1 0

    Murine Normal Colon Organoid Culture Medium

    Murine Normal Colon Organoid Culture Medium

    Catalog Number: abs9984 Brand: Absin
    Share:
    Price:
    $476 USD
    $476 USD
    Size:

    For shipping services or bulk orders, you may request a quotation.
    Secure checkout with
    View full details

    Product Details

    Product Specification

    Usage 1 , original  
    （ 1 The normal colon tissue of the mice after sampling was placed in a precooled ( 2-8°C The tissue preservation solution was quickly transported to a clean laboratory for tissue processing and stem cell separation, photographed, and information recorded.
    （ 2 Prepare several petri dishes and add... 4℃ Precooled mouse normal colon organoid tissue primary culture buffer ( abs9731 ) Backup.
    （ 3 The sample bottle was sterilized, and the tissue was placed in a petri dish. After washing three times with primary culture buffer for normal mouse colon organoids, the tissue was cut into a volume of approximately [volume missing] with ophthalmic scissors or a scalpel. 1-3mm3 Tissue blocks.
    （ 4 Primary tissue digestive solution for normal colon tissue in mice for tissue use ( abs9644 Digestion, 4℃ 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 70μm The following cell clusters were followed by the addition of three times the volume of mouse normal colon organoid tissue primary culture buffer to stop digestion.
    （ 6 ) Use 100μm 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 mouse normal colon 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 Twice the tissue volume of matrix glue was resuspended from the plates.
    （ 8 ） 24 Taking well cell culture plates as an example, each well is dispensed with gel. 25μL 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 mouse normal colon organoid 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 ) Place 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 mouse normal colon 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 digestive fluid ( abs9520 Digestion 2-3min Add normal mouse colon organoid subculture buffer to terminate digestion and centrifuge. 5min Discard the mixture, add an appropriate amount of mouse normal colon 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. 25μL Matrix glue is laid on 24 In well cell culture plates, place in an incubator. 10-15min Add 500μL Mouse normal colon organoid medium.
    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 mouse normal colon organoid subculture buffer to resuspend again, 300g centrifuge 5min Discard the liquid.
    （ 4 Add an appropriate amount of organoid cryopreservation solution, gently blow and resuspend, and 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 Mouse normal colon 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 normal mouse colon organoid subculture buffer for resuspension and transfer. 1.5mL centrifuge tube 300g centrifuge 5min 。
    （ 5 Resuspend the matrix gel, per well 25μL Matrix glue is laid on 24 In well cell culture plates, place in an incubator. 10-15min Gelation, addition 500μL Mouse normal colon organoid medium.
    Storage Temp. Store at -20°C with a shelf life of 12 months.