After conducting some research, I have come to the following conclusions and procedure regarding the use and preparation of buffered paraformaldehyde (PFA) as a fixative. However, it would be valuable to hear from experts who have hands-on experience with this method and can share their insights and recommendations.

The differences between paraformaldehyde (PFA) and formaldehyde are as follows: PFA is in a white powder state, while formaldehyde is already dissolved in a solution. Additionally, PFA does not contain methanol, which is why it is sometimes referred to as "methanol-free 10% formalin." In contrast, formaldehyde solutions typically include methanol. Methanol is added to saturated formaldehyde solutions to inhibit or slow down polymerization, which reduces the fixing power of formalin.

Due to these differences, PFA can be used to prepare buffered formalin instead of formaldehyde. However, there are certain considerations to keep in mind.

Although the chemistry is essentially the same, 4% paraformaldehyde is polymerized, which means it does not penetrate tissues as rapidly as 10% buffered formalin. Since 4% paraformaldehyde is usually refrigerated to inhibit polymerization, the chemical reactions during fixation occur more slowly at lower temperatures. To achieve the same level of protein cross-linking, you may need to increase the fixation time, such as 10 hours at 24°C or 40 hours at 4°C.

Below is a protocol describing the technique for preparing a 4% formaldehyde solution in Phosphate-Buffered Saline (PBS).

Procedure:

References:

1. chrome-extension://efaidnbmnnnibpcajpcglclefindmkaj/https://content.ilabsolutions.com/wp-content/uploads/2014/09/10pct_Buffered_Formalinvs_4pct_Paraformaldehyde.pdf

2. https://www.rndsystems.com/resources/protocols/protocol-making-4-formaldehyde-solution-pbs

I have reached some conclusions regarding the use of PFA to prepare buffered formalin. However, I have a few additional questions that I would like to ask:

It is important to have a neutral fixing liquid, because some plankton contain lime, which is soluble in an acidic environment. Have experienced that ostracods have given off gas bubbles polyvinyl-lactophenol. An acidic embedding agent.