How to target pathogen samples
Quality nucleic acid extraction
If you want to do something good, you have to do it first, and get a tool to take advantage of it. In the development of nucleic acid detection products, screening of raw materials is one of the key factors influencing the final performance of the product. The first step of the nucleic acid test, however, is the extraction of nucleic acid, which is the highest priority. For the detection of nucleic acid in infectious samples, in order to ensure the sensitivity and accuracy of the results and the availability of high-quality nucleic acid, the following issues need to be considered:
Complexity of samples
The true samples at the time of the pathogen tests are complex and the first samples are of various types, such as blood, saliva, saliva, brain spinal fluid, urine, etc., with very wide variations in their matrices. Second, even for the same type of sample state, such as pulmonary pneumatic pneumatic fluids, can vary according to the patient's individual circumstances, and the differences in treatment, use of medication and progress of the disease result in significant changes in the composition of pneumatic pneumatic pneumatic pneumatic pneumoccanate. Therefore, producers of nucleic acid extraction reagent boxes need to consider compatibility of different sample types。
Diversity of pathogens
Fungi, dna virus, rna virus, gelan positive fungus, glancella cactus, etc. The different pathogen types mean different requirements for break walls and nucleic acid releases. In the same sample taken, both the release of fungi nucleic acid and the integrity of low abundance rna virus nucleic acid are essential。
Depressants composition
The downstream application after nucleic acid extraction is mostly based on the reaction of the bioenzymes, while the optimal activity of the bioenzymes requires stricter requirements for salt ion concentrations, ph and organic solvents in the reaction buffer, so that the nucleic acid extracted requires certain purity. The inhibitor component in the nucleic acid extraction product is derived mainly from two sources, the sample and the reagent itself. Therefore, the nucleic acid extraction reagent box needs to ensure that the non-nucleic acid composition of the sample is removed, while the reagent itself does not contain enzyme inhibitors。
High flux
In order to improve the efficiency of the entire testing process, while reducing human-induced errors and human resource inputs, steps and reagents of nucleic acid extraction should match automated platforms to achieve high-throughput efficient extraction of samples when products are put into practical application。
Appropriate suppliers
Lgc biosearch technologiestm has been serving the diagnostic industry as a reliable and key group provider of oligonucleic acid, enzymes and sample preparation solutions during the new crown pandemic。
In order to better meet client needs and promote the high-quality development of molecular diagnostic products, lgc biosearch technologies released an entirely new magnetophate pathogen nucleic acid extraction reagent (i. E. Sbedextm pathogen nucleic action kit) designed to help clients achieve simultaneous and efficient extraction of different samples, different pathogens, and provide the best nucleic acid extracts for downstream applications。
Sbeadex
Sbedex pathogen
Nucleic acid peacekeeping kit
Data on performance
Sample type validation
The most common types of infections that have been verified are swabs (utm/vtm), saliva, whole blood, plasma, serum, urine, faeces, cerebral spinal fluids. As shown in figure 1 of the results of the experiment, the reagent box completes the extraction of the pathogens mixed in the different samples, while the results indicate that there is some optimization of the length of the cracking time for different samples. The pathogen nucleic acid in the sample may degrade at the breakup of 20 minutes, if the full blood rna virus extracts the best time of three minutes。
Figure 1
Compared with the mainstream reagent extraction box in the market, the four-fold diluted sars-cov-2 standard was integrated into the whole blood sample, with three gradients containing, respectively, 1200 (neat), 200 (1 in 4), 75 (1 in 16) copies. Rt-qpcr validation after extraction. As shown in figure 2, sbedex nucleic acid extraction reagent boxes perform better than similar plant products in samples of different pathogens。
Figure 2. Comparison of results with other plants for different pathogens in samples
Diversity of pathogens
A subsequent test of nucleic acid extraction with dilution into samples of common gland positive bacteria, gland vaginal bacteria and fungi is then performed. The results show a good linear relationship between the different input samples and the final qpcr results, as shown in figure 3. This indicates that the reagent box can be successfully extracted from different types of pathogens at different concentrations。
Figure 3. Extracting tests for different types of pathogens
Automation fit
The use of sbedex pathogens to extract reagent cartridges and oktopure has been successful in automating the rna virus. The optimisation programme for oktopure uses a reduced buffer volume of 50 μl, with a total extraction time of 3 hours and 9 minutes for each of the eight 96-pore plates (including tablet preparation steps). The results of the nps (sniffing) and ss (slurry) samples are detailed in table 1. Nps lod is a copy of 75 and ss 150。
Table 1. Automatic extraction of oktopure
Sbedex principle characteristics
All sbedex reagent boxes are combined with nucleic acid through a new, two-step combination mechanism using surface-modified ultra-magnetic beads (figure 4)。
First, the nucleic acid is adsorbed on the magnetic beads through polar interactions, and second, the nucleic acid is combined with magnetic beads through a proximate and force-driven nucleus. The second combined step allows the use of water-based washing solutions for final washing steps without the need for drying, avoiding damage to the quality of nucleic acids that may result from over-drying and minimizing ethanol that eventually enters the scrubber. The two-step combination mechanism and ethanol-free washing can help clients acquire high-purity high-quality nucleic acid extraction products。
Figure 4. Sbedex nuclear acid extraction rationale
Main strengths:
Experienced nucleic acid extraction programmes applicable to viruses, bacteria and fungi
2. Automation platforms applicable to the mainstream, such as kingfisher, oktopure, etc.
Three, 25 minutes quick extraction and 45 minutes standard extraction options
4. An organic solvent extraction, ethanol deposition or centrifugal steps
5. Water-based scrubbing buffers to remove inhibitors from the leachate
6. High-quality nucleic acid extracts, applicable to all downstream applications, such as pcr/qpcr and ngs
7, reagent packs, or tailoring and directed development to client needs。
