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Description
Organotial Human bladder Cancer organoid Culture KitProduct Specification Description kit composition: Name Specification Human bladder cancer organoid medium A 100mL Primary culture buffer B 250mL Human bladder cancer primary tissue digestive juice C 30mL Organoid passage digestive juice D 30mL Tissue preservation solution E 100mL Organoid cryopreservation fluid F 20mL Organoid subculture buffer G 250mL Instructions 1, primary (1) After sampling the tissue should be placed in pre cooled (2 8 C) In the
Product Specification
| Description |
kit composition:
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| Instructions |
1, primary (1) After sampling the tissue should be placed in pre-cooled (2-8° C) In the sampling bottle of tissue preservation solution E, the tissue was quickly transferred to a clean laboratory for tissue processing and cell separation, photographed and recorded. (2) Prepare several culture dishes and add primary culture buffer B precooled at 4 ° C for later use. (3) The sampling bottle was disinfected, and the tissue was placed in the Petri dish. After washing three times with primary culture buffer B, the tissue was cut into tissue blocks of about 1-3mm3in volume using ophthalmic scissors or scalpel. (4) The tissue was digested with human bladder cancer primary tissue digestion solution C, and the digestion was shaken at 37℃ for 10-20min (the digestion was observed at any time during digestion). (5) A small amount of liquid was observed under the microscope. When more single cells or cell clusters below 70um were observed under the microscope, triploid volume primary culture buffer B was added to terminate the digestion. (6) The filtrate was filtered by a sieve with a pore size of 100um, and the filtrate was collected. After enrichment and centrifugation at 300g for 5min, the supernatant was removed, and the primary culture buffer B was added and resuspended for centrifugation. (7) Matrix glue calculation: The volume of collected tissue was observed after step 6, and matrix glue (abs9495) was added 25 times the tissue volume to resuspend the paving plate. (8) For the example of 24-well cell culture plates, 25uL of tissue matrix glue mixture was dispensed to each well for paving (operation at 4 ° C). (9) The paved culture plate was placed in an incubator at 37 ° C for 10-15min to form glue, and human bladder cancer organoid medium A (restored to room temperature) was added for culture. 2, Organoid subculture (1) The medium was sucked out by pipetting gun, and 1-2mL of 4℃ organoid subculture buffer G was added to each well and placed for 2min. (2) The matrix glue was gently blown by a pipetting gun, collected in a 15mL centrifuge tube and allowed to rest for 10min at 4 ° C. (every 6-8 Wells were a group) (3) a: When the number of organoids was insufficient or the volume was small: the supernatant was discarded by centrifugation for 5min, then an appropriate amount of organoid passage culture buffer G was added and resuspended into a 1.5mL centrifuge tube, and the liquid was discarded by centrifugation at 300g for 5min for the fourth step. b: When organoids are large in number or size: The supernatant was discarded by centrifugation for 5min, then the organoid passage digestion solution D was added for 2-3min, then the organoid passage culture buffer G was added to terminate the digestion, the mixture was discarded by centrifugation for 5min, then the organoid passage culture buffer G was added and the mixture was resuspended and transferred to a 1.5mL centrifuge tube, and the liquid was discarded by centrifugation at 300g for 5min for step 4. (4) After the organoids were collected, stromal glue was added and resuspended. Each well of stromal glue was spread in A 24-well cell culture plate, and placed in an incubator for 10-15min to add 500uL human bladder cancer organoid medium A. 3. Organoids were frozen (1) The medium was removed by suction with a pipettes gun, and 1-2 ml of 4 ° C organoid passage culture buffer G was added to each well and placed for 2min. (2) The matrix glue was gently blown by a pipetting gun, collected in a 15mL centrifuge tube and allowed to rest for 10min at 4 ° C. (every 6-8 Wells were a group) (3) The supernatant was discarded by centrifugation for 5min, then an appropriate amount of organoid passage culture buffer G was added and resuspended again, and the liquid was discarded by centrifugation at 300g for 5min. (4) Add appropriate amount of organoid cryopreservation solution F, gently blow and resuspend, take 24-well cell culture plate as an example: the density is 2 Wells and freeze 1 tube, each tube volume is 1.4mL. (5) After making the labeling information and cooling, the samples were transferred to liquid nitrogen for long-term storage. 