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Description
Human Gastric Organoid Culture MediumDescription Protocol 1. Primary culture (1) The collected tissue is placed in a sampling bottle of pre cooled (2 8 C) tissue preservation solution, and quickly transferred to a clean laboratory for tissue processing and cell separation, taking photos and registering information. (2) Prepare several petri dishes, and add 4 pre cooled human normal gastric organoid tissue primary culture buffer # abs9731 for later use. (3) Sterilize the sampling bottle,
Description
Protocol
1. Primary culture
(1) The collected tissue is placed in a sampling bottle of pre-cooled (2-8 °C) tissue preservation solution, and quickly transferred to a clean laboratory for tissue processing and cell separation, taking photos and registering information.
(2) Prepare several petri dishes, and add 4 ℃ pre-cooled human normal gastric organoid tissue primary culture buffer # abs9731 for later use.
(3) Sterilize the sampling bottle, put the tissue into a petri dish, wash it three times with the primary culture buffer of human normal gastric organoid tissue, and cut the tissue into a volume of about 1-3mm with ophthalmic scissors or scalpel3The tissue block.
(4) The tissues were digested with human normal stomach primary tissue digestive juice # abs9783, and digested by shaking at 4 ℃ for 15-20min (observe the digestion at any time during the digestion process).
(5) Take a small amount of liquid and observe it under a microscope. After more single cells or cell clusters below 70um are observed under the microscope, add triploid human normal gastric organoid tissue primary culture buffer to stop digestion.
(6) Use a sieve with a pore size of 100um for filtration, collect the filtrate, enrich and centrifuge at 300g for 5 minutes, remove the supernatant, add human normal gastric organoid tissue primary culture buffer and re-suspend and centrifuge.
(7) Matrigel calculation: After step 6, observe the collected tissue volume, add 25 times the tissue volume of Matrigel # abs9495 to resuspend and plate.
(8) Take a 24-well cell culture plate as an example. Each well is dispensed with 25ul tissue matrigel mixture for plating (operation at 4 °C).
(9) Put the laid culture plate into a 37 ℃ incubator for 10-15 minutes to form a gel, and add human normal gastric organoid culture medium (restored to room temperature) for culture.
2. Organoid subculture
(1) Aspirate the medium with a pipette gun, add 1-2ml of 4 ℃ organoid subculture buffer # abs9730 to each well for 2min.
(2) Gently blow the matrigel with a pipette gun, collect it in a 15ml centrifuge tube, and let it stand at 4 °C for 10 minutes. (groups of 6-8 wells)
(3) a: When the number of organoids is insufficient or the volume is small: centrifuge for 5 minutes to discard the supernatant, add an appropriate amount of human normal gastric organoid subculture buffer to resuspend and transfer it into a 1.5 ml centrifuge tube, centrifuge 300g for 5 minutes to discard the liquid for step 4.
b: When the number of organoids is large or the volume is large: centrifuge for 5 minutes to discard the supernatant, add an appropriate amount of organoid subculture juice # abs9520 for 2-3 minutes, add human normal gastric organoid subculture buffer to terminate digestion, centrifuge for 5 minutes to discard the mixed solution, add an appropriate amount of human normal gastric organoid subculture buffer to resuspend and transfer into a 1.5 ml centrifuge tube, and centrifuge 300g for 5 minutes to discard the liquid for step 4.
(4) After organoid collection, add Matrigel for resuspension, spread 25ul of Matrigel per well in a 24-well cell culture plate, place it in an incubator for 10-15min, and add 500ul of human normal gastric organoid culture medium.
3. Organoid cryopreservation
(1) Aspirate the culture medium with a pipette gun, add 1-2ml of 4 ℃ organoid subculture buffer to each well and place it for 2 minutes.
(2) Gently blow the matrigel with a pipette gun, collect it in a 15ml centrifuge tube, and let it stand at 4 °C for 10 minutes. (groups of 6-8 wells)
(3) Centrifuge for 5 minutes to discard the supernatant, add an appropriate amount of human normal gastric organoid subculture buffer to resuspend again, and centrifuge 300g for 5 minutes to discard the liquid.
(4) Add an appropriate amount of organoid cryopreservation solution # abs9519, gently blow and resuspend, take a 24-well cell culture plate as an example: the density is 2 wells and cryopreserve 1 tube, each tube has a volume of 1.4 ml.
(5) Make good marking information, carry out program cooling, and then move it into liquid nitrogen for long-term storage.
4. Organoid resuscitation
(1) Take 10ml of human normal gastric organoid subculture buffer in a 15ml centrifuge tube.
