Rabbit Ubiquitin (Ub) Detection Kit

rabbit

6.25-400ng/mL

Each liquid component should be clear and transparent, without precipitation or floccules. The aluminum foil bag of the microplate should be vacuum packed without damage or leakage.

Used to detect the concentration of rabbit ubiquitin (Ub) in samples such as serum, plasma, and cell culture supernatant.

The kit uses enzyme-linked immunosorbent assay. Biotin-labeled rabbit ubiquitin (Ub) and purified anti-rabbit ubiquitin (Ub) antibodies are coated on microplates. In the competitive inhibition reaction, a certain amount of solid-phase antibody inhibits the competitive reaction with biotin-labeled rabbit ubiquitin (Ub) and non-labeled antigen (calibrator or specimen). The binding amount of antibody and biotin-labeled rabbit ubiquitin (Ub) is inhibited by the amount of non-labeled antigen. The more non-labeled antigen, the less binding of antibody and biotin-labeled rabbit ubiquitin (Ub), and vice versa. After the reaction is balanced, solid-phase antibody-biotinylated rabbit ubiquitin (Ub) is formed, and then enzyme-labeled avidin is added to form a solid-phase antibody-biotinylated rabbit ubiquitin (Ub)-enzyme label-avidin complex. After adding substrate for color development, the absorbance (OD value) is measured at a wavelength of 450nm using an enzyme reader. As the concentration of rabbit ubiquitin (Ub) increases, the OD value gradually decreases and shows a good linear relationship. This kit has the characteristics of high sensitivity, strong specificity, good repeatability, simple operation and rapidity, and has reliable detection performance for the reduction or increase of rabbit ubiquitin (Ub) in samples.

This kit recognizes native and recombinant rabbit ubiquitin (Ub) with no cross-reactivity to structural analogs.

The coefficient of variation within and between plates was<10%.

The recovery rate is85%-115%between.

1.Standard specification microplate reader. 2.Automatic plate washer. 3.Oscillator. 4.A series of adjustable pipettes and tips. When testing more samples at one time, it is best to use a multi-channel pipette.

1.Move all reagents to room temperature for half an hour to balance, take the concentrated washing solution, and use distilled water according to the number of tests in the batch.1:20Dilute, mix and set aside. 2.Take the pre-coated plate out of the sealed bag and set up a blank control well without adding any liquid; set up a blank control well for each calibrator.2Wells, add corresponding calibrants to each well50μl; Add the test serum or quality control product directly to each of the remaining test wells50μl. 3.Biotinylated antigen was added to all wells except blank wells.50μl, mix well, apply the sealing film, and place37Incubation60minute. 4.Manual plate washing: discard the liquid in the wells, fill each well with washing solution, and let it stand10Spin dry in seconds, repeat3Pat dry after washing. Wash the plate in a plate washer: Select Wash3Wash the plate once and pat dry. 5.Add enzyme-labeled avidin to each well50μl(Except blank control wells), mix well, apply sealing film, and place37Incubation30minute. 6.Manual plate washing: discard the liquid in the wells, fill each well with washing solution, and let it stand10Spin dry in seconds, repeat3Pat dry after washing. Wash the plate in a plate washer: Select Wash3Wash the plate once and pat dry. 7.Add colorimetric reagent to each wellA 50μl, color developerB 50μlAfter oscillation, place37℃Avoid light and color15Add stop solution to each well50μl. 8.Read the data with an enzyme-labeled instrument and take the wavelength450nm, first use the blank control well to adjust the zero point, and then measure the optical density value of each well (ODvalue).

