Exceptionally Innovative Tools and Technologies for Single Cell Analysis (R21)

Exceptionally Innovative Tools and Technologies for Single Cell Analysis (R21) is sponsored by NIH Common Fund. This opportunity supports mission-aligned projects and measurable outcomes.

Expired RFA-RM-13-021: Exceptionally Innovative Tools and Technologies for Single Cell Analysis (R21) This notice has expired. Check the NIH Guide for active opportunities and notices. of Health and Human Services Part 1.

Overview Information Participating Organization(s) National Institutes of Health ( NIH ) of Participating Organizations Office of Strategic Coordination ( Common Fund ) This Funding Opportunity Announcement (FOA) is developed as a Common Fund initiative ( http://commonfund. nih.

gov/ ) through the Office of Strategic Coordination, Division of Program Coordination, Planning, and Strategic Initiatives, Office of the NIH Director ( http://dpcpsi. nih. gov/osc/ ).

This FOA will be administered by the National Institute of Mental Health ( NIMH ) on behalf of the NIH.

Funding Opportunity Title Exceptionally Innovative Tools and Technologies for Single Cell Analysis (R21) R21 Exploratory/Developmental Research Grant Funding Opportunity Announcement (FOA) Number Companion Funding Opportunity R33 Exploratory/Developmental Grants Phase II RFA-RM-13-022 , R01 Research Project RFA-RM-13-023 , U01 Research Project Cooperative Agreements Additional Information on Eligibility .

Catalog of Federal Domestic Assistance (CFDA) Number(s) Funding Opportunity Purpose This Funding Opportunity Announcement (FOA) issued by the National Institutes of Health, solicits early stage, high-risk/high-impact applications to develop next-generation tools that distinguish heterogeneous states among cells in situ.

Applications should define the current state of technology as a benchmark against which the new tool(s) will be measured and should propose proof-of-concept testing of the tool(s) in a complex biological tissue or living organism. The new tools should provide substantially increased sensitivity, selectivity, spatiotemporal resolution, scalability of multiple global or functional measures of single cells.

A particular emphasis for this FOA is on measures that minimize cell perturbation and permit viability of cells for repeated measures over time.

These novel technologies will aid in obtaining a fine-grained, integrative and dynamic view of heterogeneous cellular states/classes and will provide innovative platforms to transform research into the cellular basis of Open Date (Earliest Submission Date) Letter of Intent Due Date(s) April 4, 2014, by 5:00 PM local time of applicant Applicants are encouraged to apply early to allow adequate time to make any corrections to errors found in the application during the submission process by the due date.

AIDS Application Due Date(s) April 4, 2014, by 5:00 PM local time of applicant Applicants are encouraged to apply early to allow adequate time to make any corrections to errors found in the application during the submission process by the due date.

Required Application Instructions It is critical that applicants follow the instructions in (R&R) Application Guide , except where instructed to do otherwise (in this FOA or in a Notice from the NIH Guide for Grants and Contracts ). Conformance to all requirements (both in the Application Guide and the FOA) is required and strictly enforced.

Applicants must read and follow all application instructions in the Application Guide as well as any program-specific instructions noted in Section IV . When the program-specific instructions deviate from those in the Application Guide, follow the program-specific instructions. Applications that do not comply with these instructions may be delayed or not accepted for review.

Part 1. Overview Information Part 2. Full Text of the Announcement Section I.

Funding Opportunity Description Section II. Award Information Section III. Eligibility Information Section IV.

Application and Submission Section V. Application Review Information Section VI. Award Administration Information Section VII.

Agency Contacts Section VIII. Other Information Full Text of Announcement Section I. Funding Opportunity Description This Funding Opportunity Announcement (FOA) issued by the National Institutes of Health, solicits early stage, high-risk/high-impact applications to develop next-generation tools that distinguish heterogeneous states among cells in situ.

