Views: 0 Author: Site Editor Publish Time: 2023-08-08 Origin: Site
01 Misconception 1: The more magnetic beads , the better the extraction effect.
Many teachers tend to increase the amount of magnetic beads when the extraction effect is not good, thinking that more magnetic beads will be able to absorb more nucleic acids. However, it must be said that this idea is not desirable.
The main feature of magnetic beads is that they can be both dispersed in liquid and separated from the liquid phase in a solid manner under the action of an applied magnetic field, any reagent system, the ratio of magnetic beads and liquid should be a certain threshold, over a certain ratio, too many magnetic beads will not be able to evenly dispersed in the liquid because of the loss of its dispersion properties, but also make the washing process can not be sufficient to increase the efficiency of the nucleic acid beads and the liquid contact.
And excessive magnetic beads will also adsorb more impurities, which has a great influence on the effect of removing impurities. There are even times when too many magnetic beads will adsorb protease, lysozyme and other functional components that play a major role in the liquid system, leading to the inefficiency of the entire kit. There are many times when the extraction effect is poor, reduce the amount of magnetic beads used, but is the optimal way to improve the extraction effect.
Usually, the reference bead dosage given by the magnetic bead method kit is slightly excessive, so there are not many cases where the bead dosage needs to be increased to improve the adsorption efficiency. However, if it is determined that the extraction effect is poor due to insufficient bead dosage, it is possible to improve the extraction effect by increasing the bead dosage within a certain range.
02 Misconception 2: The more reagents used, the better the extraction effect.
The lysis effect is not good? Add more lysis solution. The washing effect is not good? Add more washing solution. This is the habitual thinking of many customers in the use of reagent kits.
However, for the magnetic bead method, every increase in the volume of the liquid reduces the chance of collision of the magnetic beads, and the reduction of the chance of collision of the magnetic beads will lead to a significant decrease in the adsorption rate. So very often, although the increase in lysis solution and washing solution can indeed play a role in enhancing the role of lysis and washing, but the core of the magnetic bead method of extraction is the efficiency of the magnetic beads adsorption of nucleic acids, can not guarantee the efficiency of the magnetic beads collision efficiency can not guarantee the efficiency of nucleic acid extraction, so the simple increase in the use of reagents to improve the extraction effect is not necessarily completely effective.
03 Misconception 3: The more times you wash, the better the extraction effect.
When the extracted nucleic acid has too many impurities, the user will consider washing more times to get more pure nucleic acid. Increasing the number of washes is indeed conducive to the purification of nucleic acids, but taking into account that each wash will lose a certain amount of nucleic acids, and increase the possibility of hydrolysis of nucleic acid fracture, so generally speaking, the number of washes is controlled in the 2 ~ 4 times is appropriate.
04 Misconception 4: The more samples taken, the better the extraction effect
In the case of the sample is not fresh enough or the nucleic acid content itself is very small, often the nucleic acid extraction effect is not good, many teachers will use more samples to increase the amount of nucleic acid extraction.
However, simply increasing the amount of sample taken will sometimes introduce too many impurities beyond the lysate's ability to lyse, and will also reduce the extraction efficiency, so it is not recommended to achieve the goal of increasing the extraction amount by simply increasing the amount of sample taken.
If the extraction volume is indeed too low due to insufficient sample volume, it is recommended to go through an enrichment or concentration step in the pre-treatment before starting the extraction. Or increasing the completeness of the lysis to expose more nucleic acids is also a solution.
05 Misconception 5: If one type of bead is good, it should work well in all experiments
There are various types of magnetic beads, different particle size, different dispersibility, different magnetic response time, different encapsulation base matrix, different outer layer modification functional groups, different encapsulation density, different functional group arm lengths, which will lead to a wide range of magnetic bead characteristics.
Therefore, different magnetic beads are adapted to different experiments and systems. Just like the same reagents for nucleic acid extraction are not exactly the same formula, the same magnetic beads applied to nucleic acid extraction are not exactly the same in nature.
Some magnetic beads show higher adsorption efficiency in macronucleic acid extraction, while others are more suitable for trace nucleic acid extraction. Some magnetic beads are suitable for acidic series of reagent systems, and some magnetic beads are suitable for alkaline series of reagent systems. Some magnetic beads have good magnetic response but fast settling speed, which is more suitable for magnetic rod type automatic extractor; some magnetic beads have slow settling speed but long magnetic response time, which is more suitable for pipette type automatic extractor.
Rarely a magnetic beads can be suitable for all experiments, in addition to the fixed kit with, in most cases, magnetic beads and reagent systems have to do a certain amount of time with the adjustment.
06 Misconception6: Comparison with a certain kit is not good, that is, the magnetic beads are not good.
Many customers in the process of screening magnetic beads in the already mature reagent system, simply equal amount of replacement of magnetic beads, used to compare the effect of magnetic beads.
This will easily conclude that a certain magnetic bead is not effective, but in fact, due to the different magnetic beads suitable for different systems and dosage is different, often need to be adjusted in order to obtain better extraction results.
