What is Zamboni fixative?

Zamboni’s Fixative is a phosphate-buffered picric acid-formaldehyde (PAF) solution that penetrates rapidly, stabilizes cellular proteins, provides optimal tissue preservation, and is stable at room temperature. Zamboni’s Fixative can also be used as a general purpose fixative for light microscopy.

How to make plp fixative?

Periodate-Lysine-Paraformaldehyde (PLP)

1. A. Lysine HCl solution. Lysine HCl, 13.7 g.
2. B. Paraformaldehyde solution.
3. C. Final PLP solution.
4. Paraformaldehyde (from 16% stock solution), 125 ml. Picric acid (saturated aqueous), 150 ml.
5. Picric acid (saturated aqueous), 75 ml. Formalin (37-40% formaldehyde), 25 ml.

What is PLP fixative?

Periodate-lysine-paraformaldehyde (PLP) has been proposed as a fixative for glycoprotein antigens which should stabilize periodate oxidized polysaccharide chains through lysine mediated crosslinks, either directly or by the intermediation of formaldehyde.

What is the function of the fixation process in immunohistochemistry?

All samples used in IHC/ICC experiments must be fixed to preserve tissue morphology and retain the antigenicity of the target molecules. Fixation alters the chemical composition of tissues and often requires a compromise between preserving tissue structure and preserving the epitope.

What is the fixative solution for immunofluorescence in PBS?

Fixative 4% formaldehyde in PBS (freshly prepared) 3. Quenching solution: 50 mM NH4Cl in PBS or 0.1M Glycine in PBS 4. Blocking solution 1% BSA or 10% FCS (fetal calf serum) in PBS 5.

What is the protocol for immunofluorescence in vivo?

1. Prepare tissue or culture cells 2. Prepare tissue section or cells coverlip 3. Wash samples two times with PBS 4. Fix amples with 4% paraformaldehye in PBS for 15 min at room temperature (Note: Paraformaldehye is toxic, use only in fume hood)

How to prepare cells for immunofluorescence fixation?

Many cells will grow hapily on uncoated glass but some do better with coverslips coated (eg 50 µg/ml poly-lysine for 1 hr, or collagen). Transfer your cells into the plates and culture them (eg overnight) so they are well adhered and so they are 40-70% confluent when they are ready for fixation.

How to make immunofluorescence with sterile cover slips?

Place the sterile cover slips in 12 or 24 well plates; rinse the cover slips with PBS, followed by a quick rinse with culture media. Plate the cells on the cover slips at a density of ~ 10,000/ cm2 • Day 2 1. Rinse the cells with cPBS 2. Fix the cells with freshly made fixative, for 30 min 3. Wash gently with PBS, for 2 min, twice 4.