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In cell culture, a typical formaldehyde fixing procedure would involve using a 4% formaldehyde solution in phosphate buffered saline (PBS) on ice for 10 minutes. In histology and pathology specimens preparation, usually, the fixation step is performed using 10% Neutral Buffered Formalin (4% formaldehyde) for, at least, 24 hours.
The most commonly used fixative in histology is formaldehyde. It is usually used as a 10% neutral buffered formalin (NBF), that is approx. 3.7%–4.0% formaldehyde in phosphate buffer, pH 7. Since formaldehyde is a gas at room temperature, formalin – formaldehyde gas dissolved in water (~37% w/v) – is used when making the former fixative.
It was invented by French biologist Pol Bouin and is composed of picric acid, acetic acid and formaldehyde in an aqueous solution. [2] Bouin's fluid is especially useful for fixation of gastrointestinal tract biopsies because this fixative allows crisper and better nuclear staining than 10% neutral-buffered formalin.
The most common fixative is 10% neutral buffered formalin (corresponding to 3.7% w/v formaldehyde in neutral buffered water, such as phosphate buffered saline). Preparation for histology [ edit ]
If the PAS stain will be performed on tissue, the recommended fixative is 10% neutral-buffered formalin or Bouin solution. For blood smears, the recommended fixative is methanol. Glutaraldehyde is not recommended because free aldehyde groups may be available to react with the Schiff reagent, which may result in false positive staining. [4]
Formaldehyde (/ f ɔːr ˈ m æ l d ɪ h aɪ d / ⓘ for-MAL-di-hide, US also / f ə r-/ ⓘ fər-) (systematic name methanal) is an organic compound with the chemical formula CH 2 O and structure H−CHO, more precisely H 2 C=O.
If the glacial acetic acid is replaced by 5 ml of formalin (37–40% formaldehyde), the resulting solution is Helly's fixative, also sometimes called "formol-Zenker".Helly is stable for only a few hours because the formaldehyde and dichromate components react, producing formic acid and chromium(III) ions; the orange solution becomes greenish.
Buffer capacity falls to 33% of the maximum value at pH = pK a ± 1, to 10% at pH = pK a ± 1.5 and to 1% at pH = pK a ± 2. For this reason the most useful range is approximately p K a ± 1. When choosing a buffer for use at a specific pH, it should have a p K a value as close as possible to that pH.