Human Cell Biology ELISA Kits 3
Human INSL3 (Insulin Like Protein 3) CLIA Kit (HUES00290)
- SKU:
- HUES00290
- Product Type:
- ELISA Kit
- ELISA Type:
- CLIA Kit
- Size:
- 96 Assays
- Sensitivity:
- 18.75pg/mL
- Range:
- 31.25-2000pg/mL
- ELISA Type:
- Sandwich
- Synonyms:
- RLF, RLNL, ley-I-L
- Reactivity:
- Human
- Sample Type:
- Serum, plasma and other biological fluids
- Research Area:
- Cell Biology
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Human |
| Detection method: | Chemiluminescence |
| Detection range: | 31.25-2000 pg/mL |
| Sensitivity: | 18.75 pg/mL |
| Sample volume: | 100µL |
| Sample type: | Serum, plasma and other biological fluids |
| Repeatability: | CV < 15% |
| Specificity: | This kit recognizes Human INSL3 in samples. No significant cross-reactivity or interference between Human INSL3 and analogues was observed. |
This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Human INSL3. Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human INSL3 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 INSL3, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Human INSL3. The concentration of Human INSL3 in the samples can be calculated by comparing the RLU of the samples to the standard curve.
| UniProt Protein Function: | INSL3: Seems to play a role in testicular function. May be a trophic hormone with a role in testicular descent in fetal life. Is a ligand for LGR8 receptor. Defects in INSL3 seems to be a cause of cryptorchidism (CRYPTO); also known as impaired testicular descent. It is one of the most frequent congenital abnormalities in humans, involving 2-5% of male births. Cryptorchidism is associated with increased risk of infertility and testicular cancer. The frequency of INSL3 gene mutations as a cause of cryptorchidism is low. Belongs to the insulin family. |
| UniProt Protein Details: | Protein type:Secreted; Secreted, signal peptide Chromosomal Location of Human Ortholog: 19p13. 2-p12 Cellular Component: extracellular space; perinuclear region of cytoplasm; extracellular region Molecular Function:G-protein-coupled receptor binding; protease binding; hormone activity; insulin receptor binding; receptor binding Biological Process: negative regulation of cell proliferation; cell-cell signaling; in utero embryonic development; positive regulation of cAMP biosynthetic process; positive regulation of cell proliferation; male gonad development; spermatogenesis; G-protein signaling, adenylate cyclase inhibiting pathway; oocyte maturation; negative regulation of apoptosis Disease: Cryptorchidism, Unilateral Or Bilateral |
| NCBI Summary: | This gene encodes a member of the insulin-like hormone superfamily. The encoded protein is mainly produced in gonadal tissues. Studies of the mouse counterpart suggest that this gene may be involved in the development of urogenital tract and female fertility. This protein may also act as a hormone to regulate growth and differentiation of gubernaculum, and thus mediating intra-abdominal testicular descent. Mutations in this gene may lead to cryptorchidism. Alternate splicing results in multiple transcript variants. [provided by RefSeq, May 2012] |
| UniProt Code: | P51460 |
| NCBI GenInfo Identifier: | 317373369 |
| NCBI Gene ID: | 3640 |
| NCBI Accession: | P51460. 2 |
| UniProt Secondary Accession: | P51460,Q3KPI5, Q3KPI6, Q6YNB5, Q9UEA2, Q9UPH6, B4DZ72 G3XAG0, |
| UniProt Related Accession: | P51460 |
| Molecular Weight: | 16,872 Da |
| NCBI Full Name: | Insulin-like 3 |
| NCBI Synonym Full Names: | insulin-like 3 (Leydig cell) |
| NCBI Official Symbol: | INSL3 |
| NCBI Official Synonym Symbols: | RLF; RLNL; ley-I-L |
| NCBI Protein Information: | insulin-like 3; prepro-INSL3; relaxin-like factor b; leydig insulin-like peptide; leydig insulin -like hormone |
| UniProt Protein Name: | Insulin-like 3 |
| UniProt Synonym Protein Names: | Leydig insulin-like peptide; Ley-I-L; Relaxin-like factorCleaved into the following 2 chains:Insulin-like 3 B chain; Insulin-like 3 A chain |
| Protein Family: | Insulin |
| UniProt Gene Name: | INSL3 |
| UniProt Entry Name: | INSL3_HUMAN |
As the RLU 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 (pg/mL) | RLU | Average | Corrected |
| 2000 | 54160 57282 | 55721 | 55693 |
| 1000 | 22442 25506 | 23974 | 23946 |
| 500 | 11515 10549 | 11032 | 11004 |
| 250 | 5089 5499 | 5294 | 5266 |
| 125 | 2761 2455 | 2608 | 2580 |
| 62.5 | 1314 1308 | 1311 | 1283 |
| 31.25 | 665 683 | 674 | 646 |
| 0 | 27 29 | 28 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human INSL3 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human INSL3 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 (pg/mL) | 105.88 | 281.99 | 851.44 | 111.83 | 295.88 | 861.62 |
| Standard deviation | 13.02 | 22.73 | 65.48 | 12.20 | 25.80 | 96.16 |
| C V (%) | 12.30 | 8.06 | 7.69 | 10.91 | 8.72 | 11.16 |
Recovery
The recovery of Human INSL3 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) | 96-114 | 104 |
| EDTA plasma (n=5) | 92-107 | 100 |
| Cell culture media (n=5) | 93-106 | 99 |
Linearity
Samples were spiked with high concentrations of Human INSL3 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 (%) | 103-119 | 90-102 | 91-106 |
| Average (%) | 110 | 96 | 98 | |
| 1:4 | Range (%) | 98-117 | 101-116 | 97-113 |
| Average (%) | 107 | 109 | 105 | |
| 1:8 | Range (%) | 88-98 | 94-106 | 99-116 |
| Average (%) | 93 | 100 | 106 | |
| 1:16 | Range (%) | 87-102 | 101-114 | 86-99 |
| Average (%) | 94 | 106 | 91 |
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 CLIA 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 A | 1 vial, 5 mL | 4°C (shading light) |
| Substrate Reagent B | 1 vial, 5 mL | 4°C (shading light) |
| 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 100µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37°C. Protect the plate from light.
- Determine the RLU value of each well immediately.
Related Products
Human INSL3 / Insulin-like 3 ELISA Kit
