# TRD #3: An automated and streamlined pipeline for in-situ molecular microscopy

> **NIH NIH P41** · NEW YORK STRUCTURAL BIOLOGY CENTER · 2021 · $297,728

## Abstract

4. TRD#3: An automated and streamlined pipeline for in-situ molecular microscopy
Abstract
Our goal for this TRD is to integrate all of the techniques required for in situ molecular microscopy into a robust
and reliable pipeline, and make it available to the scientific community. In situ molecular microscopy, that is
reconstructing molecules inside cells or tissue, requires the integration of several challenging methods and
techniques. Regions of interest need to be identified using cryo light micrsocopy (cryoLM), the specimen
needs to be thinned down around this region of interest using a focused ion beam scanning electron
microscope (FIB-SEM) in order to make it amenable to subsequent cryo electron tomography (cryoET). We
will develop a variety of approaches for specimen preparation including some that are high-risk but of
potentially high-impact. We will expand the Appion pipeline to integrate and track images, analysis and
metadata across all three imaging modalities. Providing streamlined and transparent access to this pipeline is
an essential aspect of making these methods accessible to general users. We will incorporate the ability to
integrate light microscopy and SEM images into our image tracking database and enable relevant image
processing techniques using our web based pipeline; the database that underlies both Leginon and Appion will
be key to tracking these multiple imaging modalities and related actions and metadata. Several important
biological projects will drive improvements in the design, development, automation and streamlining of this
pipeline, and we anticipate a range of Collaborative and Service projects will make excellent use of the pipeline
as soon as it is available. We will work closely with several groups with deep experience with in situ
microscopy (Elizabeth Villa, Carolyn Larabell, and Michael Marko) to guide this project in the right direction.
We will hold an annual weeklong workshop to train users in these methods and we will help to disseminate the
overall pipeline to other labs in the scientific community, just as we do now for our Leginon and Appion
systems.

## Key facts

- **NIH application ID:** 10136616
- **Project number:** 5P41GM103310-21
- **Recipient organization:** NEW YORK STRUCTURAL BIOLOGY CENTER
- **Principal Investigator:** Jeffrey S Kieft
- **Activity code:** P41 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $297,728
- **Award type:** 5
- **Project period:** 2002-09-30 → 2023-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10136616

## Citation

> US National Institutes of Health, RePORTER application 10136616, TRD #3: An automated and streamlined pipeline for in-situ molecular microscopy (5P41GM103310-21). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10136616. Licensed CC0.

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