The Fixing Agent was first applied in optical microscopy, but the Fixing Agent used in the optical microscope was not particularly suitable for electron microscopy. The people were then introduced to the immobilization of osmium and neutral formaldehyde. The early electron microscopic biological sample chemical fixation method uses four oxidation osmium single fixed, but the osmium does not have the very good preservation glycogen and other carbohydrate, moreover the four oxidation osmium in the tissue's infiltration speed is relatively slow.
All samples used in the IHC/ICC experiment must be fixed to maintain tissue morphology and preserve antigenic characteristics of the target molecule. Fixed changes in the chemical composition of the Organization, so it is often necessary to maintain the organizational structure and retain the table between the trade-offs.
Incomplete immobilization of cells or tissues (inadequate immobilization) may result in rapid hydrolysis of the target protein in the tissue and reduce specific immune reactivity. However, excessive fixation (fixed excessive) may result in shadowing of the table or strong non-specific background staining, which may obscure the specific markings.
It is best to preserve the morphological structure of cells and tissues, and to maximize the immune activity of antigens. However, so far no standard stationary fluid can be used for various antigen immobilization. After fixation with the same stationary fluid, different tissues may exhibit distinct staining results. Therefore, different antigens and samples must be tested repeatedly to select the best retention agent.
Unlike tissue samples, the cell has a shorter fixed time and a lower concentration of immobilized fluid. For example, a 2% formaldehyde solution at room temperature for 20 minutes, often sufficient to preserve cell morphology and antigenicity.
The fixation of cultured cells is usually only removed from the medium and added to the stationary solution. However, changes in surface tension after removal of the medium may impair certain cell types. If this is the case, the fixing agent can be directly added to the medium. For example: adding 4% formaldehyde in the same volume as the medium will get 2% formaldehyde solution, which is sufficient to preload the cell. After 2 minutes, the pre-fixed medium should be replaced with a fresh amount of 2% fixed agents. The fixed steps make the cells harder, so they can withstand any possible harmful effects of surface tension changes.