Applications should define the current state of technology as a benchmark against which the new tool(s) will be measured and should propose proof-of-concept testing of the tool(s) in a complex biological tissue or living organism. The new tools should provide substantially increased sensitivity, selectivity, spatiotemporal resolution, scalability of multiple global or functional measures of single cells.

A particular emphasis for this FOA is on measures that minimize cell perturbation and permit viability of cells for repeated measures over time. These novel technologies will aid in obtaining a fine-grained, integrative and dynamic view of heterogeneous cellular states/classes and will provide innovative platforms to transform research into the cellular basis of diseases.

This initiative is part of the Single Cell Analysis Program (SCAP) and is funded through the NIH Common Fund (See http://nihroadmap. nih. gov/ ), which supports cross-cutting programs that are expected to have exceptionally high impact.

Common Fund initiatives address key roadblocks in biomedical research that impede basic scientific discovery and its translation into improved human health. In addition, these programs capitalize on emerging opportunities to catalyze progress across multiple biomedical fields.

Single cell analysis has recently emerged as an important field of research because technologies have improved in sensitivity and throughput sufficiently to begin measuring and understanding heterogeneity in complex biological systems and correlating it with changes in biological function and disease processes.

By profiling individual cells it is possible to resolve rare cells, transient cell states, and the influence of organization and environment on such cells and states, which cannot be described by ensemble measurements.

The long-term goal of the SCAP is to accelerate this move towards personalizing health to the cellular level by understanding the link between cell heterogeneity, tissue function and emergence of disease through the discovery, development and translation of innovative approaches which will dramatically change the way cells are characterized.

The SCAP is focusing on supporting work which will systematically measure, analyze and model cell-to-cell variation, and identify crucial differences and rare biological states, which may have important functional consequences. To robustly and systematically describe cell level heterogeneity, projects are expected to take a multiplexed approach with minimal perturbation, which can be applied reproducibly to any complex tissue.

Technologies and methods must also be capable of capturing spatiotemporal information to understand the organization, evolution and response of cell states as part of a functional population.

In addition, the SCAP emphasizes the application of these technologies to in situ populations of cells from multicellular organisms to link cell state measurements with complex, functional tissues which can inform our understanding of disease processes.

The SCAP has been designed as a five-year program with several components: (1) the collection, analysis and sharing of comprehensive expression datasets to understand the role of heterogeneity in tissues and systemically and identify critical parameters and states; (2) the discovery of new, innovative tools for spatiotemporal imaging, manipulation, analysis and modeling of a biologically relevant population of cells with minimal perturbation; (3) milestone-driven validation and translation of technologies for characterizing single cells in situ meeting the needs of end-users; and (4) development and coordination of a multidisciplinary research community through workshops and other collective endeavors.

Further details can be found on the Program’s website ( http://commonfund. nih. gov/singlecell/ ).

This FOA solicits applications to develop next-generation, innovative technologies to better define cell heterogeneity in situ (e.g., slices, intact organs, whole organisms). These techniques should provide new analytical measures and manipulations of cellular contents, structure and activity significantly beyond those currently available at the single cell level.

Of particular interest for this FOA are first-in-class and/or cross-cutting techniques that minimize cell perturbation and permit viability of cells for repeated measures over time. The goal of this FOA is to accelerate early stage development of promising concepts by focusing on overcoming technical challenges, building prototype systems and testing performance on a population of cells in situ (e.g., explant, whole organism).

Though applications are particularly encouraged that propose first-in-class and/or cross-cutting techniques that minimize cell perturbation and permit cell viability for repeated measurements over time , applications may address other significant gap areas or may propose early-stage, high-risk/high-impact technologies, including, but not limited to: Combinations of tools for multiplex analysis and/or manipulation of single cells in situ to maximize data content over many parameters (e.g., proteins, metabolites, organelles, electrochemical dynamics, signal secretion/reception/transduction, cytoarchitecture or migratory changes).

