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Description
Human PINP ELISA KitProduct Specification Usage Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High precision pipette and gun tips: 0. 5 10uL, 5 50uL, 20 200uL, 200 1000uL 3. 37 constant temperature box 4. Distilled water or deionized water Sample processing and requirements: 1. Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4 overnight, then centrifuge at 1000g for 20
Product Specification
| Usage |
Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High-precision pipette and gun tips: 0.5-10uL, 5-50uL, 20-200uL, 200-1000uL 3. 37℃ constant temperature box 4. Distilled water or deionized water Sample processing and requirements: 1. Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4℃ overnight, then centrifuge at 1000×g for 20 minutes, and take the supernatant, or store the supernatant at -20℃ or -80℃, but avoid repeated freezing and thawing. 2. Plasma: Collect the specimen using EDTA or heparin as an anticoagulant. Centrifuge the specimen at 1000 × g for 15 minutes at 2-8°C within 30 minutes of collection. The supernatant can be assayed or stored at -20°C or -80°C, but avoid repeated freezing and thawing. 3. Tissue homogenization: Rinse the tissue with pre-chilled PBS (0.01M, pH 7.4) to remove residual blood (lysed red blood cells in the homogenate will affect the measurement results). Weigh the tissue and mince it. Add the minced tissue to the appropriate volume of PBS (generally a 1:9 weight-to-volume ratio, e.g., 1 g of tissue sample to 9 mL of PBS. The specific volume can be adjusted according to experimental needs and recorded. It is recommended to add protease inhibitors to the PBS) in a glass homogenizer and grind thoroughly on ice. To further lyse tissue cells, the homogenate can be sonicated or repeatedly frozen and thawed. Finally, centrifuge the homogenate at 5000 × g for 5-10 minutes, and the supernatant can be assayed. 4. Cell Lysis Buffer: Gently wash adherent cells with ice-cold PBS, then trypsinize and collect cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be collected directly by centrifugation. Wash collected cells three times with ice-cold PBS and resuspend in 150-200 μL of PBS per 1×10^6 cells (it is recommended to add protease inhibitors to the PBS; if the cell count is very low, reduce the PBS volume appropriately). Disrupt the cells by repeated freeze-thaw cycles or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, and remove the supernatant for analysis. 5. Cell Culture Supernatant: Centrifuge at 1000×g for 20 minutes. Remove the supernatant for analysis or store at -20°C or -80°C, avoiding repeated freeze-thaw cycles. 6. Other biological fluids: Centrifuge at 1000xg for 20 minutes, remove the supernatant, and test. Pre-test preparation: 1. Remove the test kit from the refrigerator 10 minutes in advance and equilibrate to room temperature. 2. Prepare the standard gradient working solution: Add 1 mL of universal diluent to the lyophilized standard, let it stand for 15 minutes to completely dissolve, then gently mix (concentration is 10 ng/mL). Then dilute to the following concentrations: 10 ng/mL, 5 ng/mL, 2.5 ng/mL, 1.25 ng/mL, 0.625 ng/mL, 0.3125 ng/mL, 0.15625 ng/mL, and 0 ng/mL. Serial dilution method: Take seven EP tubes and add 500uL of universal diluent to each. Pipette 500uL of the 10ng/mL standard working solution into the first EP tube and mix thoroughly to make a 5ng/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube serves as a blank well; there is no need to pipette liquid from the penultimate tube. See the figure below for details. 3. Preparation of biotinylated detection antibody working solution: Centrifuge the concentrated biotinylated antibody at 1000×g for 1 minute 15 minutes before use. Dilute the 100× concentrated biotinylated antibody to a 1× working concentration with universal diluent (e.g., 10uL concentrate + 990uL universal diluent). Prepare and use immediately. 4. Prepare the enzyme conjugate working solution: 15 minutes before use, centrifuge the 100× concentrated enzyme conjugate at 1000×g for 1 minute. Dilute the 100× concentrated HRP enzyme conjugate to a 1× working concentration with universal diluent (e.g., 10 μL of concentrate + 990 μL of universal diluent). Prepare immediately. 5. Prepare the 1× wash solution: Dispense 10 mL of 20× wash solution into 190 mL of distilled water (concentrated wash solution removed from the refrigerator may crystallize; this is normal. Allow to stand at room temperature until the crystals have completely dissolved before preparing). Procedure: 1. Remove the desired strips from the aluminum foil bag after equilibration at room temperature for 10 minutes. Seal the remaining strips in a ziplock bag and return to 4°C. 2. Sample addition: Add 100 μL of sample or standard of varying concentrations to the corresponding wells. Add 100 μL of universal diluent to the blank wells. Cover with a film and incubate at 37°C for 60 minutes. (Recommendation: Dilute the sample to be tested at least 1-fold with universal diluent before adding it to the ELISA plate. This will reduce the impact of matrix effects on the test results. The sample concentration should be multiplied by the corresponding dilution factor when calculating the final sample concentration. It is recommended to run replicates for all test samples and standards.) 