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    Human Gliomatosis Cerebri Organoid Culture Medium

    Human Gliomatosis Cerebri Organoid Culture Medium

    Catalog Number: abs9740 Brand: Absin
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    $476 USD
    $476 USD
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    Product Specification

    Usage 1 , original  
    （ 1 The removed tissue was placed in a pre-cooled ( 2-8°C The tissue preservation solution was quickly transported to a clean laboratory for tissue processing and cell separation, photographed, and information recorded.
    （ 2 Prepare several petri dishes and add... 4℃ Precooled human glioma organoid tissue primary culture buffer #abs9731 Backup.
    （ 3 The sample bottle is sterilized, the tissue is placed in a petri dish, washed three times with human glioma organoid tissue primary culture buffer, and then the tissue is cut into a volume of approximately [size missing] using ophthalmic scissors or a scalpel. 1-3mm3 Tissue blocks.
    （ 4 Human glioma primary tissue digestive fluid for tissue use #abs9522 digestion, 37℃ oscillation digestion 10-20min (Observe the digestion situation at any time during the digestion process).
    （ 5 Take a small amount of liquid and observe it under a microscope. A large number of individual cells or 70um After the following cell clusters, digestion was terminated by adding three times the volume of human glioma organoid tissue primary culture buffer.
    （ 6 ) Use 100um The filtrate is filtered using a sieve with a certain pore size, and the filtrate is collected and then... 300g enrichment centrifugation 5min The supernatant was then removed, and human glioma organoid tissue primary culture buffer was added to resuspend and centrifuge.
    （ 7 Matrix rubber calculation: No. 6 After the step, observe the collected tissue volume and add 25 double tissue volume matrix glue #abs9495 Re-hang the slab.
    （ 8 ） 24 Taking well cell culture plates as an example, each well is dispensed with gel. 25ul Tissue matrix glue mixture for board laying ( 4℃ (Next operation).
    （ 9 Place the prepared culture plate in... 37℃ Incubator 10-15min Gelatinization was performed, and human glioma organoid culture medium was added (restored to room temperature) for culture.
    2 Organoid subculture
    （ 1 Remove the culture medium with a pipette and add to each well. 1-2ml 4℃ organoid subculture buffer #abs9730 placement 2min 。
    （ 2 Gently blow the matrix glue with a pipette to collect... 15ml In the centrifuge tube, 4℃ standing 10min 。 (each 6-8 (The holes are a group)
    （ 3 ） a : Insufficient number or small volume of organoids: centrifuge 5min Discard the supernatant, add an appropriate amount of human glioma organoid subculture buffer, resuspend and transfer. 1.5ml centrifuge tube, 300g centrifuge 5min Discard the liquid for the first 4 step.
             b When the number or volume of organoids is large: Centrifuge 5min Discard the supernatant and add an appropriate amount of organoid passage digestion solution. #abs9520 digestion 2-3min Add human glioma organoid subculture buffer to terminate digestion and centrifuge. 5min Discard the mixture, add an appropriate amount of human glioma organoid subculture buffer, resuspend and transfer. 1.5ml centrifuge tube, 300g centrifuge 5min Discard the liquid for the first 4 step.
    （ 4 After organoids were collected, matrix gel was added for resuspension, and each well was resuspended. 25ul Matrix glue is laid on 24 In well cell culture plates, place in an incubator. 10-15min Add 500ul Organoid medium for human gliomas.
    3 Organoid cryopreservation
    （ 1 Remove the culture medium with a pipette and add to each well. 1-2ml 4℃ organoid subculture buffer placement 2min 。
    （ 2 Gently blow the matrix glue with a pipette to collect... 15ml In the centrifuge tube, 4℃ standing 10min 。 (each 6-8 (The holes are a group)
    （ 3 Centrifugation 5min Discarding the supernatant, adding an appropriate amount of human glioma organoid subculture buffer for resuspension, 300g centrifuge 5min Discard the liquid.
    （ 4 Add an appropriate amount of organoid cryopreservation solution. #abs9519 Gently blowing and resuspending, 24 Taking well cell culture plates as an example: the density is 2 Individual hole cryopreservation 1 Tubes, volume per tube 1.4ml 。
    （ 5 Mark the information, cool it down procedurally, and then transfer it to liquid nitrogen for long-term storage.
    4 Organoid resuscitation
    （ 1 ) take 10ml Human glioma organoid subculture buffer 15ml In the centrifuge tube.
    （ 2 Remove the frozen organoid cells from the liquid nitrogen tank and quickly place them in... 37℃ Melt in a water bath.
    （ 3 During the water bath thawing process, the freezing tube needs to be gently shaken to ensure that the cryopreservation solution is... 1-2min It completely dissolves inside.
    （ 4 ) Rapidly transfer the lysed organoid cells to 15ml Centrifuge tube, gently blow with a pipette 6-8 next time, 300g centrifuge 5min The supernatant is then removed and the organoid cell pellet is collected. Add an appropriate amount of human glioma organoid subculture buffer to resuspend and transfer 1.5ml centrifuge tube 300g centrifuge 5min 。
    （ 5 Resuspend the matrix gel, per well 25ul Matrix glue is laid on 24 In well cell culture plates, place in an incubator. 10-15min Gelation, addition 500ul Organoid medium for human gliomas.
    Storage Temp. Store at -20°C with a shelf life of 1 year.