(2) Take out the frozen organoid cells from the liquid nitrogen tank and quickly thaw them in a 37 ℃ water bath.
(3) During the water bath melting process, the cryotube needs to be gently shaken to ensure that the cryopreservation solution is completely melted within 1-2 minutes.
(4) Quickly transfer the dissolved organoid cells to a 15ml centrifuge tube, gently pipette 6-8 times with a pipette, centrifuge at 300g for 5min, then remove the supernatant and collect the organoid cell pellet. Add appropriate amount of human normal gastric organoid subculture buffer, resuspend, transfer into 1.5 ml centrifuge tube 300g and centrifuge for 5min.
(5) Resuspend Matrigel, spread 25ul Matrigel per well in a 24-well cell culture plate, place it in an incubator for 10-15min to form a gel, and add 500ul human normal gastric organoid culture medium.
(1) The collected tissue is placed in a sampling bottle of pre-cooled (2-8 °C) tissue preservation solution, and quickly transferred to a clean laboratory for tissue processing and cell separation, taking photos and registering information.
(2) Prepare several petri dishes, and add 4 ℃ pre-cooled human normal gastric organoid tissue primary culture buffer # abs9731 for later use.
(3) Sterilize the sampling bottle, put the tissue into a petri dish, wash it three times with the primary culture buffer of human normal gastric organoid tissue, and cut the tissue into a volume of about 1-3mm with ophthalmic scissors or scalpel3The tissue block.
(4) The tissues were digested with human normal stomach primary tissue digestive juice # abs9783, and digested by shaking at 4 ℃ for 15-20min (observe the digestion at any time during the digestion process).
(5) Take a small amount of liquid and observe it under a microscope. After more single cells or cell clusters below 70um are observed under the microscope, add triploid human normal gastric organoid tissue primary culture buffer to stop digestion.
(6) Use a sieve with a pore size of 100um for filtration, collect the filtrate, enrich and centrifuge at 300g for 5 minutes, remove the supernatant, add human normal gastric organoid tissue primary culture buffer and re-suspend and centrifuge.
(7) Matrigel calculation: After step 6, observe the collected tissue volume, add 25 times the tissue volume of Matrigel # abs9495 to resuspend and plate.
(8) Take a 24-well cell culture plate as an example. Each well is dispensed with 25ul tissue matrigel mixture for plating (operation at 4 °C).
(9) Put the laid culture plate into a 37 ℃ incubator for 10-15 minutes to form a gel, and add human normal gastric organoid culture medium (restored to room temperature) for culture.
2. Organoid subculture
(1) Aspirate the medium with a pipette gun, add 1-2ml of 4 ℃ organoid subculture buffer # abs9730 to each well for 2min.
(2) Gently blow the matrigel with a pipette gun, collect it in a 15ml centrifuge tube, and let it stand at 4 °C for 10 minutes. (groups of 6-8 wells)
(3) a: When the number of organoids is insufficient or the volume is small: centrifuge for 5 minutes to discard the supernatant, add an appropriate amount of human normal gastric organoid subculture buffer to resuspend and transfer it into a 1.5 ml centrifuge tube, centrifuge 300g for 5 minutes to discard the liquid for step 4.
b: When the number of organoids is large or the volume is large: centrifuge for 5 minutes to discard the supernatant, add an appropriate amount of organoid subculture juice # abs9520 for 2-3 minutes, add human normal gastric organoid subculture buffer to terminate digestion, centrifuge for 5 minutes to discard the mixed solution, add an appropriate amount of human normal gastric organoid subculture buffer to resuspend and transfer into a 1.5 ml centrifuge tube, and centrifuge 300g for 5 minutes to discard the liquid for step 4.
(4) After organoid collection, add Matrigel for resuspension, spread 25ul of Matrigel per well in a 24-well cell culture plate, place it in an incubator for 10-15min, and add 500ul of human normal gastric organoid culture medium.
3. Organoid cryopreservation
(1) Aspirate the culture medium with a pipette gun, add 1-2ml of 4 ℃ organoid subculture buffer to each well and place it for 2 minutes.
(2) Gently blow the matrigel with a pipette gun, collect it in a 15ml centrifuge tube, and let it stand at 4 °C for 10 minutes. (groups of 6-8 wells)
(3) Centrifuge for 5 minutes to discard the supernatant, add an appropriate amount of human normal gastric organoid subculture buffer to resuspend again, and centrifuge 300g for 5 minutes to discard the liquid.
(4) Add an appropriate amount of organoid cryopreservation solution # abs9519, gently blow and resuspend, take a 24-well cell culture plate as an example: the density is 2 wells and cryopreserve 1 tube, each tube has a volume of 1.4 ml.
