Metabolomic sequencing samples and sample size requirements

In the vast field of life science research, metabolomics is gradually emerging as an emerging and dynamic discipline. Through a comprehensive analysis of metabolites in organisms, it reveals the metabolic changes of organisms under the action of different physiological states, disease courses and environmental factors. However, in order to accurately capture these metabolic changes, the sample and sample size requirements for metabolomic sequencing are particularly important.

Metabolomics sequencing samples

1. Sample type Metabolomics studies have a variety of samples, common ones include: 1. Animal or clinical samples: such as serum, plasma, urine, saliva, cerebrospinal fluid, joint fluid, lymph fluid, tissue (such as liver, kidney, muscle, tumor, etc.), cells and their culture medium, etc. 2. Plant samples: such as leaves, stems, flowers, roots, fruits, etc. 3. Microbial samples: such as bacteria, fungi and their fermentation broth, etc.

2. Sample collection and preservation. Different types of samples should be paid attention to the following points when collecting and preserving: 1. Blood samples (serum, plasma): Collect blood from suitable veins, usually the collection amount is about 10mL. Collect using lithium heparin anticoagulation tube (for plasma) or a regular serum tube. Serum or plasma are isolated by centrifugation and transfer to a freeze storage tube in aliquots (such as 0.5 mL or 0.1 mL). Store in a -80℃ refrigerator and use sufficient dry ice when shipping. 2. Urine sample: Collect mid-section urine from morning (clinical) or 1 hour of urine in the morning (animal). Aliquot directly into centrifuge tubes, about 1 mL per tube. Add sodium azide as a preservative and centrifuge as soon as possible. Store in a -80℃ refrigerator and use sufficient dry ice when shipping. 3. Tissue sample: After removing tissue from animals or humans, rinse it with normal saline. Use clean dust-free absorbent paper to absorb the moisture, then fill it into the marked centrifuge tube in equal parts (such as 20mg/tube). Quickly put it in liquid nitrogen and freeze it for at least 15 minutes, and then store it in a -80°C refrigerator. 4. Cell sample: For suspended cells, transfer them to a centrifuge tube together with the culture medium and pellet the cells by centrifugation at low speed. After pouring out the culture medium, wash it with PBS once, centrifuge it again and then pour out the PBS. The tip of the centrifuge tube was inserted into liquid nitrogen to quench the cells and then stored in a -80°C refrigerator. For adherent cells, first pour out the medium, wash it once with PBS, and then contact the bottom of the culture dish with liquid nitrogen to quench the cells. Add methanol-aqueous solution to scrape the cells and transfer them to a centrifuge tube, and store them in a refrigerator of -80°C. 5. Plant sample: Take the whole leaf, a stem, a flower or root, and quickly rinse it with PBS to remove soil impurities, etc. Place in a centrifuge tube or tin foil and mark it and quickly put it in liquid nitrogen for freezing treatment for at least 15 minutes. After removal, quickly put it in a self-sealing bag and indicate the sample information. Store in a -80℃ refrigerator and use sufficient dry ice when shipping. 6. Microbial sample: Transfer the microorganisms and culture medium to a centrifuge tube and centrifuge at low speed to precipitate the bacteria. After pouring out the culture medium, wash it once with PBS and centrifuge the bacteria to collect the bacteria. Insert the tip of the centrifuge tube into liquid nitrogen to quench the bacteria and store it in a -80°C refrigerator.

3. Sample size requirements. The sample size requirements for metabolomic sequencing are usually determined based on the experimental design and sample type. Generally speaking: 1. Blood sample: At least 2 ml is required. 2. Urine sample: The recommended amount is no less than 5 ml. 3. Tissue sample: Depending on the type of tissue and experimental needs, generally not less than 50 mg. 4. Cell sample: For mammalian cell lines, 1×10^7 cells/sample are usually required. 5. Plant and microbial samples: The sample size depends on the specific experimental needs, but it usually needs to reach a certain number to ensure the accuracy and repetition of the detection. Too small sample size may affect the accuracy and repetition of the detection to a certain extent. Therefore, when collecting samples, try to collect and freeze them as much as possible to avoid repeated freeze-thawing. At the same time, sufficient amount of dry ice should be used when sending samples to ensure the stability of the samples during transportation.

The success of metabolomic sequencing depends largely on sample selection and sample size design. Reasonable sample selection can ensure the representativeness of the data, while appropriate sample size can improve the statistical efficacy of the experiment and help researchers better discover metastatic differences.

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