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Open Access Publications from the University of California

Scholarly Works (4 results)

  • Thesis
  • Peer Reviewed
UC Irvine Electronic Theses and Dissertations (2022)

Multiplexed mRNA and protein profiling in the spatial context provides important new information enabling basic research and clinical applications. Unfortunately, most existing spatial transcriptomics and proteomics methods are limited due to either low multiplexing or assay complexity. Here, we introduce a new spatialomics technology, termed Multi Omic Single-scan Assay with Integrated Combinatorial Analysis (MOSAICA), that integrates in situ labeling of mRNA and protein markers in cells or tissues with combinatorial fluorescence spectral and lifetime encoded probes, spectral and time-resolved fluorescence imaging, and machine learning-based target decoding. This technology is the first application combining the biophotonic techniques, Spectral and Fluorescence Lifetime Imaging and Microscopy (FLIM), to the field of spatial transcriptomics and proteomics. By integrating the time dimension with conventional spectrum-based measurements, MOSAICA enables direct and highly multiplexed in situ spatial biomarker profiling in a single round of staining and imaging while providing error correction removal of background autofluorescence. We demonstrated MOSAICA’s capabilities in cell culture and Formalin-Fixed Paraffin-Embedded (FFPE) tissues while obtaining a strong correlation with sequencing data (Pearson’s r = 0.96). We then demonstrate simultaneous co-detection of protein and mRNA in colorectal cancer cells. To answer biological questions with a simple 3-4 plex immunofluorescence panel, we developed a low-cost Tissue Imager for under $9,000 and achieved a performance on par with commercial fluorescence microscopes that cost ~20x more. Additionally, another tool we developed to study the tissue microenvironment was cell-based mechanosensors to quantitively and dynamically assess the tissue mechanics. We have already used MOSAICA to study colon cancer heterogeneity, profile neurological mRNA panels in brain tissue, and profile immuno-oncology panels for skin tissue. MOSAICA represents a simple, versatile, and scalable tool for targeted spatial transcriptomics and proteomics analysis that can find broad utility in constructing human cell atlases, elucidating biological and disease processes in the spatial context, and serving as companion diagnostics for stratified patient care.

    Cover page: Developing Spatialomics Platforms to Profile BiomarkersCreative Commons 'BY-NC-ND' version 4.0 license
    • Article
    • Peer Reviewed
    UC Irvine Previously Published Works (2023)
    To image 4-plex immunofluorescence-stained tissue samples at a low cost with cellular level resolution and sensitivity and dynamic range required to detect lowly and highly abundant targets, here we describe a robust, inexpensive (<$9000), 3D printable portable imaging device (Tissue Imager). The Tissue Imager can immediately be deployed on benchtops for in situ protein detection in tissue samples. Applications for this device are broad, ranging from answering basic biological questions to clinical pathology, where immunofluorescence can detect a larger number of markers than the standard H&E or chromogenic immunohistochemistry (CIH) staining, while the low cost also allows usage in classrooms. After characterizing our platform's specificity and sensitivity, we demonstrate imaging of a 4-plex immunology panel in human cutaneous T-cell lymphoma (CTCL) formalin-fixed paraffin-embedded (FFPE) tissue samples. From those images, positive cells were detected using CellProfiler, a popular open-source software package, for tumor marker profiling. We achieved a performance on par with commercial epifluorescence microscopes that are >10 times more expensive than our Tissue Imager. This device enables rapid immunofluorescence detection in tissue sections at a low cost for scientists and clinicians and can provide students with a hands-on experience to understand engineering and instrumentation. We note that for using the Tissue Imager as a medical device in clinical settings, a comprehensive review and approval processes would be required.
      Cover page: A Low-Cost Modular Imaging System for Rapid, Multiplexed Immunofluorescence Detection in Clinical Tissues
      • Article
      • Peer Reviewed
      UC Irvine Previously Published Works (2021)
      We examined the association between endogenous opioid β-endorphin, cancer progression and pain in a transgenic mouse model of breast cancer, with a rat C3(1) simian virus 40 large tumor antigen fusion gene (C3TAg). C3TAg mice develop ductal epithelial atypia at 8 weeks, progression to intra-epithelial neoplasia at 12 weeks, and invasive carcinoma with palpable tumors at 16 weeks. Consistent with invasive carcinoma at 4 months of age, C3TAg mice demonstrate a significant increase in hyperalgesia compared to younger C3TAg or control FVBN mice without tumors. Our data show that the growing tumor contributes to circulating β-endorphin. As an endogenous ligand of mu opioid receptor, β-endorphin has analgesic activity. Paradoxically, we observed an increase in pain in transgenic breast cancer mice with significantly high circulating and tumor-associated β-endorphin. Increased circulating β-endorphin correlates with increasing tumor burden. β-endorphin induced the activation of mitogenic and survival-promoting signaling pathways, MAPK/ERK 1/2, STAT3 and Akt, observed by us in human MDA-MB-231 cells suggesting a role for β-endorphin in breast cancer progression and associated pain.
        Cover page: β-endorphin at the intersection of pain and cancer progression: Preclinical evidence
        • Article
        • Peer Reviewed
        UC Irvine Previously Published Works (2022)
        Multiplexed mRNA profiling in the spatial context provides new information enabling basic research and clinical applications. Unfortunately, existing spatial transcriptomics methods are limited due to either low multiplexing or complexity. Here, we introduce a spatialomics technology, termed Multi Omic Single-scan Assay with Integrated Combinatorial Analysis (MOSAICA), that integrates in situ labeling of mRNA and protein markers in cells or tissues with combinatorial fluorescence spectral and lifetime encoded probes, spectral and time-resolved fluorescence imaging, and machine learning-based decoding. We demonstrate MOSAICA's multiplexing scalability in detecting 10-plex targets in fixed colorectal cancer cells using combinatorial labeling of five fluorophores with facile error-detection and removal of autofluorescence. MOSAICA's analysis is strongly correlated with sequencing data (Pearson's r = 0.96) and was further benchmarked using RNAscopeTM and LGC StellarisTM. We further apply MOSAICA for multiplexed analysis of clinical melanoma Formalin-Fixed Paraffin-Embedded (FFPE) tissues. We finally demonstrate simultaneous co-detection of protein and mRNA in cancer cells.
          Cover page: Spatial transcriptomics using combinatorial fluorescence spectral and lifetime encoding, imaging and analysis
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