Over two years ago, I started a new postdoc using a new model species, zebrafish. Luckily, I was able to collaborate with an established PI with a large, well-maintained zebrafish facility.
I love working with zebrafish- they look beautiful under the microscope and are a great way to expose undergraduates to research. But I’ve learned a few lessons the hard way and wanted to hear what tips and tricks other zebrafish researchers had to share. So of course, I headed to Twitter.
Here are their responses, including a few tips of my own.
1. (Your) Hydration is key
Well #zebrafish community? What tips can you share?
For one, I find new #zfish researchers are not aware how warm the facility is kept. Hydration is key.
If it’s their first time working with fish I also emphasize patience especially with learning how to net.
— ZHA (@ZHAonline) November 21, 2017
Zebrafish researchers dress very casual, even for scientists. A warm aquatics facility may be great for the zebrafish, but it can be hard on us humans. T-shirts and shorts (if permitted) are common, and I quickly got too warm in my favorite lab coat. Plus, washing fish tanks with bleach is bound to put holes in your clothes.
@ZHAonline also recommends patience when learning how to net. I admit, some days I think the zebrafish are re-enacting a scene from Finding Nemo. “Swim down! Swim down! Swim down!”
2. Mate the most “female” looking females with the most “male” looking males.
The more a fish looks like a female, the better chance it has of laying eggs (bigger belly= more eggs). I like this mating guide.
If you are crossing two different zebrafish strains, this is especially important as sex is typically used to keep track of the source of each fish in a mating pair.
3. Just let them rest
Look at Mueller et al for ideas of equivalent brain regions, clean eggs on a dark matt and if your fish don’t lay eggs don’t panic, just let them rest for a week or two.
— Sophie Leacock (@Sophie_Leacock) November 21, 2017
You may have been told to only mate fish once a week, but you should also remember that repeatedly mating the same fish is metabolically costly. Make sure to feed your fish a little extra if you plan to mate them frequently. It will also make determining the sex of the fish next week a lot easier when they are nice and chubby!
4. Be careful in-crossing transgenic fluorescent fish
While in-crossing transgenic fluorescent zebrafish is a sure-fire way to get lots of fluorescent embryos, you may discover unexpected phenotypes. If you are not certain whether the transgene in your fluorescent zebrafish impairs the function of any genes, be cautious when generating homozygous transgenic fish. You may unexpectedly reveal a deleterious phenotype normally unobserved in heterozygotes.
5. Mighty morpholinos
For new zebrafish researchers: if you will use Morpholinos, read “Guidelines for morpholino use in zebrafish”. https://t.co/XY2rTYFd2D
— Jon D. Moulton (@jonmoulton) November 21, 2017
With the advent of zinc finger nucleases, TALENs, and CRISPRs, discrepancies between mutants and morpholinos have become apparent. The above article briefly describes the importance of comparing morpholinos with mutants and provides updated guidelines for the use of morpholinos.
6. Sometimes fluorescent protein doesn’t fix.
Never treat GFP transgenic fish with MeOH, trichloroacetic acid, or acetone if you want to retain the fluorescence. Paraformaldehyde is best, and be careful when using various antigen retrieval methods.
I sometimes have trouble observing fluorescence even after fixation in paraformaldehyde, especially with red fluorescent protein. ALWAYS check if your fluorescent protein fixes nicely before starting a big experiment.
7. Never trust a new transgenic.
You can never assume that a transgenic fluorescent fish has the expression listed on its tank before using it for the first time. On several occasions, I have found fish that do not express their transgene at my time point of interest, or do not express their fluorescent protein strongly enough for proper observation. It is best to check your fish by live imaging or immunohistochemistry when using a new transgenic.
8. It’s a team effort
Dedicated staff, consistency of care, proactive maintenance and LOTS of room to set up those crossing tanks! Make sure your photoperiod is working properly #personalexperience
— Aleah McCorry (@imjustaleah) November 21, 2017
Zebrafish research runs so much more efficiently when approached from a team perspective. Scheduling feedings over the holidays, monitoring general health of the zebrafish population, time-sensitive mating periods, and identifying stage-specific phenotypes seem unimaginable to me without a team of dedicated students and staff.
8. Look at a larval zebrafish under the microscope.
Just look at it. It is SO cool! Sometimes I take a break from sorting fish just to look at these beautiful creatures. Wonder at the blood cells traveling all around the body in front of your eyes. Check out how the notochord looks like a stack of pennies. If you are using fluorescent fish, look at the fluorescence in a different tissue than you normally focus. You may find a new perspective helpful to your research. At the least, it will remind you of the value of zebrafish in biomedical research and the respect that these animals deserve.
9. Get ready for arts and crafts.
Here are some fun things I have seen used or created while working with zebrafish:
- hand-pulled glass probes for manipulating embryos
- vacuum grease creates a seal of variable heights between your slide and coverslip
- 3d-printed food dispensors
- custom-made embryo troughs
- tiny beveled needles for cell transplants
- Scotch-Bright(TM) sponges for holding zebrafish
I think zebrafish lend themselves to creativity since they are harder to contaminate than cells and are big enough to see and handle. I can’t wait to set up my own aquatics facility!
10. Join a zebrafish community.
The Zebrafish Husbandry Association @ZHAonline was quick to offer advise for the creation of this post, and has a number of helpful resources available to new zebrafish scientists.
Our annual meeting is a great way to network, learn new ideas, & discuss common concerns in #zebrafish facilities worldwide. We also host #zfish webinars & connect members around the world. If you would like to discuss more feel free to DM or email email@example.com
— ZHA (@ZHAonline) November 21, 2017
Other helpful communities include
- Society for Developmental Biology
- International Zebrafish Society
- ZFIN, The Zebrafish Information Network
Do you have any other tips that you wish you knew when starting with zebrafish? Share them in the comments below!