3. Add Biotinylated Antibody: Remove the ELISA plate and discard the liquid without washing. Add 100 μL of Biotinylated Antibody Working Solution directly to each well. Cover with a film and incubate at 37°C for 60 minutes. 4. Wash: Discard the liquid and add 300 μL of 1x Wash Solution to each well. Let stand for 1 minute, shake off the wash solution, and pat dry on absorbent paper. Repeat this process three times (a plate washer can also be used). 5. Add Enzyme Conjugate Working Solution: Add 100 μL of Enzyme Conjugate Working Solution to each well. Cover with a film and incubate at 37°C for 30 minutes. 6. Washing: Discard the liquid and wash the plate five times as in step 4. 7. Adding substrate: Add 90 μL of substrate (TMB) to each well, cover with a sealing film, and incubate at 37°C in the dark for 15 minutes. 8. Adding stop solution: Remove the ELISA plate and add 50 μL of stop solution directly to each well. Immediately measure the OD value of each well at a wavelength of 450 nm. Calculating experimental results: 1. Calculate the average OD value of the standard and sample replicates and subtract the OD value of the blank well as a correction factor. Plot the standard curve of the four-parameter logistic function on double-logarithmic graph paper, with concentration as the horizontal axis and OD value as the vertical axis. 2. If the sample OD value is higher than the upper limit of the standard curve, dilute the sample appropriately and retest. Multiply the sample concentration by the corresponding dilution factor. |
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| Theory | This kit uses a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). Sample, standard, biotin-labeled detection antibody, and HRP conjugate are sequentially added to microwells pre-coated with a capture antibody against procollagen I N-terminal propeptide (PINP). After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by peroxidase (HRP) catalysis and to yellow by acid. The intensity of the color is positively correlated with the amount of procollagen I N-terminal propeptide (PINP) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Procollagen I N-Terminal Propeptide ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Peptides are cleaved from procollagen by proteases to form collagen. The concentrations of these two peptides in the bloodstream reflect the rate of type I collagen synthesis. Procollagen type 1 N-terminal peptide (PINP) is considered the most sensitive biomarker of bone formation. PINP exhibits diurnal variations, with higher values at night. PINP is metabolized in the liver. | |||||||||||||||||||||||||||||||||
| General Notes | 1. Strictly adhere to the specified incubation time and temperature to ensure accurate results. All reagents must be at room temperature (20-25°C) before use. Refrigerate reagents immediately after use. 2. Improper plate washing may result in inaccurate results. Ensure that all liquid in the wells is aspirated thoroughly before adding substrate. Do not allow the wells to dry out during incubation. 3. Remove any residual liquid and fingerprints from the bottom of the plate, as this will affect the OD value. 4. The substrate developer solution should be colorless or very light in color. Do not use substrate solution that has turned blue. 5. Avoid cross-contamination of reagents and specimens to prevent erroneous results. 6. Avoid direct exposure to strong light during storage and incubation. 7. Do not expose any reagents to bleaching solvents or the strong fumes emitted by bleaching solvents. Any bleaching agent will destroy the biological activity of the reagents in the kit. 8. Do not use expired products, and do not mix components with different product numbers and batches. 9. Recombinant proteins from sources other than the kit may not be compatible with the antibodies in this kit and will not be recognized. 10. If there is a possibility of disease transmission, all samples should be managed properly and samples and testing devices should be handled according to prescribed procedures. |
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| Storage Temp. | If the unopened kit is stored at 4°C, the shelf life is 6 months. | |||||||||||||||||||||||||||||||||
| Test Range | 0.15-10 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma, tissue homogenate, cell lysate, cell culture supernatant and other biological fluids |
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4.3 ★★★★★
Based on 8 reviews
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Product Reviews
★★★★★ 4
LOVE this brand. Have many of their styles. This is basically an unstuffed toy.