(5) Make good marking information, carry out program cooling, and then move it into liquid nitrogen for long-term storage.
4. Organoid resuscitation
(1) Take 10ml of human normal gastric organoid subculture buffer in a 15ml centrifuge tube.
(2) Take out the frozen organoid cells from the liquid nitrogen tank and quickly thaw them in a 37 ℃ water bath.
(3) During the water bath melting process, the cryotube needs to be gently shaken to ensure that the cryopreservation solution is completely melted within 1-2 minutes.
(4) Quickly transfer the dissolved organoid cells to a 15ml centrifuge tube, gently pipette 6-8 times with a pipette, centrifuge at 300g for 5min, then remove the supernatant and collect the organoid cell pellet. Add appropriate amount of human normal gastric organoid subculture buffer, resuspend, transfer into 1.5 ml centrifuge tube 300g and centrifuge for 5min.
(5) Resuspend Matrigel, spread 25ul Matrigel per well in a 24-well cell culture plate, place it in an incubator for 10-15min to form a gel, and add 500ul human normal gastric organoid culture medium.
Storage Temp.
Store at 4 °C, shelf life is 3 months; Store at-20 °C, shelf life 1 year.
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4.6 ★★★★★
Based on 29 reviews
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Product Reviews
★★★★★ 4
Not indestructable, but very sturdy
Size: Medium, Style: Beef
I have recently been blessed (and cursed) with my first Super Chewer dog, a year old Australian Shepherd. She will chew herself out of boredom, so it's imperative we always have sturdy toys readily available for her, preferably in every room.
Playology and Kong by and large have been the best toys we can find for her. Kong has a very good rubber ball, bone, and ring that have held up spectacularly (no noticeable damage after months of use, except maybe some pin-prick teeth marks). Dogs need a wide range of textures though for enrichment, so we've been dabbling with the Playology line (the tire, plush bone, dry-tech rope), including this ball.
The scent factor is definitely interesting. Most people know Playology for the peanut butter scents, so I was very excited to find a multitude of flavors available on Amazon, especially with the different scents clearly labeled. I choose the red or beef flavored since the only two in the medium size in stock at the time were beef and sweet potato. The scent is indeed noticeable to humans, but it's not too strong or overwhelming on its own (walking down the Playology aisle in Petsmart can be overwhelming though since they have dozens of dozens of scented toys, much like going down a candle aisle).
I can't speak for the dog on how she feels about the scent, but she took to the ball immediately, same as the normal peanut butter ones we get. The medium size is pretty much perfect for her (she's just under 50lbs). It's got decent bounce for playing fetch in the house, but not so bouncy that we're afraid to use it in the house. It's got a decent density to it as well that adds to the sturdiness.
The biggest issue is the squeaker. It's not an obnoxious squeaker by any means, and the dog enjoys it very much. However, when you put a squeaker in any toy, it reduces the density and creates a weak spot dogs can exploit. She did good with the ball at first and was able to sit and chew on it for minutes as a time without doing any real damage (her teeth leave lots of pin-prick teeth marks, but they don't do any real damage to the ball, just superficial marks).
Aussies are considered incredibly smart though, and it didn't take her long to learn she could rip out the bottom of the ball where the squeaker is sealed inside to tear the squeaker out, thus ripping the squeaker apart and compromising the structural integrity of the ball. We're debating if we want to toss the ball in the trash or attempt letting her play with it in a supervised session without the squeaker to see if it is safe.
So, overall, this is a very tough ball. It does accumulate teeth marks that are essentially harmless, but there is a weak spot in the seam where the squeaker is inserted, which means this is not indestructible. It is by and large a great ball for fetch and likely won't ever be destroyed unless a dog is left with it long enough to chew a hole through that squeaker seam. Playology balls are pretty costly though, so the risk of it becoming unusable once the squeaker is ripped out is a bit off-putting. The bright side is that if you monitor your dog, you can likely get several weeks or months of use out of the ball; just no unsupervised play sessions.
7/10, durable, bouncy, and the scent keeps the dog interested, but the ball is not good unsupervised as it can be destroyed if the squeaker is ripped out. We will continue to buy Playology toys, but we will likely not buy too many of these balls until a more durable one (maybe without a squeaker) is released.
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Reviewed in the United States on March 20, 2022
★★★★★ 1
No. Just no. My own dog shamed me…
Size: Medium, Style: Beef, Size: Medium, Style: Beef
My German Shepherd likes squeaky balls. She has lots of them. Most are similar to a tennis ball. By the way, don’t give your dog an actual tennis ball. The outer material is made for grip for tennis. It’s abrasive and is not good for your dog’s teeth enamel at all.
