Ultrasensitive proteomic quantitation of cellular signaling by digitized nanoparticle-protein counting.
Many important signaling and regulatory proteins are expressed at low abundance and are difficult to measure in single cells. We report a molecular imaging approach to quantitate protein levels by digitized, discrete counting of nanoparticle-tagged proteins. Digitized protein counting provides ultrasensitive molecular detection of proteins in single cells that surpasses conventional methods of quantitating total diffuse fluorescence, and offers a substantial improvement in protein quantitation. We implement this digitized proteomic approach in an integrated imaging platform, the single cell-quantum dot platform (SC-QDP), to execute sensitive single cell phosphoquantitation in response to multiple drug treatment conditions and using limited primary patient material. The SC-QDP: 1) identified pAKT and pERK phospho-heterogeneity and insensitivity in individual leukemia cells treated with a multi-drug panel of FDA-approved kinase inhibitors, and 2) revealed subpopulations of drug-insensitive CD34+ stem cells with high pCRKL and pSTAT5 signaling in chronic myeloid leukemia patient blood samples. This ultrasensitive digitized protein detection approach is valuable for uncovering subtle but important differences in signaling, drug insensitivity, and other key cellular processes amongst single cells.
Department of Biomedical Engineering, Oregon Health &Science University, Portland OR 97201, USA.|OHSU Center for Spatial Systems Biomedicine, Oregon Health &Science University, Portland OR 97201, USA.
Thomas
T
Jacob
Division of Hematology and Medical Oncology, The Knight Cancer Institute, Oregon Health &Science University, Portland OR 97239, USA.
Anupriya
A
Agarwal
Department of Biomedical Engineering, Oregon Health &Science University, Portland OR 97201, USA.|OHSU Center for Spatial Systems Biomedicine, Oregon Health &Science University, Portland OR 97201, USA.
Damien
D
Ramunno-Johnson
Division of Hematology and Hematologic Malignancies, Huntsman Cancer Institute, University of Utah, 2000 Circle of Hope, Salt Lake City, Utah 84112, USA.
Thomas
T
O'Hare
Department of Biomedical Engineering, Oregon Health &Science University, Portland OR 97201, USA.|Department of Industrial Engineering, Koc University, Istanbul, Turkey.
Mehmet
M
Gönen
Division of Hematology and Medical Oncology, The Knight Cancer Institute, Oregon Health &Science University, Portland OR 97239, USA.|Department of Cell Developmental &Cancer Biology, Oregon Health &Science University, Portland OR 97239, USA.
Jeffrey W
JW
Tyner
Division of Hematology and Medical Oncology, The Knight Cancer Institute, Oregon Health &Science University, Portland OR 97239, USA.|Howard Hughes Medical Institute, Portland, Oregon, USA.
Brian J
BJ
Druker
Department of Biomedical Engineering, Oregon Health &Science University, Portland OR 97201, USA.|OHSU Center for Spatial Systems Biomedicine, Oregon Health &Science University, Portland OR 97201, USA.|Division of Hematology and Medical Oncology, The Knight Cancer Institute, Oregon Health &Science University, Portland OR 97239, USA.
Tania Q
TQ
Vu
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D000069439:Dasatinib / Q000494:pharmacology*; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000473:pathology*; D053758:Nanoparticles / Q000737:chemistry*; D009363:Neoplasm Proteins / Q000032:analysis*; D009687:Nuclear Proteins / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D040901:Proteomics / Q000379:methods; D045663:Quantum Dots; D050799:STAT5 Transcription Factor / Q000378:metabolism*; D015398:Signal Transduction; D025521:Tumor Suppressor Proteins / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D000970:Antineoplastic Agents; C082869:CRKL protein; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D047428:Protein Kinase Inhibitors; D050799:STAT5 Transcription Factor; C494092:STAT5A protein, human; D025521:Tumor Suppressor Proteins; D000069439:Dasatinib