Size: Large, Color: Dinos Frills (Gray)
I have about 10 -12 different styles of the GoDog brand toys. It is like Xmas when my Malinois dogs get a new GoDog toy. Have loved this brand for many years and have some toys which have lasted 5-6 years -- minor "surgeries" stitch they back together -- until I retired them. They have never just worn out. Just to be clear:
THESE TOYS ARE NOT CHEW PROOF. NO STUFFED TOY IS CHEW PROOF.
However, this brand of stuffed toy is more chew proof than other brands. But if you are looking for a chew proof stuffed toy be advised -- IT DOES NOT EXIST!! No fabric is chew proof. Even if they invented such a fabric I would not want to ruin my dog's teeth by letting them try to chew through it. Protecting one's dog's teeth should be the highest priority.
If a dog can chew through a PLASTIC KENNEL and even METAL WIRE KENNELS (yes, it happens) then they can easily chew through a puffy, fluffy stuffed toy. The "Chew Guard Technology" language may be misleading to some customers who are disappointed in the brand. I defend the brand and have used it for many years. No, I don't get paid or get free toys.
Since I have had many styles within this brand and even have the pink version of this style I thought I knew what I was getting. This is basically a toy without stuffing which some people prefer. For whatever reasons my dogs prefer toys that are stuffed. Really stuffed with a firm density that they can sink their teeth into. So this toy will be OK. I can tell it will not be one of their beloved toys like some of the brand's other styles such as the purple brontosaurus. My fault for not reading more closely and assuming the grey version would be exactly like the pink version which is fully stuffed. If they get bored with these two I keep some of the Chew Guard toys stock piled.
BTW one can wash and dry these toys. Yes, eventually the squeakers suck up some water and die but one can replace the squeakers. Also on Amazon. So I am willing to replace the squeakers a few times in the life time of one of these toys to be able to wash and dry them occasionally. Usually takes a couple drying cycles to get the heavily stuffed toys dried completely.
If you have an aggressive chewer you may wish to look into Goughnuts dog toys or other hard rubber toys that are designed for aggressive chewers. Even then, with a determined dog, the Goughnuts toys can be chewed through. I think they offer a replacement guarantee. The other brand which has some toys of hard rubber is West Paw. An excellent brand that also offers a one time replacement. Bully Sticks are another option for chewers.
PLEASE, PLEASE, PLEASE stay away from the hard plastic chew toys like the Nyla Bones. Hard plastic toys will destroy your dog's teeth over time. I learned the hard way (no pun intended) and ruined one of my dog's teeth. Vet asked if I allowed my dog to chew ROCKS !!!!!! Every tooth was cracked or fractured. Not wishing to pull teeth I had to pay for some very expensive root canals and caps. Vet's advise: If you can't dent the toy or chew with your finger nail DO NOT offer to your dog. Good advise to follow. Even some of the Bully Sticks (Costco Brand) are so hard they can chip or fracture teeth.
Best for you and your dog.
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Reviewed in the United States on September 1, 2019
★★★★★ 5
Well made, minor issue was quickly resolved
Size: Large, Color: Dinos Bruto (Purple)
I purchased the Dino Bruto Dog toy for my labradoodle who is quite fond of squeaky, squawky stuffed toys and oddly enough has shown a preference for purple. I opened the package with my hound standing by waiting for his gift. I detached the tags and attempted to squeak the toy. It seemed difficult to squeak or honk. I tossed the purple Dino Bruto to my dog who excitedly chomped to squeak. The toy squawked a couple times and then just quit. My poor hound was staring up at me with that "what's up with this" look on his face. I tracked down the website for the GoDog toys, not going through the Amazon return process. I called the good people at GoDog and stated simply that my dog's favorite part of the toy failed quite quickly and I was wondering about guarantees. I was told to go to the website and file a replacement request. The replacement request required that I include a photo which I did as directed. I received an reply email within a couple hours which stated that a new toy would be sent out shortly.