Tools that provide significant advances in sensitivity, selectivity or spatiotemporal resolution of molecules/structures/activities within single cells and between ostensibly similar cells in situ (e.g., high resolution imaging of molecular interactions within single cells).

Automated and scalable assays for high-throughput analysis of single cells in situ, including scalability of measured parameters in parallel, cell numbers and/or speed of processing. Systems-level single cell dataset analysis or modeling, including computational approaches, in the context of tissues or whole organisms (e.g., multiscale models that link cell states to organ or circuit function).

Note: Applications should avoid proposing studies that are adequately addressed by currently funded SCAP grants (e.g., projects focused on RNA expression / transcriptomics); refer to the Common Fund Single Cell Analysis Program web site for information on currently supported projects ( http://commonfund. nih. gov/singlecell/fundedresearch.

aspx ). Applicants are strongly advised to solicit prior advice from the Scientific/Research contact below in determining the responsiveness of their application.

Research designs should focus on tool development, but tools may be tested in the context of heterogeneous biological systems, such as: identifying spatiotemporal transitions in cellular states (e.g., progenitor lineage determination, cellular aging); detection of rare cells in a population (e.g., stem cells, drug resistant cells); elucidating the molecular signatures or functional consequences of stochastic variation in cellular states (e.g., genomic stability, clonal selection and evolution, asymmetric division, cell specification); or characterizing heterogeneous cell responses to environmental changes (e.g. homeostatic perturbation, modulation of niche/microenvironment, morphogens or cell-to-cell signaling, toxicological exposure, experience-dependent plasticity, host cell responses to infectious, immunological or allergic challenges).

Applicants are expected to explain the significance of the specific technical approach with respect to characterizing a heterogeneous cell population in situ. However, any proposed biological assay(s) may be chosen strictly for utility (rather than biological novelty) in proof-of-concept testing of the innovative technique.

NIH understands and accepts that early stage, high-risk/high-impact applications that are responsive to this FOA will be much riskier than for most other FOAs. Applications will be considered based on the expected technological impact relative to risk. All applications must explicitly address considerations detailed in Section IV.

2 below (Application and Submission Information). Note that for grants funded through this FOA, the PD/PI is expected to attend the annual SCAP workshop meeting (likely to be held in the Bethesda area), which will be used to build synergy among the supported projects. Section II.

Award Information Grant: A support mechanism providing money, property, or both to an eligible entity to carry out an approved project or activity. Application Types Allowed Glossary and the SF424 (R&R) Application Guide provide details on Funds Available and Anticipated Number of Awards The number of awards is contingent upon NIH appropriations and the submission of a sufficient number of meritorious applications.

NIH intends to fund an estimate of 8-10 awards, corresponding to a total of $2,000,000, for fiscal year 2014. Future year amounts will depend on annual Direct costs shall not exceed $275,000 for the total project period or $200,000 in any single year. The maximum project period is 2 years.

described in the NIH Grants Policy Statement will apply to the applications submitted and awards made in response to this FOA.

Higher Education Institutions Public/State Controlled Institutions of Higher Education Private Institutions of Higher Education The following types of Higher Education Institutions are always encouraged to apply for NIH support as Public or Private Institutions of Higher Education: Black Colleges and Universities (HBCUs) Controlled Colleges and Universities (TCCUs) Native and Native Hawaiian Serving Institutions American Native American Pacific Islander Serving Institutions (AANAPISIs) Nonprofits Other Than Institutions of Higher Education Nonprofits with 501(c)(3) IRS Status (Other than Institutions of Nonprofits without 501(c)(3) IRS Status (Other than Institutions For-Profit Organizations (Other than Small Businesses) City or Township Governments Special District Governments Indian/Native American Tribal Governments (Federally Recognized) Indian/Native American Tribal Governments (Other than Federally Eligible Agencies of the Federal Government - NIH Intramural U.S. Territory or Possession Independent School Districts Public Housing Authorities/Indian Housing Authorities Native American Tribal Organizations (other than Federally recognized tribal governments) Faith-based or Community-based Organizations Non-domestic (non-U.S.) Entities (Foreign Institutions) Non-domestic (non-U.S.) Entities (Foreign Institutions) are eligible to apply.