Anyway, for $14.99, I thought I’d give this one a try. The pictured packaging alone was an eye grabber. The picture you see on Amazon is a cardboard with item specific info on the card: flavor, size, what it is (squeaky ball). The pictured product also has a hole in the card to secure and present the item the card describes. Even two color coded red zip ties securing the ball in the card hole. Awesome presentation and frankly that’s what made me give this a shot. So, as far as the presented product/packaging on Amazon, great job! I even sent a link to a buddy and he bought one too.
Well, I cannot speak for everyone, but what I received was what’s pictured with my review. A red ball in a clear bag, along with a card that describes the product line. Nothing specific to what I actually bought (like the slick item-specific packaging on Amazon. To find that, I had to read the condensed description on the Amazon specific bar code that is right on top of a different barcode. This kind of thing displeases me. I want what grabbed my attention on amazon. No, this was not a return. This is how they come.
I figured with item specific info on the packaging, the company is putting some money into packaging that makes you feel like you bought something specific. Pictured one says all natural beef. The chicken one says chicken. Pictured one says the size. Heck, I’d of been happy to at least have the ball attached to a card that at least says it’s a squeaky ball. Nope.
Cost cutting is not ok when you are selling a beef flavored, medium size squeaky ball for $14.99.
What arrives is an advertisement for the whole line, nothing that says what it is, the flavor, the size (unless you try to make out the Amazon bar code desc). Just a ball in a bag with an advertisement to buy other stuff they make.
Maybe you received what’s pictured. I texted my buddy and asked him what he received. The exact same thing I did.
In my opinion, when one cuts corners on packaging, I have no doubt corners could have been cut with the product itself. The squeak isn’t as easy as kong ball squeaker. Takes effort. So if your dog likes to play by walking/running around while constantly biting and getting that pleasing squeak each time, that didn’t happen here. Yes, I know the tennis ball type squeakers are different. But this says it’s a squeaker ball. Not just a squeaker ball, but a flavored squeaker ball. Mine smelled like rubber. Where’s the beef?
I threw it a few times. My dog chased it a bit, did her usual “what’s this” evaluation, bit into it a few times with her very strong jaws, only to have no squeak. God as my witness, she dropped it, looked at me with what I insist is a look of disappointment, and walked away. She then went to the big bin of a huge variety of squeaky balls and grabbed one. She then went upstairs, with a squeak every second or so as she went up.
I went and picked this one up with slobber and all. To get this to squeak requires opposing thumbs. So I suppose it could be marketed as a hand strengthener that squeaks.
Friends lab had zero interest as well. So, I’m gong to actually return a dog toy. This was a disappointment. If it had come carded the way it shows on Amazon, I would not be as frustrated. I won’t take a chance when corners are cut when it comes to my dog. Update the picture to what I received is my advice to the company. I have a feeling what I got wouldn’t sell as easily.
Topping it off, I noticed something odd. It doesn’t squeak when squeezing it, only when it is released to go back to its regular shape. So it’s a reverse reward. Weird. All the squeaker balls we have make a noise both when squeezing and releasing.
Sorry Olina (my pooch).
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Reviewed in the United States on April 25, 2023
★★★★★ 5
Best Toy EVER!
Size: Jumbo, Style: Peanut Butter
I was a bit skeptical. We have had NO joy in finding toys for "aggressive" chewers that actually last more than a day for our 1 year old Staffordshire/Pittie mix. He has jaws of iron! (or so it seems) Nothing holds up. He is literally eating them, and subsequently gets them taken away, within 24 to 48 h ours. IF that long. He finds it a challenge to get to the center of whatever toy he has, whether it is supposed to have a center or not! We bought this in April of last year!! Yes, the squeaker is no longer squeaking (a blessing in disguise) but the ball itself is still intact, with no nicks, dings, pieces missing, etc. It still bounces well, and he still loves it. 1 year later, still going strong! I didn't realize it had been that long, until today. So I had to write a review, and buy a couple more Playology Jumbo toys. I hope they hold up 1 tenth as well!
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Reviewed in the United States on December 20, 2025
★★★★★ 3
Hit or miss
Size: Medium, Style: Beef, Size: Medium, Style: Beef
I’ve purchased this item multiple times. Twice now, I got duds. Within minutes of opening the package, the ball was destroyed. The other times, it lasted months! I don’t know how to explain it.
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Reviewed in the United States on April 22, 2026
★★★★★ 5
Chew toy that will last!
Color: Blue&Red Fish
Great toy for a real chewer. It's been in the house for about 5 days and the toy is still intact and the squeaker is still squeaking.
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Reviewed in the United States on May 28, 2026