Using Fluorescence Stereo Microscopy to Sort Drosophila Larvae


Fluorescent proteins are powerful tools for diverse applications in Life Science research.  In some cases they are used to visualize organs or other targeted features within organisms, while in others they are used as a convenient visual tool for selecting offspring with desired genetic traits.

One of the very first customers for NIGHTSEA’s economical Model SFA Stereo Microscope Fluorescence Adapter faced a daunting selection challenge. The researcher had funding for an experiment that required efficient sorting of thousands of GFP-transgenic Drosophila larvae.

Everything was lined up, including a team of undergraduates to work at microscope stations to do the sorting, with one exception - the budget was not large enough to acquire multiple fluorescence stereo microscopes. She found the NIGHTSEA system when it was first introduced and was able to outfit four existing stereo microscopes to get the job done.

The Problem

The objective of a research program headed by Dr. Laura Reed (Dept. of Biological Sciences, University of Alabama, Tuscaloosa) was to examine if mutations in specific genes in fruit flies (Drosophila melanogaster) affected triglyceride storage. To collect enough material for the study, she needed 200 or more larvae of each of 84 different genotypes.

Since not all offspring will have the mutations, a special strain of fruit flies was genetically engineered to express an actin promoting green fluorescent protein (GFP) (Figure 1). Only the flies without the mutations fluoresced. The difference between fluorescent and non-fluorescent larvae made them easy to sort.

Figure 1. Non-mutant Drosophila melanogaster expressing GFP (Image courtesy of C. Mazel)

To get the best results, the larvae had to be gathered, sorted and frozen when they were at their largest, but before they pupated. This stage lasts for only about six hours. Dr. Reed had a large group of undergraduates to carry out the sorting, but the major remaining challenge was that she only had access to borrowed time on another laboratory’s research-level fluorescence stereo microscope.

The Practical Solution

Dr. Reed saw NIGHTSEA’s Stereo Microscope Fluorescence Adapter (SFA) system (Figure 2) during her visit to the NIGHTSEA booth at the annual Drosophila Research conference, the meeting at which the system was first introduced.

She immediately realized the potential of putting her undergraduates to work with four existing lab-grade stereo microscopes, each outfitted with an SFA system with Royal Blue excitation for use with GFP (Figure 2). With existing microscopes and a limited budget, the SFA provided a practical and economical solution (Figure 3).

Figure 2. The NIGHTSEA SFA.


Figure 3. (L) Katie Bray and (R) Dana Davis sort larvae using NIGHTSEA’s SFA.

SFA Advantages

NIGHTSEA’s Stereo Microscope Fluorescence Adapters are advantageous in many ways. First, they do not need any modifications to the existing microscope. They just click into place, which makes them simple to use and easy to exchange, either on one microscope or between various microscopes present in the lab.

The SFAs are expandable and economical. Dr. Reed needed to buy only one version of the SFA as she worked only with GFP (green fluorescence/ blue excitation). However, she could readily add additional wavelength combinations as lab requirements evolved.

Lastly, SFA’s exceptional barrier filters and bright illumination often enables conducting fluorescence work under near-ambient lighting as demonstrated by Nick Izor (Figure 4). For this application the overhead lights were turned off and the blinds remained closed, but a special darkened room, as is commonly used for fluorescence microscopy work, was not needed .

Figure 4. Nick Izor demonstrates larval sorting under ambient lighting.

Thanks to NIGHTSEA’s SFA, Dr. Reed had shifts of two to four undergraduates at a time sorting Drosophila larvae. 84 genotypes? 200 larvae per experiment? Problem solved!

For more information on this source, please visit NIGHTSEA.


NIGHTSEA develops economical solutions for viewing fluorescence at scales ranging from stereo microscopy to whole organisms. The product range includes a simple system for adding fluorescence to existing stereo microscopes; fluorescence-exciting flashlights and filter glasses; photography accessories, and more. With both off-the-shelf and customized equipment we help thousands of customers around the world in a wide range of applications in research, education, industry, forensic sciences, exploration and more.

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Last updated: Feb 27, 2018 at 8:42 AM

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