present a single-molecule microfluidic sensing platform for digital protein biomarker detection in solution, termed digital immunosensor assay (DigitISA). DigitISA is based on microchip electrophoretic separation combined with single-molecule detection and enables absolute number-concentrations quantification of proteins in a single, solution-phase step. Applying DigitISA to a range of targets including amyloid aggregates, exosomes, and biomolecular condensates, we demonstrate that the assay provides information beyond stoichiometric interactions, and enables characterization of immunochemistry, binding affinity, and protein biomarker abundance. Together, DigitISA constitutes a new experimental paradigm for the digital sensing of protein biomarkers, and enables analyses of targets that would otherwise be hard or impossible to address by conventional immuno-sensing techniques.

biophysics↗