# Optimization of vitrification methods for Drosophila embryos

> **NIH NIH R21** · MASSACHUSETTS GENERAL HOSPITAL · 2020 · $224,088

## Abstract

PROJECT SUMMARY
Drosophila research labs routinely maintain hundreds to thousands of unique strains, where each new
generation must be transferred to fresh food every 4 to 6 weeks. Not only is this practice labor and resource
intensive, but there is a risk that valuable strains will be lost or acquire additional mutations. This limitation would
be overcome by the development of Drosophila cryopreservation methods. Thus, the overall goal of this proposal
is to develop a simple, robust method to cryopreserve Drosophila embryos. Based on literature review and our
expertise in cryobiology and Drosophila biology, we anticipate that vitrification is the optimal approach to
cryopreservation. Vitrification is an `ice-free' method of cryopreservation where cells are loaded with high
concentrations of cryoprotective agents (CPAs, e.g. dimethyl sulfoxide, propylene glycol, etc.) and rapidly cooled
through the glass transition. The result is formation of an amorphous glassy state as opposed to crystalline ice.
Nearly thirty years ago, a vitrification protocol for wild-type Drosophila was reported. However, this protocol is
quite complex and results in a loss of nearly 70% of embryos upon development into adults. However, this
breakthrough method represents an excellent starting point for us here, where we will take a comprehensive
approach to both simplify and optimize the vitrification procedure by incorporating concepts and technologies
from industrial chemistry, engineering and cryobiology. In Aim 1, we will simplify the embryo permeabilization
procedure and then automate the liquid handling steps of the protocol by incorporating microfluidics. Automation
will reduce user-error and decrease labor demands. In Aim 2, we will optimize the vitrification procedure by
systematically evaluating a series of CPAs and CPA cocktail solutions. This information will then be used to
increase the intraembryonic concentration of CPAs, thereby reducing the risk of ice crystallization upon cooling
or rewarming. We will finally merge results from both Aims to validate this simplified and optimized vitrification
procedure, where our goal is to achieve ≥60% survival of thawed embryos to adults. To better ensure the
successful completion the work described here, we have assembled a multidisciplinary team including
cryobiologists, engineers and Drosophila biology experts. The successful completion of this exploratory proposal
will result in the development of a simplified and robust vitrification method for Drosophila embryos that can be
quickly implemented into laboratories and resource centers.

## Key facts

- **NIH application ID:** 9957490
- **Project number:** 1R21GM137186-01
- **Recipient organization:** MASSACHUSETTS GENERAL HOSPITAL
- **Principal Investigator:** Rebecca Sandlin
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $224,088
- **Award type:** 1
- **Project period:** 2020-06-01 → 2022-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9957490

## Citation

> US National Institutes of Health, RePORTER application 9957490, Optimization of vitrification methods for Drosophila embryos (1R21GM137186-01). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9957490. Licensed CC0.

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