Non-domestic (non-U.S.) components of U.S. Organizations are eligible Foreign components, as defined in the NIH Grants Policy Statement , are allowed. Applicant organizations must complete and maintain the following registrations as described in the SF 424 (R&R) Application Guide to be eligible to apply for or receive an award. All registrations must be completed prior to the application being submitted.

Registration can take 6 weeks or more, so applicants should begin the registration process as soon as Policy on Late Submission of Grant Applications states that failure to complete registrations in advance of a due date is not a valid reason for a Universal Numbering System (DUNS) - All registrations require that applicants be issued a DUNS number.

After obtaining a DUNS number, applicants can begin both SAM and eRA Commons registrations. The same DUNS number must be used for all registrations, as well as on the grant application. System for Award Management (SAM) (formerly CCR) Applicants must complete and maintain an active registration, which requires renewal at least annually .

The renewal process may require as much time as the initial registration. SAM registration includes the assignment of a Commercial and Government Entity (CAGE) Code for domestic organizations which have not already been assigned a Government Entity (NCAGE) Code Foreign organizations must obtain an NCAGE code (in lieu of a CAGE code) in order to register in SAM.

must have an active DUNS number and SAM registration in order to complete the eRA Commons registration. Organizations can register with the eRA Commons as they are working through their SAM or Grants. gov registration.

eRA Commons requires organizations to identify at least one Signing Official (SO) and at least one Program Director/Principal Investigator (PD/PI) account in order to must have an active DUNS number and SAM registration in order to complete the Directors/Principal Investigators (PD(s)/PI(s)) All PD(s)/PI(s) must have an eRA Commons account and should work with their organizational officials to either create a new account or to affiliate an existing account with the applicant organization’s eRA Commons account.

If the PD/PI is also the organizational Signing Official, they must have two distinct eRA Commons accounts, one for each role. Obtaining an eRA Commons account can take up to 2 weeks.

Eligible Individuals (Program Director/Principal Any individual(s) with the skills, knowledge, and resources necessary to carry out the proposed research as the Program Director(s)/Principal Investigator(s) (PD(s)/PI(s)) is invited to work with his/her organization to develop an application for support.

Individuals from underrepresented racial and ethnic groups as well as individuals with disabilities are always encouraged to apply for NIH support. For institutions/organizations proposing multiple PDs/PIs, visit the Multiple Program Director/Principal Investigator Policy and submission details in the Senior/Key Person Profile (Expanded) Component of the SF424 (R&R) Application Guide.

This FOA does not require cost sharing as defined in the NIH Grants Policy Statement . 3. Additional Information on Eligibility Applicant organizations may submit more than one application, provided that each application is scientifically distinct.

NIH will not accept any application that is essentially the same as one already reviewed within the past thirty-seven months (as described Grants Policy Statement ), except for submission: To an RFA of an application that was submitted previously as an investigator-initiated application but not paid; Of an investigator-initiated application that was originally submitted to an RFA but not paid; or Of an application with a changed grant activity code.

Section IV. Application and Submission Information Applicants must download the SF424 (R&R) application package associated with this funding opportunity using the Apply for Grant Electronically button in this FOA or following the directions provided at Grants. gov .

2. Content and Form of Application Submission It is critical that applicants follow the instructions in (R&R) Application Guide , except where instructed in this funding opportunity announcement to do otherwise. Conformance to the requirements in the Application Guide is required and strictly enforced.

Applications that are out of compliance with these instructions may be delayed or not accepted for For information on Application Submission and Receipt, visit Frequently Asked Questions Application Guide, Electronic Submission of Grant Although a letter of intent is not required, is not binding, and does not enter into the review of a subsequent application, the information that it contains allows IC staff to estimate the potential review workload and By the date listed in Part 1.

