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Description
Rat N-MID-OT 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: 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: 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. 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. 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. 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. 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. 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. Preparation of standard gradient working solution: Add 1mL of universal diluent to the lyophilized standard, let it stand for 15 minutes to completely dissolve, then gently mix (concentration is 4000pg/mL). Then dilute to the following concentrations: 4000pg/mL, 2000pg/mL, 1000pg/mL, 500pg/mL, 250pg/mL, 125pg/mL, 62.5pg/mL, and 0pg/mL. Serial dilution method: Take seven EP tubes and add 500uL of universal diluent to each tube. Pipette 500uL of the 4000pg/mL standard working solution into the first EP tube and mix thoroughly to make a 2000pg/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube is used 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 utilizes 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 N-terminal mid-segment osteocalcin (N-MID-OT). After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by HRP peroxidase and to yellow by acid. The intensity of the color is positively correlated with the amount of N-terminal mid-segment osteocalcin (N-MID-OT) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat N-terminal mid-segment osteocalcin ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Osteocalcin, also known as bone R-hydroxyglutamic acid protein (GLa protein), is a 49-amino acid protein. It is a non-collagenous acidic glycoprotein and a vitamin K-dependent calcium-binding protein. It is primarily synthesized by osteoblasts and odontoblasts, with some synthesis by proliferating chondrocytes. In its uncarboxylated form, osteocalcin acts as a hormone in the body, sending signals to the pancreas, fat, muscle, testicles, and brain, playing a key role in regulating bone calcium metabolism. | |||||||||||||||||||||||||||||||||
| 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 | 62.5-4000pg/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma, tissue homogenate, cell lysate, cell culture supernatant and other biological fluids |
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4.4 ★★★★★
Based on 1827 reviews
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Product Reviews
★★★★★ 5
You want this one, for power and quality and saving on AA battery costs
Color: Black
Yes. You want this one, for power and quality and saving on AA battery costs — buy it now. Don’t waste any more time comparing products.
After hours, days, weeks, of researching and comparing, I ordered this one. I do not regret it. It is powerful. It was less than $25 when I bought it. It has a USB charger port and it feels solid in your hand. It has a turn-knob at the top to control the variable speed immersion blender aka milk frother aka wand frother. Charge battery fully for 2hrs before first use and then you’re all set. It works very well. It functioned as expected and then even better. Yes. It exceeded expectations. I hope it lasts a long time. It feels like it is very well made and high quality. Classiest and best wand frother I have ever owned. And I’ve owned over 3-4 in the past 10yrs.
I got this one at $25 instead of the highest-rated one for $9.99 because those require AA batteries and this frother however is USB-C rechargeable!
So in 1-2 years it will have paid for itself in battery costs alone.
But there’s no price I can put on how enjoyable it is to use this super powerful, super smooth, high quality, wand frother.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on September 20, 2025
★★★★★ 5
Quality Design
Color: Gray
Excellent frother/mixer
Pros:
- Quality feels premium.
- Plenty of power to mix even with a drink packed with ice and peanut butter. Basically a mini handheld mixer.
- Ease of use is great. I appreciated that this had a longer brother so I can mix drinks in taller cups. Also the grip is fantastic and it is easy very easy to change the speed with just one hand so you can hold the drink with the other.
Longevity - So far it's been holding up and hasn't needed a battery replacement.
Cons:
None so far.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on December 10, 2025
★★★★★ 5
Highly recommend!
Color: Gray
Love this frother!!! Purchased to replace an Aerolatte that stopped working after a reasonable period of use. Right out of the box, it was charged and ready to go. I’ve been using it daily for about 5 weeks on the same charge it had on arrival and it’s still going strong. Love the variable speed and easy to work with one hand. And this baby can froth! It froths far better than our old Aerolatte. Also love the rechargeable feature so no more AA batteries wasted. This handy kitchen tool makes my morning coffee a true highlight. Highly recommend the Maestri milk frother!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on February 18, 2026
★★★★★ 5
Best Frother Ever!
Color: Black
Great little frother and very powerful after just one charge. I haven’t charged it since the first day and it is still going strong after 3 weeks. I use it daily for coffee froth, but have used it for mixing other things as well. It works great on mixing drink powders into water and for making salad dressing from scratch. It’s one of the best things I’ve purchased on Amazon for kitchen tools. My old frother ate through batteries and was 1/10th as powerful. This one really is amazing. Great quality heavy duty but still small machine. Extra attachment heads are also a bonus.
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Reviewed in the United States on May 23, 2026
★★★★★ 4
Very good frother that gets the job done extreamly well
Color: White
I hvae used this frother primarily for smoothies and coffees. Each time, it does a wonderful job.
Deducted a star for the location of the speed control which is ontop of the device. It's akward to use one handed while attempting to control the speed, athough it's advertised as making it easier. It's not!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on November 27, 2025
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