Human PLCgamma2 (Phospholipase C Gamma 2, Phosphatidylinositol Specific) CLIA Kit (HUES00573)
Human PLCgamma2 (Phospholipase C Gamma 2, Phosphatidylinositol Specific) CLIA Kit
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 PLCgamma2 . 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 PLCgamma2 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 PLCgamma2, 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 PLCgamma2. The concentration of Human PLCgamma2 in the samples can be calculated by comparing the RLU of the samples to the standard curve.
|Detection range||62.50-4000 pg/mL|
|Sample type||Serum, plasma and other biological fluids|
|Repeatability||CV < 15%|
This kit recognizes Human PLCgamma2 in samples. No significant cross-reactivity or interference between Human PLCgamma2 and analogues was observed.
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.
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human PLCgamma2 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human PLCgamma2 were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||10.64||11.77||6.04||8.63||9.64||10.66|
The recovery of Human PLCgamma2 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
|Sample Type||Range (%)||Average Recovery (%)|
|EDTA plasma (n=5)||85-99||90|
|Cell culture media (n=5)||85-97||90|
Samples were spiked with high concentrations of Human PLCgamma2 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)|
Kit Components & Storage
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.
|Micro CLIA Plate(Dismountable)||8 wells X12 strips||-20'C, 6 months|
|Reference Standard||2 vials|
|Concentrated Biotinylated Detection Ab (100X)||1 vial, 120 uL|
|Concentrated HRP Conjugate (100X)||1 vial, 120 uL||-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 (25X)||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|
Human PLCgamma2 (Phospholipase C Gamma 2, Phosphatidylinositol Specific) CLIA Kit (HUES00573) Assay procedure
- 1. 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.
- 2. Aliquot 100µl of standard solutions into the standard wells.
- 3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- 4. Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids.) into test sample wells.
- 5. Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- 6. 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.
- 7. 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.
- 8. Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- 9. Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- 10. 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.
- 11. Determine the RLU value of each well immediately.
Human PLCgamma2 (Phospholipase C Gamma 2, Phosphatidylinositol Specific) CLIA Kit (HUES00573) Protein Information
|UniProt Protein Function:||PLCG2: a calcium dependent phosphatidylinositol-specific phospholipase C. The activated enzyme produces the second messenger molecules diacylglycerol and inositol 1,4,5-trisphosphate. It is a crucial enzyme in transmembrane signaling. Phosphorylated and activated by tyrosine kinases in response to signaling through a variety of growth factor receptors and immune system receptors.|
|UniProt Protein Details:|
Protein type:Carbohydrate Metabolism - inositol phosphate; Phospholipase; EC 220.127.116.11; Motility/polarity/chemotaxis
Chromosomal Location of Human Ortholog: 16q24.1
Cellular Component: plasma membrane; cytosol
Molecular Function:signal transducer activity; protein binding; phospholipase C activity; phosphoinositide phospholipase C activity
Biological Process: platelet activation; Wnt receptor signaling pathway; inositol phosphate metabolic process; phospholipid catabolic process; calcium-mediated signaling; response to lipopolysaccharide; T cell receptor signaling pathway; activation of store-operated calcium channel activity; B cell receptor signaling pathway; regulation of gene expression; inositol trisphosphate biosynthetic process; B cell differentiation; release of sequestered calcium ion into cytosol; negative regulation of programmed cell death; phosphatidylinositol biosynthetic process; innate immune response; follicular B cell differentiation; blood coagulation
Disease: Autoinflammation, Antibody Deficiency, And Immune Dysregulation, Plcg2-associated; Familial Cold Autoinflammatory Syndrome 3
|NCBI Summary:||The protein encoded by this gene is a transmembrane signaling enzyme that catalyzes the conversion of 1-phosphatidyl-1D-myo-inositol 4,5-bisphosphate to 1D-myo-inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG) using calcium as a cofactor. IP3 and DAG are second messenger molecules important for transmitting signals from growth factor receptors and immune system receptors across the cell membrane. Mutations in this gene have been found in autoinflammation, antibody deficiency, and immune dysregulation syndrome and familial cold autoinflammatory syndrome 3. [provided by RefSeq, Mar 2014]|
|NCBI GenInfo Identifier:||215274231|
|NCBI Gene ID:||5336|
|UniProt Secondary Accession:||P16885,Q3ZTS2, Q59H45, Q969T5, D3DUL3,|
|UniProt Related Accession:||P16885|
|NCBI Full Name:||1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma-2|
|NCBI Synonym Full Names:||phospholipase C, gamma 2 (phosphatidylinositol-specific)|
|NCBI Official Symbol:||PLCG2|
|NCBI Official Synonym Symbols:||FCAS3; APLAID; PLC-IV; PLC-gamma-2|
|NCBI Protein Information:||1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma-2; phospholipase C-IV; phosphoinositide phospholipase C-gamma-2|
|UniProt Protein Name:||1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma-2|
|UniProt Synonym Protein Names:||Phosphoinositide phospholipase C-gamma-2; Phospholipase C-IV; PLC-IV; Phospholipase C-gamma-2; PLC-gamma-2|
|Protein Family:||1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase|
|UniProt Gene Name:||PLCG2|
|UniProt Entry Name:||PLCG2_HUMAN|