Overview Information , prospective applicants are asked to submit a letter of intent that includes the following information: Descriptive title of proposed activity Name(s), address(es), and telephone number(s) of the PD(s)/PI(s) Names of other key personnel Participating institution(s) Number and title of this funding opportunity The letter of intent should be sent to: David M. Panchision, Ph. D.

Division of Neuroscience and Basic Behavioral Science National Institute of Mental Health 6001 Executive Boulevard, Room 7186, MSC 9641 Telephone: (301) 443-5288 All page limitations described in the SF424 Application Page Limits must be followed. Required and Optional Components The forms package associated with this FOA includes all applicable components, required and optional.

Please note that some components marked optional in the application package are required for submission of applications for this FOA. Follow all instructions in the SF424 (R&R) Application Guide to ensure you complete all appropriate optional components.

Instructions for Application Submission The following section supplements the instructions found in the SF424 (R&R) Application Guide and should be used for preparing an All instructions in the SF424 (R&R) Application Guide SF424(R&R) Project/Performance Site Locations All instructions in the SF424 (R&R) Application Guide SF424(R&R) Other Project Information All instructions in the SF424 (R&R) Application Guide SF424(R&R) Senior/Key Person Profile All instructions in the SF424 (R&R) Application Guide All instructions in the SF424 (R&R) Application Guide PHS 398 Cover Page Supplement All instructions in the SF424 (R&R) Application Guide All instructions in the SF424 (R&R) Application Guide must be followed, with the following additional instructions: Strategy: All applications should include the following: (1) Investigators should specifically define the current state of technology as a benchmark against which their proposed technology will be measured.

Describe how the project addresses barriers to single cell analysis in situ that are due to the lack of tools to identify and target cells rigorously, how it provides a major change in practice from current methods for single cell analysis, and to what extent the resulting tool(s) provide first-in-class, cross-cutting or significantly advanced capacity for single cell analysis of heterogeneity in situ (e.g., explant, whole organism).

Describe how the project will transform the field by generating foundational resources (e.g., tools, methods) that will be widely used throughout the basic and translational research community (2) At least one proof-of-concept test, involving analysis of individual cells among a heterogeneous population using an in situ (e.g., explant, whole organism) biological system, should be proposed during the project period.

Applicants are expected to explain the significance of the specific technical approach with respect to characterizing cell heterogeneity in situ. However, the specific biological assay(s) to be utilized in the application may be chosen primarily for utility (rather than biological novelty) in proof-of-concept testing of the exceptionally innovative technique.

The design should indicate how tools be made sufficiently sensitive, selective or appropriate for the intended use in single cell analysis in situ, and how the methodologies lead to a more rigorous distinction between potentially subtle cell subtypes, developmental stages and/or functional states (i.e., cell heterogeneity).

If using multiplex analysis, the design should indicate how multiple or cross-cutting approaches will be utilized to characterize heterogeneous cellular states in a more integrative manner.

If using computational approaches, the design should indicate how these will yield a more comprehensive systems-level view of cell heterogeneity by modeling or (3) Preliminary/feasibility data are not required or expected for early-stage, high-risk projects. However, a sound rationale should be provided as to why the approach proposed is the most appropriate and likely to generate an exceptionally high impact if successful.

In these cases, more emphasis should also be placed in details of the approach, particularly feasibility-testing. In all cases, there should be a particular focus throughout the application on risk management, including alternative strategies if the original plan fails to reach expected performance.

(4) Milestones: In lieu of a timeline, the Research Strategy should include a distinct final section, entitled Milestones , that briefly proposes indicators of progress at critical junctures. These should be tailored to the unique scope of each project and written concretely enough to evaluate what exactly will have been achieved (e.g., crucial steps in tool making) during the course of the project.

Given that projects are likely to be early stage and high-risk in nature, this should include the specific proof-of-concept test(s) that will indicate whether/how a proposed tool actually works , along with alternative strategies should that effort fail to perform as expected.

