Dec 10, 2022
Eggs injections
This protocol is a draft, published without a DOI.
- FishFloorUCL1
- 1University College London
Protocol Citation: FishFloorUCL 2022. Eggs injections. protocols.io https://protocols.io/view/eggs-injections-ckbgusjw
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: In development
Created: December 09, 2022
Last Modified: March 22, 2023
Protocol Integer ID: 73800
Abstract
Protocol for microinjections of 0-dpf eggs @FishFloorUCL.
Set-up breeding pairs
Set-up breeding pairs
Set up the breeding pairs the night before or order them from the fish facility. Arrive early in the morning, 9AM for single cell, 9:30 for fish facility injections.
Make sure to book the injector in advance.
Calibrate the needle
Calibrate the needle
Open the red valve for your injector, you should see the dial turn to ~40
Power on switch
Set the time to 14-15ms
Take a needle from the large Petri dish on the side and stick in the blue tack above the microscope
Add what you want to inject (less is better, eg: 1-3 nL)
Place the black measuring slide under the microscope, add a drop or two of mineral oil and focus on the little ruler.
When tilting the whole needle base use both hands. When not using make sure it is tilted up
Tilt the needle into the oil
Break the needle with forceps and press the pedal until the correct amount is coming out of your needle (using the table on the wall to calibrate)
Collect eggs
Collect eggs
Take the dividers out of the fish boxes
wait 6-7 minutes
Transfer the fish to new boxes and replace the dividers for another round of eggs
Collect the eggs by pouring the water through a small sieve and transferring this to a Petri dish, clearly labelled
Inject the eggs
Inject the eggs
Use a Pasteur pipette to transfer eggs to a new pipette
Use a thin ended pipette to remove all the water and then line them up along the edge of a glass slide placed in the Petri dish
Go down the line of cells and inject one at a time
Collect a new clutch and repeat.
Turn everything off/clean after yourself
Turn everything off/clean after yourself
Turn off the red tap.
Turn time to minutes, press pedal until the dial has run completely down, flick it back to ms
Turn off the power switch
Wipe down everything with ethanol.