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Description
Rat NSMASE ELISA KitProduct Specification Usage Required experimental equipment: 1. Microplate reader (450nm) 2. High precision pipettes and pipette tips: 0. 5 10uL, 5 50uL, 20 200uL, 200 1000uL 3. 37C incubator 4. Distilled or deionized water Sample preparation and requirements: 1. 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
Product Specification
| Usage | Required experimental equipment: 1. Microplate reader (450nm) 2. High-precision pipettes and pipette tips: 0.5-10uL, 5-50uL, 20-200uL, 200-1000uL 3. 37°C incubator 4. Distilled or deionized water Sample preparation and requirements: 1. 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 and mince the tissue. Add the minced tissue to the appropriate volume of PBS (generally a 1:9 weight-to-volume ratio, e.g., 1g of tissue sample to 9mL 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 collect the supernatant for analysis. 2. Cell Lysis Buffer: Adherent cells should be gently washed with pre-chilled PBS, then trypsinized and harvested by centrifugation at 1000×g for 5 minutes. Suspension cells can be harvested directly by centrifugation. Collected cells should be washed three times with pre-chilled PBS and resuspended 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 freezing and thawing or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, and collect the supernatant for analysis. 3. 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 sphingomyelin phosphodiesterase 2 (NSMASE) capture antibody. After incubation and washing, the assay is developed using the substrate TMB. TMB converts to blue under the catalysis of HRP and to yellow under the action of acid. The intensity of the color is positively correlated with the amount of sphingomyelin phosphodiesterase 2 (NSMASE) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Sphingomyelin phosphodiesterase 2 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Neutral sphingomyelinase (NSMASE), also known as sphingomyelin phosphodiesterase 2 (SMPD2), is an enzyme encoded by the SMPD2 gene. This gene encodes a protein that was initially identified as a sphingomyelinase based on sequence similarity between bacterial sphingomyelinases and yeast proteins. Subsequent studies have shown that its biological function is unlikely to be a sphingomyelinase, but rather a lysophospholipase. | |||||||||||||||||||||||||||||||||
| 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 | 0.156-10 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates, cell lysates, and other biological fluids |
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4.2 ★★★★★
Based on 12 reviews
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Product Reviews
★★★★★ 5
Outstanding Indie Fantasy! You NEED to read this!
I have started and deleted my intro to my review multiple times at this point. My mind is sufficiently BLOWN. This book has landed itself into my top favorite romantasies right alongside ACOTAR and FBAA. ITS SO DANG GOOD. I literally woke up, popped it open on my kindle app (Thanks author and R&R Book Tours for the arc!) and got to reading and just didn’t stop. I couldn’t stop. From the first dang page, Menard dragged my butt in and tied me down. I. Was. HOOKED.
Writing Style: I absolutely love the writer’s style! Told from mainly Milla’s POV, with the occasional glance into Nico’s POV when its important. Her world building is rich and immersive, sticking firmly into the steampunk/gas lamp fantasy atmosphere. Her magic system—while feeling familiar—still feels unique and blends beautifully with the world she’s created.
Characters: I can’t begin to describe how much I love the characters Menard has created for this story. Milla is a firestorm. She’s passionate, fiesty, loyal, and self confident. She doesn’t back down, no matter the odds. For me, personally, she’s ranking right up there with Feyre (ACOTAR), Poppy (FBAA), and Sera (FBAA prequel) as one of my favorite female leads. Despite her fierceness, she’s not invincible and she’s still vulnerable—which unveils itself more as the story goes on.
Nico, mmmmmmmm, Nico. He’s cocky, and strong. His heart is for his blood and family. There’s nothing he won’t do for those he cares for. He’s incredibly insightful which leads to a lot of me squeeing and highlighting his quotes. Pairing him up with Milla is a riot I will watch again and again.
The side characters are all phenomenal, ranging wide in personalities and giving the world a richer vibrancy. They compliment both main characters, bringing out the best and worst in them, helping to bring secrets to light and keep the plot going. Not a single character felt out of place, or ever out of character.
Relationship: Y’all know I love a good slow burn and this burns so good. The push and pull of their hate/love relationship is a thrill to witness. Watching them fall for each other is a joy and the spice (when we get there) is HAWT.
Plot: This story had me hooked, and that’s largely in thanks to how well planned out the story’s plot is. Menard did an excellent job, masterfully balancing her foreshadowing and reveals. While I halfway guessed the big reveal, i definitely did not expect the big reveal. (Read the book, then you’ll understand lmao). The pacing is fantastic. Nothing about the story felt rushed, not the relationship, not the reveals, not the over arching plot line. Just absolutely phenomenal.
Enjoyment: 100/10 will read again. I need a physical copy in my life. I HIGHLY recommend this book.
