Method and kit for extracting exosomes and biomacromolecules combined with them
A macromolecule stabilizer and exosome technology, which is applied in diagnostic kits and cell derivation, specifically in the field of detection and identification, can solve the problems of high cost, damage to the integrity of exosomes, and labor consumption.
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Embodiment 1
[0097] Example 1: Extraction of cell-derived vesicles from biological fluids ( figure 2 )
[0098] In a reaction tube, fill any biological fluid less than 4 milliliters, more suitably less than 3 milliliters of biological fluid, optimally at least 50 microliters of biological fluid, or at least 100 microliters, at least 200 microliters liters, 500 microliters, at least 1 milliliter, or at least 2 milliliters of biological fluid. Add 1% phosphate buffer saline at a ratio of 40% (v / v) to the biological sample, and mix well. Then add 1.5% (w / w), 2% (w / w), or 2.5% (w / w) of the macromolecular stabilizer, wherein the macromolecular stabilizer is most preferably the general formula of C 6 h 12 o 7 The sodium salt of the organic compound was mixed by pipetting up and down. Then, a volume-exclusion polymer containing low molecular weight polyethylene glycol was added to the mixture, and mixed with a pipette gun up and down to obtain a mixture. Wherein, the volume exclusion polym...
Embodiment 2
[0099] Example 2: Extraction of cell-derived vesicles from cell culture medium ( image 3 )
[0100] In a reaction tube, less than 4 milliliters of conditioned cell culture medium is filled, optimally at least 50 microliters, or at least 100 microliters, at least 200 microliters, at least 500 microliters, at least 1 milliliter, or at least 1 milliliter. Add 1.5% (w / w), 2% (w / w), or 2.5% (w / w) of a macromolecular stabilizer to the biological sample, wherein the macromolecular stabilizer is optimally characterized by a general formula of C 6 h 12 o 7 The sodium salt of the organic compound was mixed by pipetting up and down. Then, a volume exclusion polymer containing low molecular weight polyethylene glycol was added to the mixture, and mixed by pipetting up and down to obtain a mixture. Wherein, the volume exclusion polymer preferably contains polyethylene glycol with a molecular weight not greater than 5000Da and not less than 3000Da, and contains at least 20% (w / v), 22% ...
Embodiment 3
[0101] Example 3: Estimating the content of exosomal proteins in the extracted cell-derived vesicles ( Figure 4 )
[0102] refer to Figure 4 , shows the protein yields obtained by extraction in some embodiments. Among them, in these embodiments, the relevant steps of the prior art in the related field are improved according to the above-mentioned method, and the above-mentioned experimental steps are followed, and the amount of exosome protein obtained is 75 micrograms to 500 micrograms per milliliter of biological samples, Wherein said biological sample includes cell culture medium of a cell line or a plurality of cell lines.
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