Dec 11, 2023

Public workspacePreparation of fresh paraformaldehyde for mouse perfusion and tissue fixation

DOI
dx.doi.org/10.17504/protocols.io.6qpvr3qbbvmk/v1
  • 1UCSD
amanda schneeweis
Open access
DOI: dx.doi.org/10.17504/protocols.io.6qpvr3qbbvmk/v1
Protocol Citation: Thomas Hnasko 2023. Preparation of fresh paraformaldehyde for mouse perfusion and tissue fixation. protocols.io https://dx.doi.org/10.17504/protocols.io.6qpvr3qbbvmk/v1
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: Working
We use this protocol and it's working
Created: December 11, 2023
Last Modified: December 11, 2023
Protocol Integer ID: 92134
Abstract
Hnasko lab - preparation of fresh PFA for ms perfusion and tissue fixation
Materials
paraformaldehyde (PFA, EMS, cat# 19210)
10X PBS (Fisher bioreagents, cat# BP399-1)
vacuum filtration flask (ThermoScientific, cat# 567-0020).
Safety warnings
Attention
Caution: Paraformaldehyde is toxic. Please read the MSDS before working with this chemical. Gloves, mask and safety glasses should be worn and solutions made inside a fume hood.
1. For Amount1 L of 4% paraformaldehyde, warm up Amount700 mL of ddH2O in a glass beaker using microwave until Temperature40-60 °C .

2. Place on heated stir plate in a chemical fume hood, add a magnetic stir bar and start stirring and insert thermometer into solution.
3. Weigh Amount40 g of granular paraformaldehyde (PFA, EMS, cat# 19210).

4. Add PFA to the heated ddH2O solution and place on a stir plate in a chemical fume hood. Insert thermometer. Stir and heat to approximately Temperature60 °C (paraformaldehyde flash point = 71°C). Avoid overheating.

5. The powder will not immediately dissolve into solution. Slowly raise the pH by adding about Amount1 mL NaOH 10N until the solution clears.

6. Once the paraformaldehyde is dissolved, add Amount100 mL 10X PBS (Fisher bioreagents, cat# BP399-1).

7. Cool solution to TemperatureRoom temperature before adjusting pH, can place on ice to speed cooling.

8. Adjust pH to ~Ph7.4 with small amounts of 12N HCl.

9. Adjust the volume of the solution to Amount1 L with ddH2O.

10. Filter the solution using a vacuum filtration flask (ThermoScientific, cat# 567-0020). Label and store in sealed container at Temperature4 °C .