Tissue flotation bath is required after the step of cutting paraffin sections and before theseare placed on slides. It lets tissues relax and smoothen before being mounted on slide as well as makes paraffin stick to the slide. This ensures removal of wrinkles and folds before sections are placed on the slide.
Tissue flotation baths provideexact temperature regulation, thus ensuring evenly stretched cuttings that are clearly visible in the anodized bath. It is ideal for use in histological, chemical, clinical, and bacteriological laboratories that require stretching and drying of cut tissues.It also has applications for processes that require less distortion and wrinkle free tissue samples, such as tissue culture and enzyme reactions fermentation tissue section processing.
Water bath is too cold
Sections never properly flatten in such cases and may remain wrinkled. Slightly warmer water could help overcome this problem.
Water bath is too hot
Paraffin melts and tissue morphology changes due to overexpansion, resulting in cell and tissue damage. It can be microscopically seen asshriveled, pyknotic nuclei and extensive cracking.
Placing for extended time
This can lead to overexpansion and cell and tissue damage, especially to delicate samples such as lymphoid tissue. This can be overcome by placing samples on the bath for just enough time to flatten.
Air bubbles in the bath
If ignored, air bubbles sometimes get trapped under the section and may seem to disappear as the section dries; however, these distort the section and usually float off during staining. Therefore, air bubble formation in the flotation bath should be avoided and any visible bubbles should be dislodged before the sections are laid on water.
Out of focus areas
This occurs when slides are not properly drained before horizontal drying. As a result, sections sometimes move on the slide and do not dry flat, resulting in raised, out of focus appearance under the microscope. To overcome this problem, sections should be briefly drained before being placed in the drier.
Inconsistent adhesion
This occurs when protein based section adhesives, such as gelatin, glue, or starch are used in the water bath. It results in accumulation of reagents below the lifting sections leading to blocking of the surface of charged slide causing inconsistent adhesion and uneven staining, microscopically seen as a thick line adjacent to the section.
Tissue flotation baths should be filled with distilled water; 95% alcohol is sometimes added to water as it lowers the surface tension and removes wrinkles. Adding adhesives to water can also sometimes improve slide adhesion.
The temperature of tissue flotation bath should be optimized depending on the sample as well as the type of paraffin used; fatty tissues, such as brain or breast tissues, require lower temperature. The optimum temperature for a tissue flotation bath is 40°C–50°C, which is 5°C–10°C below that of paraffin’s melting point. This temperature is optimum for most paraffins since they are soft and would otherwise disintegrate at higher temperatures; hard paraffins, on the contrary need higher water temperature.
Machine cleanliness and maintenance should be regularly followed. Water should be kept clean and changed daily. Reservoirs should also be cleaned daily and paraffin should be removed from gauze. Empty reservoirs should be covered do avoid contamination from dust and fibers.
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