Human Immunology ELISA Kits 12
Human FBLN3 (Fibulin 3) ELISA Kit (HUES02671)
- SKU:
- HUES02671
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- Q12805
- Sensitivity:
- 0.19ng/mL
- Range:
- 0.31-20ng/mL
- ELISA Type:
- Sandwich
- Synonyms:
- EFEMP1, DHRD, DRAD, FBNL, FIBL-3, MLVT, MTLV, S1-5
- Reactivity:
- Human
- Sample Type:
- Serum, plasma and other biological fluids
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Human |
| Detection Method: | Colormetric |
| Detection Range: | 0.31-20 ng/mL |
| Sensitivity: | 0.19 ng/mL |
| Sample Volume Required Per Well: | 100µL |
| Sample Type: | Serum, plasma and other biological fluids |
| Specificity: | This kit recognizes Human FBLN3 in samples. No significant cross-reactivity or interference between Human FBLN3 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 FBLN3. 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 FBLN3 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 FBLN3, 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 FBLN3. The concentration of Human FBLN3 in samples can be calculated by comparing the OD of the samples to the standard curve.
| UniProt Protein Function: | EFEMP1: Binds EGFR, the EGF receptor, inducing EGFR autophosphorylation and the activation of downstream signaling pathways. May play a role in cell adhesion and migration. May function as a negative regulator of chondrocyte differentiation. In the olfactory epithelium, it may regulate glial cell migration, differentiation and the ability of glial cells to support neuronal neurite outgrowth. Defects in EFEMP1 are a cause of Doyne honeycomb retinal dystrophy (DHRD); also known as malattia leventinese (MLVT) (ML). DHRD is an autosomal dominant disease characterized by yellow-white deposits known as drusen that accumulate beneath the retinal pigment epithelium. Belongs to the fibulin family. 4 isoforms of the human protein are produced by alternative splicing. |
| UniProt Protein Details: | Protein type:Secreted; Secreted, signal peptide Chromosomal Location of Human Ortholog: 2p16. 1 Cellular Component: extracellular matrix; extracellular region; extracellular space Molecular Function:epidermal growth factor receptor activity; epidermal growth factor receptor binding; protein binding Biological Process: camera-type eye development; embryonic eye morphogenesis; epidermal growth factor receptor signaling pathway; negative regulation of chondrocyte differentiation; peptidyl-tyrosine phosphorylation; post-embryonic eye morphogenesis; regulation of transcription, DNA-dependent; visual perception Disease: Doyne Honeycomb Retinal Dystrophy |
| NCBI Summary: | This gene encodes a member of the fibulin family of extracellular matrix glycoproteins. Like all members of this family, the encoded protein contains tandemly repeated epidermal growth factor-like repeats followed by a C-terminus fibulin-type domain. This gene is upregulated in malignant gliomas and may play a role in the aggressive nature of these tumors. Mutations in this gene are associated with Doyne honeycomb retinal dystrophy. Alternatively spliced transcript variants that encode the same protein have been described. [provided by RefSeq, Nov 2009] |
| UniProt Code: | Q12805 |
| NCBI GenInfo Identifier: | 9973182 |
| NCBI Gene ID: | 2202 |
| NCBI Accession: | Q12805. 2 |
| UniProt Secondary Accession: | Q12805,Q541U7, A8K3I4, B4DW75, D6W5D2, |
| UniProt Related Accession: | Q12805 |
| Molecular Weight: | 39,195 Da |
| NCBI Full Name: | EGF-containing fibulin-like extracellular matrix protein 1 |
| NCBI Synonym Full Names: | EGF containing fibulin like extracellular matrix protein 1 |
| NCBI Official Symbol: | EFEMP1 |
| NCBI Official Synonym Symbols: | DHRD; DRAD; FBNL; MLVT; MTLV; S1-5; FBLN3; FIBL-3 |
| NCBI Protein Information: | EGF-containing fibulin-like extracellular matrix protein 1 |
| UniProt Protein Name: | EGF-containing fibulin-like extracellular matrix protein 1 |
| UniProt Synonym Protein Names: | Extracellular protein S1-5; Fibrillin-like protein; Fibulin-3; FIBL-3 |
| UniProt Gene Name: | EFEMP1 |
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 |
| 20 | 2.265 2.319 | 2.292 | 2.221 |
| 10 | 1.537 1.541 | 1.539 | 1.468 |
| 5 | 0.861 0.825 | 0.843 | 0.772 |
| 2.5 | 0.437 0.441 | 0.439 | 0.368 |
| 1.25 | 0.253 0.247 | 0.25 | 0.179 |
| 0.63 | 0.173 0.155 | 0.164 | 0.093 |
| 0.31 | 0.11 0.128 | 0.119 | 0.048 |
| 0 | 0.069 0.073 | 0.071 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human FBLN3 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human FBLN3 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) | 1.04 | 2.62 | 7.12 | 1.10 | 2.61 | 7.22 |
| Standard deviation | 0.07 | 0.13 | 0.23 | 0.06 | 0.12 | 0.34 |
| C V (%) | 6.73 | 4.96 | 3.23 | 5.45 | 4.60 | 4.71 |
Recovery
The recovery of Human FBLN3 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) | 94-105 | 100 |
| EDTA plasma (n=5) | 95-107 | 101 |
| Cell culture media (n=5) | 95-109 | 100 |
Linearity
Samples were spiked with high concentrations of Human FBLN3 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 (%) | 92-106 | 93-107 | 92-106 |
| Average (%) | 100 | 101 | 98 | |
| 1:4 | Range (%) | 94-107 | 85-94 | 86-99 |
| Average (%) | 100 | 89 | 93 | |
| 1:8 | Range (%) | 94-106 | 83-95 | 84-98 |
| Average (%) | 100 | 89 | 91 | |
| 1:16 | Range (%) | 91-106 | 83-96 | 88-101 |
| Average (%) | 97 | 87 | 93 |
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.
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