I am happy to report that we received a replacement toy, not the purple Dino, but a colorful wood duck. The squawker device works well and is still performing like new more than a month later.
It appears that the squeaker is not those little plastic squeakers but the bigger heavy plastic bag, bladder, with what I visualize as a duck call sort of sound device. I am guessing because I haven't seen the inside of the GoDog toys. I have seen the guts of other stuffed toys which use this type of noise device. While our dog seems to enjoy the sound of the plastic bag, bladder set-up squawkers, they seem to fail quickly and I tend to avoid them because of this. The toys using this type of squawker that I have seen, use what looks like a cheap lightweight plastic bag which is not securely attached to the "voice box".
Since the GoDog replacement toy is holding up so well, I am guessing GoDog uses a heavier grade plastic bladder coupled with reinforced construction. The purple Dino Bruto toy still looks new even after multiple times of our hound grabbing the purple toy, shaking it at us with his invite to give chase or play keep away. Since the squawker failed so quickly with the new toy, I figure it was just a defective item which does not reflect the general quality of toys from GoDog.
One evening our neighbor was visiting while his pitbull was playing in our yard with our dog. Our patio door was open so our dog ran into the house and grabbed his purple Dino Bruto toy. He is a bit of a show-off with new toys. Our dog then went racing back into the yard, shaking his GoDog toy at the pitbull. A quick game of tug-of-war ensued. Since this is a new toy, our neighbor leaped into the game and took the toy away from the dogs. In that short time of tugging between the dogs, the toy held together and did not even pull apart at any of the seams. There were no puncture marks either.
This is a game our dog has initiated before with the neighbor pitbull. Normally, the stuffed toys are ripped apart almost immediately. I was surprised and impressed with the quality of the sewing which appears to employ some sort of reinforced construction or heavier material.
I plan to make more purchases from GoDog toys. The toys are a little more expensive than some stuffed dogs toys but they hold up so well I figure they are worth a few more dollars. They are great toys that hold up well and the company provides quick responses to inquiries. I consider this a winning combination.
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Reviewed in the United States on June 15, 2013
★★★★★ 5
Good quality
Size: Large, Color: Dinos Frills (Gray)
we’ve had many godog toys over the years and although my dog eventually rips through them I do find that they last longer than other cheap toys. Overall very well made!
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Reviewed in the United States on May 11, 2026
★★★★★ 5
Cute and fairly long lasting
Size: Large, Color: Dinos Frills (Red)
Lasted from Nov 13th, 2024-August 20th, 2025 lol very impressive. She played with it nearly daily and loved the big squeak. The squeak box was just ripped out so she probably won’t play with it anymore. Happy to order another!
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Reviewed in the United States on August 21, 2025
★★★★★ 4
He was OBSESSED!!!
Size: Large, Color: Dinos Frills (Gray), Size: Large, Color: Dinos Frills (Gray)
Our year and a half old lab was excited the moment he saw it come out of the box ! He had many toys that he tore into shreds within a day so this was his first “heavy toy “ that wasn’t easy to tear .
I think he loves the design and feel of it , as he carried this thing EVERYWHERE and even to bed . Him and his sister share toys but this was his , he claimed it lol
Now if it wasn’t for us telling him to stop trying to kill the Dino he could have easily tore into this thing within a couple of days . However since he started chewing gently he became obsessed with the squeaker , and used that as an alarm that he was up every morning in bed 😂
The quality of the product is actually pretty good but it’s not bullet proof, and our smart dog was able to sneakily tear a whole in the nose and some how pull out the stuffing .
All in all I would definitely buy another toy from this company because they are made with quality for dogs , and lasted about three weeks longer than his average toys ! It was his favorite toy 💕
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Reviewed in the United States on September 3, 2023