Document text
Principal Investigator: Russell H Cole
Organization: SCRIBE BIOSCIENCES, INC.
Fiscal Year: 2023
Award: $300,000
Funding agency: National Institute of Allergy and Infectious Diseases
ABSTRACT
Scribe Biosciences are leading experts in the field of droplet microfluidics and have developed a best-in-class
droplet manipulation platform, Microenvironment on Demand (MOD), that can currently assemble >100k
paired-cell assays in <3 hours, with proven proof of concept. Using this innovative technology, this SBIR Phase
1 project proposes the development and quantification of assay methods to be used for single-cell functional
screening workflows to enable large scale screening of therapeutic antibody (Ab) candidates. The development
of such a workflow to reliably, consistently, and repeatably identify large and diverse pools of B cell hits would
offer a significant advantage over the classical but inefficient hybridoma method. Porting direct B-cell assays to
microfluidics is a natural fit because short lived B-cells can rapidly generate significant secreted Ab
concentrations when incubated in appropriately small volumes; current attempts are limited by cost and
scalability, and none offer high throughput (HT) assays against target cells, sensitive assays, or integrated HT
sequencing. MOD represents an evolutionary advancement in the capability to build droplet-based cell assays
with precision and scale, effectively integrating assay construction, readouts, hit selection, and sample prep into
a single workflow and instrument. MOD co-encapsulates Ab-secreting and target cells in the same microfluidic
droplet, which enables building an assay based on the target cell, since it will carry along the Ab-secreting cell
and therefore the RNA that is available to identify the Ab in a subsequent sequencing step. MOD utilizes flow
cytometry-style detection and sorting, so it is readily scalable for HT. The approach for this project has been
informed by previous work developing assays on the MOD platform. In the first aim, two assays will be developed
to detect Ab binding against membrane protein targets. The first will adopt an existing bead-based no wash
assay scheme for use with high copy number targets, and the second will develop a more sensitive assay for
low copy number targets with a wash step, and will explore the appropriate method for creating a durable physical
linkage between the cells that will last through FACS sorting or re-encapsulation. The second aim will test and
quantify the system with B-cells from immunized mice for a real-world demonstration of Ab discovery. B-cells will
be sourced from standard 4-week immunization protocols on groups of 3 mice using SARS-CoV-2 as the antigen,
and will be used to explore the parameters of primary B-cell culture in droplets and other factors associated with
porting B-cell biology on to the MOD platform. A small batch (50-100k) of B-cell/target cell assays will be tested,
and assuming that the HT of the platform will correlate with a high number of hits (~1000 positive assays), a
small number (~10) of Ab candidates will be bioinformatically selected for subsequent re-cloning and hit
validation. Successful MOD-enabled antibody screening would introduce a new paradigm in the capabilities of
researchers to identify a larger and more diverse field of Ab candidates, bypassing current limitations of cost,
scalability, commercial availability, or technical complexity, and ultimately leading to better therapies.
Terms: <2019 novel corona virus><2019 novel coronavirus><2019-nCoV><2019-nCoV S protein><2019-nCoV spike glycoprotein><2019-nCoV spike protein><Adopted><Antibodies><Antibody Therapy><Antibody-Secreting Cells><Antigen-Presenting Cells><Antigens><Assay><B blood cells><B cell><B cells><B-Cells><B-Lymphocytes><B-cell><BALB C Mouse><BALB/c><Binding><Bio-Informatics><Bioassay><Bioinformatics><Biologic Assays><Biologic Sciences><Biological Assay><Biological Sciences><Biology><Bioscience><Biotin><Bypass><CHO Cells><COVID-19 S protein><COVID-19 spike glycoprotein><COVID-19 spike protein><COVID-19 virus><COVID19 S protein><COVID19 spike glycoprotein><COVID19 spike protein><COVID19 virus><Cancers><Cardiology><Cell Body><Cell Culture Techniques><Cell