California Institute of Technology

Other

Long Cai

Sheel Mukesh Shah

Eric Lubeck

Chang Ho Sohn

Timur Zhiyentayev

Ting-Fang He

Ahmet F Coskun

Niles A Pierce

Michael B Elowitz

Mark William Budde

James Linton

Wen Jie Zhou

Harry Ming Tak Choi

Naeem S Husain

Amjad Askary

Carlos Lois

Luis Sanchez Guardado

Yandong Zhang

Ahmet Coskun

Chee Huat Eng

The present invention, among other things, provides technologies for detecting and/or quantifying nucleic acids in cells, tissues, organs, or organisms. Pre-designed barcodes are associated with specific molecular targets through sequential hybridization experiments. A pseudo-color based barcoding scheme is described that overcomes the limitations in the previous generation of the technology. The current method can be applied to both in vitro and in situ analysis.

Sequential probing of molecular targets based on pseudo-color barcodes with embedded error correction mechanism

The present invention, among other things, provides technologies for detecting and/or quantifying nucleic acids in cells, tissues, organs or organisms. In some embodiments, through sequential barcoding, the present invention provides methods for high-throughput profiling of a large number of targets, such as transcripts and/or DNA loci.

Multiplex labeling of molecules by sequential hybridization barcoding

The present application describes compositions and methods for identifying and quantitating molecular targets within a cellular environment. Specifically, provided herein are compositions and methods for separately identifying and quantifying each of one or more molecular targets from a single cell. More specifically, provided herein are compositions and methods for separately identifying and quantifying the same molecular target from a single cell.

Quantitative analysis of single-cell proteins by identification and quantification separation (duet)

A multi-channel line microscope for single molecule Fluorescence In Situ Hybridization (FISH) imaging of a sample. A microscope stage moves a sample across two or more reflected excitation lines positioned relative to each other so that each excitation line excites a spatially distinct horizontal line in the image plane of the sample. A sample is imaged by moving the microscope stage across two or more reflected excitation lines positioned relative to each other so that each excitation line excites a spatially distinct horizontal line in the image plane of the sample. The apparatus and methods of use are suitable for a broad range of applications.

Multi-channel line scanner for fast large field and confocal imaging

Disclosed herein include systems, methods, compositions, and kits for in situ readout of barcodes, such as DNA barcodes. Barcode constructs containing a promoter (e.g., a phage promoter) that is inactive in live cells can be integrated in the genomes of cells. Cells can be fixed, and phage RNA polymerase can be used for transcription of the barcode to RNA transcripts. The RNA transcripts can be detected using, for example, fluorescent imaging and used to determine barcode sequences.

In situ readout of DNA barcodes

Disclosed herein are methods and systems for analyzing visual data from multiple rounds of hybridization interactions where the same molecular target is detected by probes with different detectable labels. In particular, disclosed herein are methods and systems for analyzing sequential hybridization images for molecular profiling, where the images are obtained using multiplex fluorescence in situ hybridization (FISH).

Multiplex analysis of molecules in single cells by image correlation

The present disclosure relates to methods involving HCR reactions that involve using trigger oligos to activate probes that initiate HCR.

Immunohistochemistry via hybridization chain reaction

Disclosed herein are methods and systems for detecting and/or quantifying cellular targets such as nucleic acids in cells, tissues, organs or organisms. Through sequential barcoding, it is possible to perform high-throughput profiling of a large number of targets, such as transcripts and/or DNA loci. In some embodiments, error correction is implemented through use of barcodes that can tolerate mistakes and missing data during sequential hybridization of probes to selected targets.

Error correction of multiplex imaging analysis by sequential hybridization

Methods and systems are provided for creating molecular barcodes or indicia for cellular constituents within single cells and for resolving such barcodes or indicia with super resolution technologies such as super resolution microscopy. By this approach, numerous molecular species that can be measured simultaneously in single cells. It has been demonstrated that multiple mRNA transcripts can be labeled with a spatially ordered sequence of fluorophores, and that barcode can be resolved. The methods and systems can be used for genome-wide transcriptional profiling in individual cells by super-resolution barcoding and suggest a general strategy to bring large-scale-omics approach into single cells.

Molecular indicia of cellular constituents and resolving the same by super-resolution technologies in single cells

Methods and systems are provided for creating molecular barcodes or indicia for cellular constituents within single cells and for resolving such barcodes or indicia with super-resolution technologies such as super-resolution microscopy. By this approach, numerous molecular species that can be measured simultaneously in single cells. It has been demonstrated that multiple mRNA transcripts can be labeled with a spatially ordered sequence of fluorophores, and that barcodes can be resolved. In addition, alternative splicing events can be characterized by identifying and quantifying mRNA isoforms in an individual cell.

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Pasadena, CA, United States of America

Long Cai