Tests should include a comparison against existing benchmark technologies; if a tool is truly first-in-class, comparisons may be done against a nearest neighbor technology. Investigators should briefly note how results will be used to inform future phases of tool development, validation or implementation beyond this R21 project. This section should be included within the page limit of the Research Strategy.

If funded, investigators should describe progress toward milestones in the final report of the grant.

Sharing Plan : Individuals are required to comply with the instructions for the Resource Sharing Plans (Data Sharing Plan, Sharing Model Organisms, and Genome Wide Association Studies (GWAS)) as provided in the SF424 (R&R) Application Guide, with the following modification: While it is understood that many tools will be at an early proof-of-concept stage, a central goal of this FOA is to generate transformative tools that will be widely used throughout the research community.

Applications that propose to generate such tools are expected to include a detailed plan for sharing these resources and should include the following key elements consistent with achieving the goals of this program: Project management of resource sharing; Description of what specific resources will be shared (e.g., model organisms, reagents, completed tools or repurposable components thereof); Schedule/timeline for availability of resources to other users; Persons who will have access to the resources (written as broadly as possible to the extent consistent with applicable laws, regulations, rules, Plan for post award disposition of resources.

Do not use the Appendix to circumvent page limits.

Follow all instructions for the Appendix as described in the SF424 (R&R) Application Planned Enrollment Report When conducting clinical research, follow all instructions for completing Planned Enrollment Reports as described in the SF424 (R&R) PHS 398 Cumulative Inclusion Enrollment Report When conducting clinical research, follow all instructions for completing Cumulative Inclusion Enrollment Report as described in the SF424 (R&R) Application Guide.

Foreign (non-U.S.) institutions must follow policies described in the NIH Grants Policy Statement , and procedures for foreign institutions described throughout the SF424 (R&R) Application Guide. Part I. Overview Information contains information about Key Dates.

Applicants are encouraged to submit applications before the due date to ensure they have time to make any application corrections that might be necessary for successful submission. Organizations must submit applications to Grants. gov (the online portal to find and apply for grants across all Federal agencies).

Applicants must then complete the submission process by tracking the status of the application in the eRA Commons , NIH’s electronic system for grants administration. NIH and Grants. gov systems check the application against many of the application instructions upon submission.

Errors must be corrected and a changed/corrected application must be submitted to Grants. gov on or before the application due date. If a Changed/Corrected application is submitted after the deadline, the application will be considered late.

are responsible for viewing their application before the due date in the eRA Commons to ensure accurate and successful submission. Information on the submission process and a definition of on-time submission are provided in the SF424 (R&R) Application Guide. 4.

Intergovernmental Review This initiative is not subject to intergovernmental All NIH awards are subject to the terms and conditions, cost principles, and other considerations described in the NIH Grants Pre-award costs are allowable only as described in the NIH Grants Requirements and Information Applications must be submitted electronically following the instructions described in the SF424 (R&R) Application Guide.

Paper applications will not be accepted. Applicants must complete all required registrations before the application due date. Section III.

Eligibility Information contains information about registration. For assistance with your electronic application or for more information on the electronic submission All PD(s)/PI(s) must include their eRA Commons ID in the Credential field of the Senior/Key Person Profile Component of the SF424(R&R) Application Package .

Failure to register in the Commons and to include a valid PD/PI Commons ID in the credential field will prevent the successful submission of an electronic application to NIH. See Section III of this FOA for information on registration requirements.

The applicant organization must ensure that the DUNS number it provides on the application is the same number used in the organization’s profile in the eRA Commons and for the System for Award Management. Additional information may be found in the SF424 (R&R) Application Guide. tips for avoiding common errors.

Upon receipt, applications will be evaluated for completeness by the Center for Scientific Review and responsiveness by components of participating organizations , NIH. Applications that are incomplete and/or nonresponsive will not be reviewed.

