Images of Connor's Work

  • Cor4U cells stained with MitoTracker dye and a nuclear stain

Connor Jacobson started working in the Harvard Program in Therapeutic Science in August 2015. He began his research working on a project, sponsored by the FDA, exploring cardiotoxic effects seen in patients treated with tyrosine-kinase inhibitors (TKIs). TKIs have been seen to be effective in reducing cancerous symptoms as first- and second-pass therapeutics but have been noted to cause cardiotoxicity such as myocardial infarctions and abnormal heart beating patterns, among others (the paper can be found here). As a member of both the LSP and Sorger Labs, Connor initially studied the longitudinal effects of multiple drugs dosed to cell types of interest (IPS-derived Cor4U cardiomyocytes) and investigated the ways in which these drugs effect Cor4U cells as well as various cancer cell lines. By employing a pre-existing protocol of cyclic immunofluorescence (CycIF; developed by Dr. Jia-Ren Lin of the Sorger Lab), over 200 unconjugated and direct;y conjugated antibodies have been validated. CycIF is traditional indirect immunofluorescence, however, once secondary antibodies have been added for imaging, a photo-bleaching solution (H2O2-based) is added to quench the fluorophores of the secondary antibodies. Following washes, the cells are re-stained with directly conjugated antibodies and re-imaged as subsequent “cycles.”

After an initial battery of optimization and troubleshooting experiments, Connor established a finely-tuned plate-based CycIF (p-CycIF) workflow in cardiomyocytes. By exploring how particular biomarkers clustered over time and dose at a single-cell resolution, Connor helped reveal which epitopes are indicative of cardiotox at the mechanistic and signaling-pathway level. The hope is to predict cardiotox when these TKIs are used in cancer patients and to offer combination therapies that may alleviate unwanted cardiotox.

Connor’s efforts and assay development testing over 200 directly conjugated and unconjugated antibodies now serves as the foundation for an open-source, real-time antibody validation database known as Reagent Tracker. Additionally, with the in-place work flow, Connor continued technical development with newer technologies to enable greater multi-plex and high-dimensionality such as Thermo’s Zenon-labeling. By matching species and isotype, one is able to label unconjugated antibodies with whichever fluorophore is available for that particular species, allowing multiple antibodies of the same species to be tested in the same cycle as long as the isotypes and fluorophore of the antibody are distinct. Furthermore, Connor has optimized the incorporation of an Edu-label into p-CycIF through “click-chemistry” which allows the tracking of cells’ progression through the cell cycle enabling the gating of cells in the G1, S, and G2 cell-cycle phases. Connor has also optimized and incorporated MitoTracker dyes staining which serves as a vital stain to gauge cell health. Connor has also tested and quantified cell loss, bleaching intensity and signal decay of over 10 fluorophores, as well as which dilutions of antibodies work best in particular conditions/cell types.

Since p-CycIF has been established...

Connor works tirelessly on establishing a standardized protocol for all cyclic immunofluorescence assays from initial experimental design to image registration and processing. By refining the current CycIF protocols down to the most minute details, Connor has streamlined the ways in which we perform high-throughput plate-based microscopy across multiple systems in HiTS. If you would like to perform a CycIF experiment or have specific questions as to how to best proceed, email him at:

With a degree in Neuroscience, Connor is interested in further exploring academic research, namely in the fields of immune-oncology and neuro-oncology in hopes of one day transitioning to the pharmaceutical industry.