Description
Human RI (Ribonuclease Inhibitor) ELISA Kit
The Human RI (Ribonuclease Inhibitor) ELISA Kit is specifically designed for the accurate measurement of ribonuclease inhibitor levels in human serum, plasma, and cell culture supernatants. This kit is known for its high sensitivity and specificity, ensuring reliable and reproducible results for a variety of research applications.Ribonuclease inhibitor is a key protein that plays a crucial role in RNA metabolism by binding to and regulating the activity of ribonucleases. Dysregulation of ribonuclease inhibitor has been linked to various diseases such as cancer, autoimmune disorders, and viral infections, highlighting its importance as a potential biomarker for studying these conditions and developing targeted therapies.
With easy-to-follow instructions and quick assay times, the Human RI (Ribonuclease Inhibitor) ELISA Kit is a valuable tool for researchers looking to accurately quantify ribonuclease inhibitor levels in human samples for their studies.
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Human |
| Detection Method: | Colormetric |
| Detection Range: | 1.25-80 ng/mL |
| Sensitivity: | 0.75 ng/mL |
| Sample Volume Required Per Well: | 100µL |
| Sample Type: | Serum, plasma and other biological fluids |
| Specificity: | This kit recognizes Human RI in samples. No significant cross-reactivity or interference between Human RI and analogues was observed. |
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human RI. Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human RI and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human RI, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Human RI. The concentration of Human RI in samples can be calculated by comparing the OD of the samples to the standard curve.
| UniProt Protein Function: | RNH1: Ribonuclease inhibitor which inhibits RNASE1, RNASE2 and ANG. May play a role in redox homeostasis. |
| UniProt Protein Details: | Protein type:Inhibitor Chromosomal Location of Human Ortholog: 11p15. 5 Cellular Component: angiogenin-PRI complex; cytoplasm Molecular Function:protein binding; ribonuclease inhibitor activity Biological Process: negative regulation of catalytic activity; mRNA catabolic process; regulation of angiogenesis |
| NCBI Summary: | Placental ribonuclease inhibitor (PRI) is a member of a family of proteinaceous cytoplasmic RNase inhibitors that occur in many tissues and bind to both intracellular and extracellular RNases (summarized by Lee et al. , 1988 [PubMed 3219362]). In addition to control of intracellular RNases, the inhibitor may have a role in the regulation of angiogenin (MIM 105850). Ribonuclease inhibitor, of 50,000 Da, binds to ribonucleases and holds them in a latent form. Since neutral and alkaline ribonucleases probably play a critical role in the turnover of RNA in eukaryotic cells, RNH may be essential for control of mRNA turnover; the interaction of eukaryotic cells with ribonuclease may be reversible in vivo. [supplied by OMIM, Jul 2010] |
| UniProt Code: | P13489 |
| NCBI GenInfo Identifier: | 132573 |
| NCBI Gene ID: | 6050 |
| NCBI Accession: | P13489. 2 |
| UniProt Secondary Accession: | P13489,Q8IZK8, Q96FD7, Q9BQ80, Q9UDK6, |
| UniProt Related Accession: | P13489 |
| Molecular Weight: | |
| NCBI Full Name: | Ribonuclease inhibitor |
| NCBI Synonym Full Names: | ribonuclease/angiogenin inhibitor 1 |
| NCBI Official Symbol: | RNH1 |
| NCBI Official Synonym Symbols: | RAI; RNH |
| NCBI Protein Information: | ribonuclease inhibitor; placental RNase inhibitor; placental ribonuclease inhibitor |
| UniProt Protein Name: | Ribonuclease inhibitor |
| UniProt Synonym Protein Names: | Placental ribonuclease inhibitor; Placental RNase inhibitor; Ribonuclease/angiogenin inhibitor 1; RAI |
| Protein Family: | Ribonuclease inhibitor |
| UniProt Gene Name: | RNH1 |
| UniProt Entry Name: | RINI_HUMAN |
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e. g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
| Concentration (ng/mL) | O.D | Average | Corrected |
| 80 | 2.346 2.388 | 2.367 | 2.281 |
| 40 | 1.69 1.73 | 1.71 | 1.624 |
| 20 | 0.939 0.909 | 0.924 | 0.838 |
| 10 | 0.481 0.499 | 0.49 | 0.404 |
| 5 | 0.292 0.278 | 0.285 | 0.199 |
| 2.5 | 0.192 0.18 | 0.186 | 0.1 |
| 1.25 | 0.134 0.142 | 0.138 | 0.052 |
| 0 | 0.076 0.096 | 0.086 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human RI were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human RI were tested on 3 different plates, 20 replicates in each plate.
| Intra-assay Precision | Inter-assay Precision | |||||
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean (ng/mL) | 4.36 | 9.52 | 33.52 | 4.08 | 10.44 | 32.52 |
| Standard deviation | 0.29 | 0.55 | 1.58 | 0.28 | 0.62 | 1.05 |
| C V (%) | 6.65 | 5.78 | 4.71 | 6.86 | 5.94 | 3.23 |
Recovery
The recovery of Human RI spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
| Sample Type | Range (%) | Average Recovery (%) |
| Serum (n=5) | 93-108 | 100 |
| EDTA plasma (n=5) | 89-102 | 94 |
| Cell culture media (n=5) | 86-98 | 91 |
Linearity
Samples were spiked with high concentrations of Human RI and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
| Serum (n=5) | EDTA plasma (n=5) | Cell culture media (n=5) | ||
| 1:2 | Range (%) | 92-109 | 94-106 | 97-114 |
| Average (%) | 99 | 101 | 104 | |
| 1:4 | Range (%) | 89-102 | 81-94 | 82-95 |
| Average (%) | 95 | 88 | 88 | |
| 1:8 | Range (%) | 87-102 | 87-100 | 87-101 |
| Average (%) | 93 | 93 | 92 | |
| 1:16 | Range (%) | 89-102 | 84-97 | 87-101 |
| Average (%) | 96 | 89 | 94 |
An unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
| Item | Specifications | Storage |
| Micro ELISA Plate(Dismountable) | 8 wells ×12 strips | -20°C, 6 months |
| Reference Standard | 2 vials | |
| Concentrated Biotinylated Detection Ab (100×) | 1 vial, 120 µL | |
| Concentrated HRP Conjugate (100×) | 1 vial, 120 µL | -20°C(shading light), 6 months |
| Reference Standard & Sample Diluent | 1 vial, 20 mL | 4°C, 6 months |
| Biotinylated Detection Ab Diluent | 1 vial, 14 mL | |
| HRP Conjugate Diluent | 1 vial, 14 mL | |
| Concentrated Wash Buffer (25×) | 1 vial, 30 mL | |
| Substrate Reagent | 1 vial, 10 mL | 4°C(shading light) |
| Stop Solution | 1 vial, 10 mL | 4°C |
| Plate Sealer | 5 pieces | |
| Product Description | 1 copy | |
| Certificate of Analysis | 1 copy |
- Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- Aliquot 100µl of standard solutions into the standard wells.
- Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37°C.
- Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
