Human Cell Biology ELISA Kits 4
Human Slit2 (Slit Homolog 2) CLIA Kit (HUES00554)
- SKU:
- HUES00554
- Product Type:
- ELISA Kit
- ELISA Type:
- CLIA Kit
- Size:
- 96 Assays
- Sensitivity:
- 18.75pg/mL
- Range:
- 31.25-2000pg/mL
- ELISA Type:
- Sandwich
- 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 Slit2 in samples. No significant cross-reactivity or interference between Human Slit2 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 Slit2. 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 Slit2 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 Slit2, 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 Slit2. The concentration of Human Slit2 in the samples can be calculated by comparing the RLU of the samples to the standard curve.
| UniProt Protein Function: | SLIT2: Thought to act as molecular guidance cue in cellular migration, and function appears to be mediated by interaction with roundabout homolog receptors. During neural development involved in axonal navigation at the ventral midline of the neural tube and projection of axons to different regions. SLIT1 and SLIT2 seem to be essential for midline guidance in the forebrain by acting as repulsive signal preventing inappropriate midline crossing by axons projecting from the olfactory bulb. In spinal chord development may play a role in guiding commissural axons once they reached the floor plate by modulating the response to netrin. In vitro, silences the attractive effect of NTN1 but not its growth- stimulatory effect and silencing requires the formation of a ROBO1-DCC complex. May be implicated in spinal chord midline post- crossing axon repulsion. In vitro, only commissural axons that crossed the midline responded to SLIT2. In the developing visual system appears to function as repellent for retinal ganglion axons by providing a repulsion that directs these axons along their appropriate paths prior to, and after passage through, the optic chiasm. In vitro, collapses and repels retinal ganglion cell growth cones. Seems to play a role in branching and arborization of CNS sensory axons, and in neuronal cell migration. In vitro, Slit homolog 2 protein N-product, but not Slit homolog 2 protein C-product, repels olfactory bulb (OB) but not dorsal root ganglia (DRG) axons, induces OB growth cones collapse and induces branching of DRG axons. Seems to be involved in regulating leukocyte migration. 3 isoforms of the human protein are produced by alternative splicing. |
| UniProt Protein Details: | Protein type:Secreted; Secreted, signal peptide; Motility/polarity/chemotaxis; Extracellular matrix Chromosomal Location of Human Ortholog: 4p15. 2 Cellular Component: extracellular space; cell surface; membrane; cytoplasm; plasma membrane; extracellular region Molecular Function:heparin binding; laminin-1 binding; identical protein binding; proteoglycan binding; protein binding; Roundabout binding; protein homodimerization activity; chemorepellent activity; calcium ion binding; GTPase inhibitor activity Biological Process: negative regulation of small GTPase mediated signal transduction; axon guidance; positive regulation of apoptosis; negative chemotaxis; motor axon guidance; negative regulation of axon extension; negative regulation of smooth muscle cell migration; induction of negative chemotaxis; corticospinal neuron axon guidance through the spinal cord; negative regulation of cell proliferation; cell-cell adhesion; response to cortisol stimulus; negative regulation of protein amino acid phosphorylation; ureteric bud development; negative regulation of leukocyte chemotaxis; negative regulation of cell migration; negative regulation of actin filament polymerization; in utero embryonic development; chemorepulsion involved in postnatal olfactory bulb interneuron migration; dorsal/ventral axon guidance; cellular response to hormone stimulus; axon extension involved in axon guidance; chemorepulsion involved in embryonic olfactory bulb interneuron migration; branching morphogenesis of a tube; cell migration during sprouting angiogenesis; negative regulation of catalytic activity; negative regulation of vascular permeability; positive regulation of axonogenesis; negative regulation of cell growth; metanephros development; retinal ganglion cell axon guidance |
| NCBI Summary: | This gene encodes a member of the slit family of secreted glycoproteins, which are ligands for the Robo family of immunoglobulin receptors. Slit proteins play highly conserved roles in axon guidance and neuronal migration and may also have functions during other cell migration processes including leukocyte migration. Members of the slit family are characterized by an N-terminal signal peptide, four leucine-rich repeats, nine epidermal growth factor repeats, and a C-terminal cysteine knot. Proteolytic processing of this protein gives rise to an N-terminal fragment that contains the four leucine-rich repeats and five epidermal growth factor repeats and a C-terminal fragment that contains four epidermal growth factor repeats and the cysteine knot. Both full length and cleaved proteins are secreted extracellularly and can function in axon repulsion as well as other specific processes. Alternative splicing results in multiple transcript variants. [provided by RefSeq, Sep 2015] |
| UniProt Code: | O94813 |
| NCBI GenInfo Identifier: | 33112440 |
| NCBI Gene ID: | 9353 |
| NCBI Accession: | O94813. 1 |
| UniProt Secondary Accession: | O94813,O95710, Q17RU3, Q9Y5Q7, B7ZLR5, |
| UniProt Related Accession: | O94813 |
| Molecular Weight: | 168,893 Da |
| NCBI Full Name: | Slit homolog 2 protein |
| NCBI Synonym Full Names: | slit homolog 2 (Drosophila) |
| NCBI Official Symbol: | SLIT2 |
| NCBI Official Synonym Symbols: | SLIL3; Slit-2 |
| NCBI Protein Information: | slit homolog 2 protein |
| UniProt Protein Name: | Slit homolog 2 protein |
| Protein Family: | Slit homolog 2 protein |
| UniProt Gene Name: | SLIT2 |
| UniProt Entry Name: | SLIT2_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 | 54388 57054 | 55721 | 55693 |
| 1000 | 22791 25157 | 23974 | 23946 |
| 500 | 11857 10207 | 11032 | 11004 |
| 250 | 4819 5769 | 5294 | 5266 |
| 125 | 2788 2428 | 2608 | 2580 |
| 62.5 | 1369 1253 | 1311 | 1283 |
| 31.25 | 640 708 | 674 | 646 |
| 0 | 28 28 | 28 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human Slit2 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human Slit2 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) | 99.07 | 293.09 | 997.64 | 106.84 | 267.01 | 986.60 |
| Standard deviation | 11.86 | 31.24 | 98.27 | 10.73 | 23.28 | 97.67 |
| C V (%) | 11.97 | 10.66 | 9.85 | 10.04 | 8.72 | 9.90 |
Recovery
The recovery of Human Slit2 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) | 86-98 | 91 |
| EDTA plasma (n=5) | 99-115 | 107 |
| Cell culture media (n=5) | 96-113 | 104 |
Linearity
Samples were spiked with high concentrations of Human Slit2 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 (%) | 97-115 | 97-115 | 98-112 |
| Average (%) | 105 | 105 | 104 | |
| 1:4 | Range (%) | 88-102 | 99-115 | 92-105 |
| Average (%) | 95 | 106 | 99 | |
| 1:8 | Range (%) | 89-105 | 101-115 | 94-108 |
| Average (%) | 97 | 106 | 102 | |
| 1:16 | Range (%) | 100-113 | 98-113 | 93-104 |
| Average (%) | 105 | 105 | 99 |
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.
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