Clinical laboratory techniques based on histology have endured as gold standards in clinical diagnostics, especially in determining the pathological nature of neoplasms, and monitoring the progression and treatment of metastatic diseases. A major portion of the histology toolbox relies on immunohistochemistry (IHC), where biomolecules expressed on cell surfaces interact with antibody-coupled dyes. Typically, these IHC-stained samples are qualitatively analyzed using optical microscopy. Although IHC can paint a more complete picture of cellular proliferation and phenotype than is possible with morphological assessment alone, pathologist interpretation techniques are not standardized and diagnosis is subjective. The quantitation of molecular expression in tissue samples can have paradigm shifting effects on cancer patient care. Quantitative results are objective and will significantly reduce inconsistencies that can occur during IHC preparation and pathologist assessments. In addition to standardized diagnosis, quantitative IHC can improve the treatment planning of molecular-based therapies by assessing the susceptibility of both cancerous (treatment success) and normal healthy tissues (side-effects) to molecularly-targeted drugs. In recent years, these benefits have spurned several quantitative IHC strategies, primarily based on analysis of optical microscopy images. However, these techniques cannot reliably account for competing endogenous contrast (dominated by optical scattering) that diminishes their accuracy. In addition, these techniques suffer from photobleaching or fluorescence quenching, contributing to additional inaccuracy. Overall, the limitations of current quantitative IHC techniques have restricted their suitability for clinical use. We propose a fundamentally differen and commercially viable quantitative IHC approach that is based on quantitative photoacoustic (qPA) microscopy, which we recently developed and demonstrated. Our quantitative IHC approach uses molecularly-targeted nanoparticles (NPs) to provide temporally consistent qPA contrast. Our bioconjugated NPs are used to provide molecularly-specific labeling of histological tissue sections without the need for blocking steps, making sample preparation much faster. After NP labeling, the tissues (either formalin fixed or cryopreserved) are rinsed of unbound NPs and prepared for qPA imaging. Automated qPA acquisition is used to produce spatially resolved quantitative image of molecular expression that is unaffected by endogenous contrast. By providing accurate and reproducible molecular quantitation, our proposed qPA- IHC platform can standardize pathological diagnoses and be used for improved treatment planning of molecular-based therapies. By providing the results quickly, our technique can also be used for guidance during surgeries, when biopsies cannot be obtained a priori.
Public Health Relevance Statement: Public Health Relevance: We propose the development of a new clinical quantitative immunohistochemistry platform based on photoacoustic microscopy and bioconjugated nanoparticles. Our nanoparticle-based immunohistochemistry sample preparation can be performed up to 30x faster than current clinical practices, and photoacoustic microscopy provides a way to quantify cancer biomarker expression. Overall, quantitative photoacoustic immunohistochemistry can improve diagnosis and treatment planning for cancer patients.
Project Terms: Accounting; Adverse effects; Amendment; Antibodies; antibody conjugate; base; Benchmarking; Biological Markers; Biopsy; cancer biomarkers; Cancer Patient; Cancerous; Cell Proliferation; Cell surface; Clinical; Clinical Laboratory Techniques; clinical practice; Coupled; design; Development; Diagnosis; Diagnostic; Disease; Drug Targeting; Dyes; Fluorescence; Formalin; Goals; Gold; Histology; Image; Immunohistochemistry; improved; Label; Laboratories; Malignant Neoplasms; Methods; Microscope; microscopic imaging; Microscopy; Molecular; Monitor; nanoparticle; Nanosphere; Nature; Neoplasms; Operative Surgical Procedures; Optics; Paint; Pathologist; Patient Care; Phase; Phenotype; photoacoustic imaging; Photobleaching; Predisposition; Preparation; Protocols documentation; prototype; public health relevance; quantitative imaging; research study; Sampling; Signal Transduction; Silicon Dioxide; Small Business Innovation Research Grant; Staining method; Stains; success; System; Techniques; Time; Tissue Sample; Tissues; treatment planning; Variant; Work