Applications considered nonresponsive to this FOA include: generating tools that are duplicative or an incremental improvement of existing technologies; applications to validate and translate technologies already demonstrated in practice; addressing a biological question as a main focus rather than as a proof-of-concept test of a new technology; studying single cell organisms (e.g., bacteria, yeast).

While the main goal of this FOA is in situ application of technologies, in vitro approaches may be appropriate as an initial step in Involving the NIH Intramural Research Program The requests by NIH intramural scientists will be limited to the incremental costs required for participation.

As such, these requests will not include any salary and related fringe benefits for career, career conditional or other Federal employees (civilian or uniformed service) with permanent appointments under existing position ceilings or any costs related to administrative or facilities support (equivalent to Facilities and Administrative or F&A costs).

These costs may include salary for staff to be specifically hired under a temporary appointment for the project, consultant costs, equipment, supplies, travel, and other items typically listed under Other Expenses. Applicants should indicate the number of person-months devoted to the project, even if no funds are requested for salary and fringe benefits. If selected, appropriate funding will be provided by the NIH Intramural Program.

NIH intramural scientists will participate in this program as PD/PIs in accord with the Terms and Conditions provided in this FOA. Intellectual property will be managed in accord with established policy of the NIH in compliance with Executive Order 10096, as amended, 45 CFR Part 7; patent rights for inventions developed in NIH facilities are NIH property unless NIH waives its rights.

Should an extramural application include the collaboration with an intramural scientist, no funds for the support of the intramural scientist may be requested in the application. The intramural scientist may submit a separate request for intramural funding as described above. Post Submission Materials Applicants are required to follow the instructions for post-submission materials, as described in NOT-OD-13-030 .

Section V. Application Review Information Only the review criteria described below will be considered in the review process.

As part of the NIH mission , all applications submitted to the NIH in support of biomedical and behavioral research are evaluated for scientific and technical merit through the NIH peer For this particular announcement, note the following: The R21 exploratory/developmental grant supports investigation of novel scientific ideas or new model systems, tools, or technologies that have the potential for significant impact on biomedical or biobehavioral research.

An R21 grant application need not have extensive background material or preliminary information. Accordingly, reviewers will focus their evaluation on the conceptual framework, the level of innovation, and the potential to significantly advance our knowledge or understanding.

Appropriate justification for the proposed work can be provided through literature citations, data from other sources, or, when available, from investigator-generated data. Preliminary data are not required for R21 applications; however, they may be included if available.

NIH understands and accepts that early stage, high-risk/high-impact applications that are responsive to this solicitation will be much riskier than for most other R21 solicitations. Any scarcity in preliminary/feasibility data should be balanced by greater emphasis on rationale and approach. Reviewers should balance the risk with the likelihood of the new tools having an exceptionally high impact on the field, if successful.

Applicants are expected to explain the significance of the specific technical approach with respect to characterizing cell heterogeneity in situ. However, research designs should focus on innovative tool development for in situ analysis of single cell heterogeneity, rather than innovative biological concepts about cellular states or heterogeneity.

Any proposed biological assay(s) should be chosen strictly for utility (rather than biological novelty) in proof-of-concept testing of the innovative technique.

Reviewers will provide an overall impact score to reflect their assessment of the likelihood for the project to exert a sustained, powerful influence on the research field(s) involved, in consideration of the following review criteria and additional review criteria (as applicable for the Reviewers will consider each of the review criteria below in the determination of scientific merit, and give a separate score for each.

An application does not need to be strong in all categories to be judged likely to have major scientific impact. For example, a project that by its nature is not innovative may be essential to advance a field. Does the project address an important problem or a critical barrier to progress in the field?

If the aims of the project are achieved, how will scientific knowledge, technical capability, and/or clinical practice be improved? How will successful completion of the aims change the concepts, methods, technologies, treatments, services, or preventative interventions that drive this field? How will the project address barriers to single cell analysis in situ (e.g., explant, whole organism) that are due to