Name: Jovanovic-Talisman Lab
Keywords: Jovanovic-Talisman Lab

Research Lab Overview

The Jovanovic-Talisman Lab within Beckman Research Institute of City of Hope employs novel, quantitative imaging techniques and nanobiology to investigate biological mechanisms, develop new diagnostic assays, and advance therapeutics.

Quantitative super-resolution fluorescence microscopy method development

Talisman Lab Figure1 — Golfetto, Wakefield, et al., J Am Chem Soc. 2018,140,12785
Rogacki, Golfetto et al., *Traffic*, 2018, 19, 690

To investigate biological processes that drive the progression of cancer and other diseases, we employ super-resolution fluorescence microscopy methods such as quantitative single molecule localization microscopy (qSMLM). This cutting-edge imaging method evaluates single molecules with nanoscale precision. In typical qSMLM experiments, target molecules are detected with fluorescent reporters (e.g., optical highlighter proteins or antibodies labeled with photoswitchable dyes). Because these imaging probes exhibit complex photophysical behaviors (cycling between “on” and “off” states), we developed a Surface Assay for Molecular Isolation (SAMI) to achieve robust molecular counting. We apply SAMI and other methodological innovations to elucidate key biological mechanisms and to support the development of novel diagnostic and therapeutic strategies.

Advancing super-resolution fluorescence microscopy for molecular diagnostics

We advance super-resolution microscopy methods to assess patient specimens, including cells from excised tissues and extracellular vesicles and particles from biofluids.

Extracellular vesicles and particles (EVPs) released by cells play critical roles in both normal physiological processes and the pathogenesis of various diseases. Since these nanoscopic structures carry molecular signatures reflective of their cells of origin, respond rapidly to changes in cellular status, and can be readily isolated from accessible biofluids, they represent promising candidates for disease biomarkers. However, EVPs are highly heterogeneous in size, composition, and biogenesis. Further confounding their analysis, biofluids are complex mixtures that contain EVPs derived from multiple cell types.

To address key challenges in the EVP field, we are developing advanced imaging platforms. We recently integrated affinity capture and super-resolution microscopy to enable characterization of individual EVPs. The Single Extracellular VEsicle Nanoscopy (SEVEN) and SEVEN-Universal Protocol (SEVEN-UP) platforms can detect both “general” EVP subpopulations and tissue- or disease-enriched EVPs carrying specific molecular markers. SEVEN and SEVEN-UP offer high capture efficiency, a broad linear dynamic range for EVP quantification, and compatibility with crude biofluids. Moreover, these platforms provide molecular sensitivity for EVP cargo detection, resolve EVP morphology across their full biological size spectrum, and reveal population heterogeneity. Collectively, these capabilities advance EVPs in biomarker research and mechanistic studies.

Touch Prep — Tobin, Wakefield et al., Sci. Rep., 2018, 11, 15154
Wakefield, Tobin et al., Methods Mol. Biol., 2022, 2394, 231
Maddox et al., Cancers, 2022, 14, 2795

We apply advanced qSMLM methods to quantify both the density and nano-organization of membrane receptors. Our current efforts focus on analyzing transmembrane glycoproteins. Using fluorescently labeled therapeutic antibody, trastuzumab, we recently quantified human epidermal growth factor receptor 2 (HER2) expression in breast cancer cells. By combining qSMLM with tissue touch preparation, we enabled high-resolution analysis of human tumor samples. We also developed a streamlined analytical pipeline and custom algorithms for rapid data analysis, allowing quantitative results to be obtained within a single day. Results from these studies demonstrated a significant correlation between classical HER2 screening method (i.e., fluorescence in situ hybridization, FISH) and qSMLM-based HER2 quantification. Moreover, our findings demonstrated that therapy response was associated with high HER2 density and receptor clustering. In the future, qSMLM could complement existing diagnostic standards by providing molecular-level insights into receptor organization and therapeutic response.

Principal Investigator: Tijana Jovanovic-Talisman, Ph.D.

Associate Professor, Department of Molecular Medicine

Research Focus