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Description
Human RAB7A 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: 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
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: 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. 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. 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 20 ng/mL). Then dilute to the following concentrations: 20 ng/mL, 10 ng/mL, 5 ng/mL, 2.5 ng/mL, 1.25 ng/mL, 0.625 ng/mL, 0.3125 ng/mL, and 0 ng/mL. Serial dilution method: Take 7 EP tubes and add 500 μL of universal diluent to each tube. Pipette 500 μL of the 20 ng/mL standard working solution into the first EP tube and mix thoroughly to make a 10 ng/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube serves directly as a blank well; there is no need to aspirate the liquid from the penultimate tube. See the figure below for details. 3. Preparation of Biotinylated Antibody Working Solution: 15 minutes before use, centrifuge the concentrated biotinylated antibody at 1000×g for 1 minute. Dilute the 100× concentrated biotinylated antibody to a 1× working concentration using universal diluent (e.g., 10µL concentrate + 990µL universal diluent). Prepare immediately before use. 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 the Ras-related protein Rab-7a (RAB7A). 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 Ras-related protein Rab-7a (RAB7A) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Ras-related protein Rab-7a ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | RAB7A, also known as RAB7A, a member of the RAS oncogene family, is a protein encoded by the RAB7A gene. It is involved in endocytosis, the process of bringing substances into cells. Endocytosis involves the folding of the cell membrane around extracellular material, forming a vesicle. The vesicle is then brought into the cell and cleaved from the cell membrane. It plays an important role in vesicle entry and trafficking. It has been shown to play a fundamental role in the cellular vacuolation induced by the Helicobacter pylori cytotoxin VacA. | |||||||||||||||||||||||||||||||||
| 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 | 0.312-20 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates, cell lysates, and other biological fluids |
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4.4 ★★★★★
Based on 435 reviews
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Product Reviews
★★★★★ 5
Impressed so far
Style: Standard Grip, Size: 25ft, Color: Chartreuse
I have a large back yard, a pool and a stationary hose reel bolted to the house that holds around 250 feet of hose (depending on the hose). During the summer especially, we use this hose a lot. The reel had 200 feet of pretty good quality, heavy duty residential hose on it. That hose lasted over 20 years but was starting to show it's age and had been repaired several times. What's more, my wife hated the stuff. Her primary complaint was how heavy it was. She really struggled to drag it around the yard or wind it back on the reel. So it was time to replace it. My criteria: brass fittings, reasonably heavy duty and durable, decent price. Her criteria, lighter! So I searched the reviews and looked at damn near everything amazon sells and picked this stuff. I ended up buying two 50 foot hoses and one 100 foot hose.
So, here's what I think. The hose ends are heavy duty brass and well crimped. The little bits of plastic sleeve on each end are just branding and don't provide any other useful purpose. This isn't a criticism, just an observation. I cut them off. As others have mentioned, the female fittings are crimped a little tight. This makes it somewhat difficult to tighten them without twisting the hose. I put a little white lithium grease on mine and held the things in my hand - twisting back and forth until they lubed up and loosened up. No harm, no foul, I'd rather a little tight than loose. Once that was done, I connected all the sections and stretched the whole thing out in the yard. I connected one end to the faucet (hose reel), slapped a nozzle on the other end and turned on the water. It was a warm summer day, I let the hose fill with water and then went down the entire 200 foot length untwisting and straightening it all out. Then I left it bake in the sun for a couple hours under pressure. After that I turned off the water, and started inspecting the hose. I have about 80lbs of water pressure, left under pressure and cooking in the sun, the hose definitely swelled a little diameter wise, but no bulges or anything untoward. Connections all looked good, no leaks or malfunctions. So, I turned off the water and took off the nozzle and started winding it up on the reel. Now right away I could tell that this stuff was more pliable and significantly lighter then the hose it replaced. I carefully spooled it up on the reel and threaded a brass shutoff and brass quick connector on the end. That was the middle of June, 2023. It's now December 2023.
The verdict. I like the stuff, and more importantly, so does my wife. From my perspective, I like the color, it's easy to see and find in the grass, I don't trip over it or worry about not seeing it before I run over it while cutting the grass (you can laugh, but that accounted for one of the previous "repairs" I mentioned earlier). Now, when you pull hose off a reel, it tends to come off pretty straight, which automatically alleviates a bunch of twisting and kinking issues. Still, as you drag this stuff around the yard it keeps its shape pretty well and doesn't twist up or kink. It's flexible and yet "stiff" enough to drag around corners without issue. The hose itself has definitely faded a little colorwise, the end that gets the most use is less orange then the hose that spends most of its time wrapped on the reel, but not unduly so. After a summer of use, I'd have to say it's held up well, without any signs of terminal wear and tear. What does my wife think? She loves the stuff. It's much easier for her to use and wind back up. She really is the primary user, she does 90% of the pool "stuff", plant tending, gardening and watering. If she likes it, I'm good. So, we'll see, in a couple more years, if this hose doesn't hold up, I'll come back and let you know. For now I'd have to say, if you need a new hose, try this stuff.
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Reviewed in the United States on October 12, 2024
★★★★★ 5
Great hose!
Style: Standard Grip, Size: 100ft, Color: Chartreuse
5 stars without hesitation. This Flexzilla Garden Hose 5/8 in. x 100 ft. is legitimately one of the best hoses I’ve used. It’s actually flexible like advertised, doesn’t fight you every time you move it around, and it feels way more durable than the cheap stiff hoses from big box stores. I honestly don’t understand a lot of the complaints, mine has held up great, doesn’t kink nearly as bad as others I’ve owned, and is super easy to coil up. Solid hose.
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Reviewed in the United States on May 10, 2026
★★★★★ 5
If you want to buy your last hose then flexzilla is your hose.
Style: Standard Grip, Size: 50ft, Color: Chartreuse
O.K. if you are reading my review I can honestly say that I don't think anyone knows what they are buying with this hose like I do as Amazon has the record of when I bought 100 feet of this hose. Long Long Long term user=purchased in 2017. I used this hose today and it looks-feels-works like it did the day I hooked it up on my reel. So flexible you can use it for a tourniquet if required. Hot-cold it don't matter this hose is the best. I am 70 yrs. old and it is the best hose I ever used in my life. Buy this hose once and then be done. Never kinks and not to heavy when moving around. Buy some and get on with whatever work you need to wash and then roll it up and forget about it.
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Reviewed in the United States on May 22, 2026
★★★★★ 4
Hose kinks.
Style: Standard Grip, Size: 100ft, Color: Chartreuse
I bought this hose 2 months ago and really like how light it is and easy to pull around the yard. However, I am having a problem with it kinking all of the time. It is really frustrating to have to go back and straighten it out when I am watering.
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Reviewed in the United States on April 19, 2026
★★★★★ 5
Excellent product.
Style: Standard Grip, Size: 50ft, Color: Chartreuse
I've dealt with many different brands of hoses, took into account quality and price. After many disappointing purchases I can finally say "I got the right one this time". It is exactly as advertised, lightweight, flexible and very easy to handle. You won't go wrong with Flexzilla garden hose!!!
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Reviewed in the United States on May 24, 2026