In histology, biology, and pathology, the importance of RNA fixation cannot be overlooked. Fixation mainly helps preserve the samples effectively to ensure its usefulness for the studies that are to be conducted in the future. There are various types of fixation methods used in different laboratories. Some of the popularly used fixation methods are listed below.
Guide to Physical Methods
To ensure optimum RNA protect, physical fixation methods can be used. Physical fixation includes heating the sample at a high temperature or freezing it quickly. When the chemical fixation techniques can affect the structure of the samples, physical fixation method is used. The commonly used physical fixation methods are:
1. Fixation by Heating
In histology, fixation by heating is not generally used as it has the potential to cause damage to the tissues. Heat can result in the dissolving of lipids as well as the coagulation of proteins. However, when it comes to the fixation of the microorganisms in the laboratory, fixation by heating proved to be an ideal fixative method.
It helps in preserving their morphology and ensures effective identification. With the recent advancements and developments, the technique of fixation by heating is also being used in combination with chemical fixation. It helps in reducing the time taken for fixation and enhances the speed of fixation.
2. Quick Freezing
Quick freezing is another popular physical technique that is widely used for RNA fixation. Without the use of any external chemical compound, quick freezing is considered as an ideal technique for maintaining the different molecular features of the required samples.
However, when using this physical technique for fixation, it is important to ensure that the freezing is done fast. Faster freezing will help prevent the formation of ice crystals that might affect the structure of the molecule. Quick freezing will also avoid the autolysis of the RNA molecules.
If possible, pre-treatment of the samples can help in preventing possible damages to the molecules. The quick-freezing technique enables avoiding the chemical reactions in the aqueous environments and allows the long-lasting as well as effective preservation of the molecules.
Guide to Chemical Methods
In the chemical methods of RNA fixation, aqueous solutions are used. The aqueous solutions that contain the fixative substances help in creating a bridge between the molecules. This leads to the immobilization of the molecular compounds and optimally prevents the degradation of the samples.
However, the use of chemical fixation has a significant impact on the physical as well as chemical characteristics of the RNA. Some of the most common physical impacts include hardening, retraction, and distension. In the laboratories, the mainly used chemical fixation methods include perfusion and immersion.
In the immersion technique, the sample molecules are immersed in a specific fixative solution. This results in the diffusion of the fixative into the sample. When using the immersion technique for fixation, there are several factors that need to be considered for better outcomes.
The size of the sample needs to small and less thick in order to ensure effective penetration of the fixative solution. The speed of penetration usually depends on the type of fixation. The other important features of the sample also need to be taken into consideration.
During immersion, the volume of the fixative solution must be higher than that of the taken sample. The pH of the fixative solution must be quite close or similar to the physiological Ph of the RNA sample. The osmolarity of the fixative solution, as well as the sample, also needs to be quite similar in order to avail of enhanced results.
Perfusion is another widely used fixation technique for RNA protect. The method of fixation through perfusion is considered to be more effective in comparison to the immersion technique. However, perfusion can be performed for plants as well as many biopsies. In the case of perfusion, the fixative solution comes in contact with the sample molecules much faster than the immersion technique.
On the basis of the requirement, any of these fixation techniques may be used by the laboratories. When using fixative solutions in case of chemical method, an ideal fixative solution must be used. Some of the characteristics of an ideal fixative solution are:
1. It should be able to penetrate the sample much faster.
2. It should be non-allergic as well as non-toxic in nature.
3. It should be capable of preserving the volume of the sample.
5. It must prevent the short term as well as long term destruction of the sample.
6. It must be cost-effective.
The effective fixation of RNA makes it more useful for carrying out different types of research studies in the future. Choosing the right fixation method and an appropriate fixative solution is essential to ensure optimum outcomes.