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Reviewed in the United States on October 28, 2023
★★★★★ 5
One of the best reading experiences I’ve ever had
Format: Audiobook
This was truly AMAZING!!!!!!!!!!!! In every possible way.
- the setting. I loved the steampunk/gaslamp setting with trains, overcast weather, and mafia vibes
- the magic. I still don’t entirely understand the magic system and descendants as much and it took awhile for me to get my footing in that department- and I am intrigued beyond measure, though it in no way lessened my enjoyment of the story. I just really wish I had a better grasp of it.
- the plot itself. Beautiful. The tropes were troping in the absolute best way. Every single thing that happened I was like YES MORE!!!
- the writing. I found myself grinning ear to ear uncontrollably during my entire read through and full on choking on my gasps, again, like the entire read through. And I read this too at times when I was in an awful mood so the way this book was an INSTANT pick me up really speaks to how well this was written. I was having the absolute best time, non stop.
- the characters. The best part oh my gosh. Camilla is SUCH a badass. Her and Nico are supposed to be low 20s and I gotta tell you they read more like 30s from how capable and mature they are (not that that age bracket is incapable or immature, just comparing them to other characters written in that age range). She is the type of cool as heck character that everyone would be cos-playing as and if it was a kid movie, every single girl would dress as her for Halloween. She’s that awesome.
Nico is top freaking tier. I gotta say his hairstyle is my undoing and with the addition of the metal arm- before I even started reading the book I was in love with him. And he was EVEN BETTER than my lofty expectations. This man. Perfect in every single way. Every. Single. Way. If I start now I won’t stop so I’m just gonna leave it at that. Camilla and Nico are power couple of the century though and the way they interacted was the most toe-curling/squealing/kicking your feet banter I’ve read in a while. PLUS the way they fell for each other was a gem. I was falling in love with their relationship right along with them.
- a shout out to the side characters too. Aramis, Gideon, Aunt Fran, Esme, Luther, Adler [honestly just insert all the supporting cast they were freaking phenomenal] brought so much life and personality to the story and helped round out Nico and Camilla. I really really really would absolutely adore if Adler got some more spotlight time in the future because he and Aramis are up there with “let’s give these side characters their own spinoffs”.
Also, the villains were great too. I love a great villain in a story and sometimes I don’t think they get highlighted as much in books as in movies or TV that I hear of.
If it wasn’t obvious already- this was even greater than a 5 star read. This was perfection- in quality and in vibes- and it was one of the most enjoyable reading experiences I have ever had and I’m not throwing that around lightly. I have scenes from this book playing in my mind unbidden on repeat and I know this world has so much more to be explored. I will read anything Alexis L Menard puts out in the future
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Reviewed in the United States on March 5, 2025
★★★★★ 4
Great mix of maffia and magic
Format: Kindle
This one felt really unique to me, with a great and successful blend of Twenties-feeling maffia and inventive magic. I liked both the main characters a lot, even though I found myself almost shouting at the MFC sometimes. She really should learn not to trust everyone just because they're family!
All in all a good read, with lots of romance, yearning and some spice as well.
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Reviewed in the United States on November 15, 2025
★★★★★ 3
So close to being good
I really wanted to like this book. It had all the makings of a strong, interesting story, but the plot got so convoluted that it was hard to follow. Also, for a "badass" FMC, Milla was amazingly inept. The writing got really stilted at times too. I will probably still read the next one anyway, because I'm curious to see where the plot goes.
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Reviewed in the United States on May 3, 2024
★★★★★ 5
FASCINATING world, great characters and swoony slow burn!
House of Bane and Blood by Alexis Menard
-Order and Chaos Book 1-
4.5/5⭐️
1/3🌶️*
2/3🦋
•My Thoughts•
What a RIDE! This world was absolutely fascinating with its 1920s mafia vibes meets magic. The characters and enemies to lovers storyline hooked me from the beginning - add in a marriage of convenience and a dash of mystery and I ate👏🏻it👏🏻up👏🏻. Milla’s growth was so good and I was cheering for her as she became more confident! I also loved Niko and his back story and resilience. These are imperfect characters you find easy to love, root for and I became enamored with their slow but steadfast (if reluctant) support of each other. I did find the government structure a little confusing but didn’t want to slow down enough stop and figure it out (if anyone reads it an wants to make a chart, please share it with me). This book was violent but also mesmerizing and full of hope for a better future. I loved the big family dynamics and the never ending plot twists that kept me guessing. This is a completed duology and with that ending, you BET I’m immediately reading book 2.
*I rated this a 1/3🌶️ because there were only a few scenes with on page intimacy. However, I would like to note that the spice involves kink: breath play.
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Reviewed in the United States on June 10, 2024
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