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Notes on sample processing in ELISA experiments
There are various types of samples detected by ELISA kits, such as serum, plasma, urine, cell culture supernatant, or homogenate, etc. The pre-treatment methods for different types of test samples are different. The processing of experimental samples can be considered one of the key processes in ELISA testing, so research friends must pay more attention. Below we will give you a detailed explanation!
1、 Homogenicity
① Arrange samples: Cut meat and liver food into small pieces, repeat grinding with a meat grinder, and mix evenly.
② Aquatic sample: Remove the non edible part of the sample, cut the edible part finely, and homogenize with a homogenizer; When the surface of the material is dirty, it needs to be cleaned with distilled water appropriately.
③ Eggs: Remove the shell of fresh eggs, mix the yolk and protein well.
④ Fruits and vegetables: Wash the sediment with water first, then remove the external moisture, and take the edible part.
2、 Vibration extraction
Add the extraction solvent to the stoppered container containing the sample, vibrate it to allow sufficient contact between the extraction solvent and the sample inside the container to penetrate deeper into the sample arrangement and extract the components to be tested.
① Vibration method: Perform up and down, reciprocating vibration, and manual up and down vibration on the vibrator.
② When arranging samples for extraction between organic solvents, the ELISA kit should be vibrated at the same time to prevent clumping, which is not conducive to extraction.
3、 Emulsification phenomenon
If emulsification occurs during the extraction process using organic solvents, the treatment methods include:
① Gently mix with a thin head, damage the emulsion, and then repeat centrifugation.
② Add an appropriate amount of extractant and vibrate again. Please ensure that the dilution ratio of the sample remains unchanged after centrifugation.
4、 Concentrated
Because the solvents introduced during the purification process may reduce the concentration of the tested components or be unsuitable for direct analysis, it is necessary to remove all organic solvents. During the pre-treatment process of the reagent kit, blow dry the sample under nitrogen at 60 ℃, and then dissolve the dry residue in a complex solution.
Concentration method: Nitrogen blowing for impurity removal, compressed air blowing for impurity removal.
Please note:
① Before drying the sample, clean the needle with methanol to prevent impurities from disturbing.
② When blowing samples, the needle should be above the liquid level to prevent contact with the sample and prevent cross contamination.
③ After drying the sample, it should be immediately removed to prevent the blowing time from being too long and affecting the final test results.
④ Different drugs have different shelf life of samples after air drying. ELISA kits advocate for immediate re dissolution of samples after they return to room temperature.
5、 Purification
The extracted components to be tested generally contain impurities that can disturb the antigen antibody response in the reagent kit or impurities with similar structures to the tested substance. The process of separating the tested component from impurities is called purification of the sample in the reagent kit. The common purification method involved in our existing pre-treatment methods for reagent kits is the liquid-liquid distribution method. For example, before dissolving the dry residue with a complex solution, add an appropriate amount of n-hexane. If emulsification occurs after participating in n-hexane and complex solution turbulence, it can be heated in a 60 ℃ water bath until the emulsification phenomenon disappears or weakens. Then, increase the centrifugal speed for centrifugation.
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