4, organoid resuscitation (1) 10mL organoid subculture buffer G was placed in a 15mL centrifuge tube. (2) The frozen organoid cells were removed from the liquid nitrogen tank and quickly thawed in a 37℃ water bath. (3) During the water bath melting process, it is necessary to gently shake the freezing tube to ensure that the frozen solution is completely melted within 1-2min. (4) The dissolved organoid cells were quickly transferred to a 15mL centrifuge tube, gently blown with a pipette for 6-8 times, centrifuged at 300g for 5min, and then the supernatant was removed and the organoid cell precipitate was collected. An appropriate amount of organoid passage culture buffer G was added and resuspended into a 1.5mL centrifuge tube and centrifuged at 300g for 5min. (5) Matrigel was resuspended, 25uL of Matrigel per well was spread in A 24-well cell culture plate, placed in an incubator for 10-15min to gel, and 500uL of human bladder cancer organoid culture medium A was added. |
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| Storage Temp. | Stored at 4 ° C, valid for 3 months; Stored at -20 ° C, valid for 1 year, see label for details. |
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4.6 ★★★★★
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Product Reviews
★★★★★ 5
Better than the others
Size: Large
I was in search of a ball that would hold up after the brand I’d bought for years suddenly lasted less than a month. This one is almost 3 months in use. Some damage done after playing with it almost non-stop, but still going.
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Reviewed in the United States on June 15, 2024
★★★★★ 5
Aggressive chewers have met their match.
Size: 10.5 inch Rubber Flex Tread
We purchased this toy in 2022, it finally broke last night. Four years of rough play with 2 boxers who are aggressive chewers. They would tug together, gnaw on it for hours, and often leave it outside exposed to the elements, but this toy just kept going. It was by far their favorite toy, it kept them busy for hours. I just ordered a replacement. Well worth the money, perfect size. Highly recommend.
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Reviewed in the United States on April 1, 2026
★★★★★ 5
Durable, fun, and safe for my dog
Size: 10.5 inch Rubber Flex Tread
I have a 10 month old, 50 lb mutt who has the musculature of a pit bull and the muzzle of a GSD. Buttercup’s favorite past-time is shredding toys into teeny tiny bits, but sometimes those bits don’t come back out of her mouth. She can’t have tennis balls, stuffies, anything with a squeaker, silicone toys, plastic toys, rubber toys with spikes or small appendages, anything with rope, or most chews & bones. She chewed her first and only $8+ XL bully stick like a corn chip. Finding indoor enrichment is a challenge. Buttercup is extremely pleased with this tire tug thingy, it is very ergonomic for both her and me to get a handle on it, it has more movement than any other toy she can safely have, and it’s very durable. After more than a week, there’s only a little bit of tread missing, which is definitely the best-case outcome with a super chewer. I think this is a good alternative to GoNut toys because it’s solid & dense while also a little bit easier & more interesting to play with, and very unlike GoNuts, this Tonka toy is very reasonably priced. I also think this it’s a good size for almost any breed of dog—I could definitely picture a little Boston or Frenchie dangling from one end. Highly recommended!
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Reviewed in the United States on December 20, 2024
★★★★★ 5
Dog loves it
Size: 10.5 inch Rubber Flex Tread
I would buy this again and again. It’s extremely durable and has held up great with my 9 month old Saint Bernard puppy. We even call it his “pacifier” because every night he grabs it and chews on it until he falls asleep. Definitely a favorite in our house!
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Reviewed in the United States on March 20, 2026
★★★★★ 4
Tough toy, but the yellow tire broke at the seam
Size: 10.5 inch Rubber Flex Tread
It was fun while it lasted and it got a lot of use, but it finally gave out after 6+/- months? Great tug toy. My dog is a 70# mutt with a strong bite and he's chewed through pretty much every other toy he's ever had. This one lasted longer than any of the others by quite a bit. He did zero chew damage to the tires, but the yellow tire in the middle broke at the seam. No tear, the seam just came undone at 90 degree angles. I'm going to look for something else this time but this wasn't bad considering how long it lasted and it wasn't very expensive. If I don't find anything tougher, I may purchase another one of these down the road.
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Reviewed in the United States on March 18, 2024
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