1)This kit is for in vitro research use only and is not intended for clinical diagnosis. 2)Please wear lab coats and latex gloves for protection during the test. Especially when testing blood or other body fluid samples, please follow the national biological laboratory safety protection regulations. 3)Strictly follow the specified time and temperature to incubate to ensure accurate results. All reagents must reach room temperature before use. 20-25℃. Refrigerate the reagent immediately after use. 4)Improper plate washing can lead to inaccurate results. Be sure to aspirate as much liquid as possible before adding substrate. Do not allow the wells to dry out during incubation. 5)Eliminate residual liquid and fingerprints on the bottom of the board, otherwise it will affect ODvalue. 6)The substrate developer solution should be colorless or very light in color. 7)Avoid cross contamination of reagents and specimens to avoid erroneous results. 8)Avoid direct sunlight during storage and incubation. 9)The sealed bag was opened after equilibration to room temperature to prevent water droplets from condensing on the cold strips. 10)Any reagent should not be exposed to bleaching solvents or the strong gases emitted by bleaching solvents. Any bleaching components will destroy the biological activity of the reagents in the kit. 11)The ELISA reader used for the test needs to be installed with 450±10nmWavelength filters with optical density range of 0-3.5It is recommended to use in advance 15Minutes to preheat. 12)Do not mix or substitute reagents in this kit with reagents from other lots or other sources. 13)The test used EPBoth the tube and the pipette tip are for single use and mixing is strictly prohibited. 14)Do not use expired reagents.

①Problem description: Standard curve gradient is poorPossible causes and corresponding countermeasures: 1.Inaccurate aspiration or addition of liquid——Check the pipette and tips 2.Equilibration time is too short——Ensure adequate balancing time 3.Incomplete washing——Ensure the washing time, washing times and liquid volume per well ②Problem description: The color is very weak or colorlessPossible causes and corresponding countermeasures: 1.Incubation time is too short——Ensure adequate incubation time 2.Incorrect test temperature——Use the recommended test temperature3.Insufficient reagent volume or missed addition/Incorrect dilution——Check the aspiration and addition process to ensure that all reagents are added in order and in sufficient quantities. 4.Enzyme marker inactivation or substrate failure——Mix the enzyme conjugate and substrate and check and judge by rapid color development ③Problem description: Low readingsPossible causes and corresponding countermeasures: 1.The microplate reader is not set up correctly——Check wavelength and filter settings on the microplate reader/Turn on the ELISA reader in advance to preheat ④Problem description: Large coefficient of variationPossible causes and corresponding countermeasures: 1.Incorrect addition of liquid——Check the filling status ⑤Problem description: High background valuePossible causes and corresponding countermeasures: 1.The working concentration of the detection antibody is too high——Use the recommended dilution factor 2.Incomplete washing of ELISA plate——Ensure that each step is cleaned completely; if using an automatic plate washer, check whether all outlets are blocked; whether the washing solution provided in the kit is used 3.The washing liquid is contaminated——Prepare new lotion ⑥Problem description: Low sensitivityPossible causes and corresponding countermeasures: 1.ELISAImproper storage of the test kit——Store relevant reagents according to the instructions 2.Not terminated before reading——ODStop solution should be added to each well before reading.

1.Due to the limitations of existing conditions and scientific and technological levels, it is not possible to conduct comprehensive identification and analysis of all raw materials. This product may have certain quality and technical risks. 2.This kit removed/Some endogenous interfering factors in biological samples have been reduced, but not all possible influencing factors have been removed. 3.The final experimental results are closely related to factors such as the effectiveness of the reagents, the experimenter's related operations, and the experimental environment at the time. Our company is only responsible for the kit itself and is not responsible for the sample consumption caused by the use of the kit. Please fully consider the possible usage of the sample before use and reserve sufficient samples. 4.In order to achieve good experimental results, please only use the reagents provided in our kits, do not mix with products from other manufacturers, and strictly follow the instructions. 5.Abnormal results may occur due to incorrect reagent preparation and incorrect parameter settings of the microplate reader during the operation. Please read the instructions carefully and adjust the instrument before the experiment. 6.Even if the same person performs the same operation, different results may be obtained in two independent experiments. To ensure the reproducibility of the results, it is necessary to control the operation of each step in the experimental process. 7.The test kit will undergo strict quality inspection before shipment. However, due to factors such as transportation conditions, differences in experimental equipment, etc., the user's test results may be inconsistent with the factory data. 8.This test kit has not been compared with similar test kits from other manufacturers or products that detect the same target using different methods, so inconsistent test results cannot be ruled out. 9.The kit is for research use only. If it is used for clinical diagnosis or any other purpose, our company will not be responsible for any problems arising therefrom and will not bear any legal liability.