Line><Cell membrane><Cell secretion><CellLine><Cells><Cellular Assay><Cellular Secretion><Cellular biology><Chinese Hamster Ovary Cell><Clinical Treatment Moab><Cloning><CoV-2><CoV2><Computer Analysis><Cytoplasmic Membrane><DNA Molecular Biology><Detection><Development><Devices><Disparate><Encapsulated><Engineering><Ensure><Flow Cytofluorometries><Flow Cytofluorometry><Flow Cytometry><Flow Microfluorimetry><Flow Microfluorometry><G Protein-Complex Receptor><G Protein-Coupled Receptor Genes><G-Protein-Coupled Receptors><GPCR><Genomics><High Throughput Assay><High-Throughput Nucleotide Sequencing><High-Throughput Sequencing><Hour><Hybridomas><Hydrogels><Immunization><Immunize><Immunoglobulin-Secreting Cells><Inbred BALB C Mice><Incubated><Infection><Investigators><Jurkat Cells><Life Sciences><Malignant Neoplasms><Malignant Tumor><Medical><Membrane Protein Gene><Membrane Proteins><Membrane-Associated Proteins><Methods><Mice><Mice Mammals><Microfluidics><Molecular Biology><Molecular Interaction><Monoclonal Antibodies><Murine><Mus><Neurology><Non-Polyadenylated RNA><Oligo><Oligonucleotides><Performance><Permeability><Phase><Plasma Membrane><Population><Preparation><Process><Property><Proteins><Protocol><Protocols documentation><RNA><RNA Gene Products><Reagent><Research><Research Personnel><Researchers><Ribonucleic Acid><Running><SARS corona virus 2><SARS-CO-V2><SARS-COVID-2><SARS-CoV-2><SARS-CoV-2 S protein><SARS-CoV-2 spike glycoprotein><SARS-CoV-2 spike protein><SARS-CoV2><SARS-CoV2 S protein><SARS-CoV2 spike glycoprotein><SARS-CoV2 spike protein><SARS-associated corona virus 2><SARS-associated coronavirus 2><SARS-coronavirus-2><SARS-related corona virus 2><SARS-related coronavirus 2><SARSCoV2><SBIR><Sampling><Scheme><Series><Severe Acute Respiratory Coronavirus 2><Severe Acute Respiratory Distress Syndrome CoV 2><Severe Acute Respiratory Distress Syndrome Corona Virus 2><Severe Acute Respiratory Distress Syndrome Coronavirus 2><Severe Acute Respiratory Syndrome CoV 2><Severe Acute Respiratory Syndrome-associated coronavirus 2><Severe Acute Respiratory Syndrome-related coronavirus 2><Severe acute respiratory syndrome associated corona virus 2><Severe acute respiratory syndrome coronavirus 2><Severe acute respiratory syndrome coronavirus 2 S protein><Severe acute respiratory syndrome coronavirus 2 spike glycoprotein><Severe acute respiratory syndrome coronavirus 2 spike protein><Severe acute respiratory syndrome related corona virus 2><Small Business Innovation Research><Small Business Innovation Research Grant><Sorting><Source><Speed><Staining method><Stains><Strains Cell Lines><Surface><Surface Proteins><Suspension substance><Suspensions><System><T-Cells><T-Lymphocyte><Techniques><Technology><Testing><Therapeutic><Therapeutic antibodies><Titrations><Validation><Variant><Variation><Vitamin H><Work><Wuhan coronavirus><accessory cell><antibody and antigen binding><antibody assay><antibody based detection><antibody based test><antibody based therapies><antibody detection><antibody test><antibody treatment><antibody-based therapeutics><antibody-based treatment><assay development><cell assay><cell biology><cell culture><cell cultures><coenzyme R><combinatorial><commercialization><computational analyses><computational analysis><computer analyses><coronavirus disease 2019 S protein><coronavirus disease 2019 spike glycoprotein><coronavirus disease 2019 spike protein><coronavirus disease 2019 virus><coronavirus disease-19 virus><cost><cultured cell line><detect antibodies><developmental><experiment><experimental research><experimental study><experiments><flexibility><flexible><flow cytophotometry><hCoV19><high throughput screening><immunogen><innovate><innovation><innovative><innovative technologies><instrument><lentiviral-transduced><lentivirally transduced><lentivirus transduced><mAbs><malignancy><microfluidic technology><monoclonal Abs><nCoV2><neoplasm/cancer><new technology><novel technologies><oligos><plasmalemma><preparations><scRNA-seq><screening><screenings><single cell RNA-seq><single cell RNAseq><single cell expression profiling><single cell transcriptomic profiling><single-cell RNA sequencing><success><thymus derived lymphocyte><validations><µfluidic><µfluidic technology>