Discovery Lab

Interrogation of cancer cells, stromal cells and immune cells in the tumor microenvironment and blood is crucial to developing novel therapeutic strategies and resolve limitations in existing cancer therapies.

The 10X genomics platform is a facile and powerful platform to discover novel immune subsets and transform our understanding the tumor microenvironment. We apply this high throughput state-of-the-art 10X genomics technology to interrogate cellular heterogeneity, intrinsic variation in TCR repertoire, define cellular states, characterize HLA genotype, and examine functional dynamics across cell types in many disease states.

In the Discovery Lab, we utilize a variety of cancer tissues, core biopsies, and blood to prepare genomic libraries for gene expression, V(D)J (TCR and Ig). The Discovery Lab uses an Agilent Bioanalyzer system for genomic library quality control, and DNA/RNA quantitation.

We are using combined cellular indexing of transcriptomes and epitope by sequencing ( CITE-seq), which provides additional ability to resolve proteome and transcriptome in greater detail.

We utilize the MiSeq Illumina sequencing system perform next generation DNA sequencing. It integrates cluster generation, sequencing and data analysis. MiSeq workflow allows upto 30 million reads in single run. The MiSeq system also combines proven sequencing by synthesis (SBS) technology with a workflow from DNA to data analysis in few hours. The sequencing data has broad range of applications in immunotherapy.

This sequencing platform allows flexibility and quick access for targeted re-resequencing, 16S metagenomics, small genome sequencing.

We perform TCR sequencing and HLA typing for collaborative projects or as a Core service.

Multicolor flow cytometry is one of the most robust and powerful technologies for immunology. It simultaneously utilizes a diverse range of fluorescent markers to discover cellular subsets, regulatory proteins, and detects changes in cell surface and intracellular proteins in diverse range of cell types.

We utilize a new 5 LASER BD FACSymphonyTM, machine to detect up to 23 fluorescent color simultaneously across millions of cells. This resides in the Immunomonitoring laboratory of CITI. Discovery Lab collaborations center work that required non-standard panels not already offered by the Immunomoitoring laboratory.

Our FACSymphonyTM, is also equipped with a high-throughput sampler to efficiently run samples in 96 well plates.

We routinely utilize this technology to interrogate a range of cells, including rare neoantigen specific T cells, and develop novel immunomonitoring assays. We measure cell activation states in in-vitro experiments, and perform massive immuno-profiling of immuno-therapy treated cancer patients.

Cancer neoantigen-specific T cells play a dominant role in determining the efficacy of PD-1-, CTLA-4- and PD-L1-blocking therapeutic antibodies in several cancers. Due to neo-antigens' potential to develop robust and specific T cell responses, they are considered high value targets of personalized vaccine candidate. In the Discovery Lab, we screen and validate neoantigens.

We utilize a tandem minigene approach to clone and prepare transcripts of all detected mutations in patients. We transfect RNA containing mutations into patients' dendritic cells and detect the activation of patient's mutation specific T cells in autologous settings. We also utilize mass spectrometry methods to detect and validate antigen specific T cells.

In several immunological diseases, cellular function is defined by cellular gene expression and by the location of immune cells within the tissue architecture. Spatial transcriptomics reveals the location of active genes in immune cells found in specific regions and disease tissue. In addition to single-cell-based RNA profiling in spatial transcriptomics, we interrogate simultaneously the expression of gene sets in distinct regions of clinical specimens (in development).

CD8 cytotoxic T lymphocytes (CTL) and natural killer (NK) cells play an important role in the control of microbial infections and tumors. Cytolysis of antigen-specific 51Cr-labeled targets in the standard assay help to quantify, using traditional gamma counters, 51Cr released into the supernatant and collected from CTL cultures.

Our lytic assay system reduces the amount of dry and mixed radioactive waste generated while using the same instrument for gamma- and beta-emitting isotopes. We utilize Lumaplates to substantially reduce radioactive waste for environmental concern. This high-throughput and safe assay methodology enables us to perform most critical immunological test in our cancer neoantigen vaccine discovery program.

Immunotherapy modulates immune cell types and several soluble factors. The immune cell activation states, cross-talk and cellular proliferation is highly dependent on soluble factors.

We utilize the robust Luminex™ 200™ Instrument System to analyze soluble factors in clinical specimens. The Luminex 200 Instrument System also sets the standard for multiplexing, providing the ability to perform up to 100 different tests in a single reaction volume on a flow cytometry-based platform. The Luminex 200 system is also compatible with Procarta Plex multiplex immunoassays and QuantiGene Plex multiplex gene expression assays.

The open architecture of Luminex xMAP technology uses imaging, microspheres, digital signal processing and traditional chemistry, combining proven technologies in a unique way.

Our Luminex platform utilizes small sample input and enables fast, reproducible results from favorable kinetics of the liquid bead array approach.

Our Luminex provides broad coverage of applications, including protein expression and gene expression profiling in clinical trial specimens and can reveal novel biomarkers to understand disease scenario and therapeutic outcomes.